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Sample records for haemolytica cytotoxin purification

  1. Purification of the receptor for the N-acetyl-D-glucosamine specific adhesin of Mannheimia haemolytica from bovine neutrophils.

    Science.gov (United States)

    De la Mora, Alfonso; Suárez-Güemes, Francisco; Trigo, Francisco; Gorocica, Patricia; Solórzano, Carlos; Slomianny, Marie-Christine; Agundis, Concepción; Pereyra, M Ali; Zenteno, Edgar

    2007-10-01

    The GlcNAc-specific adhesin from Mannheimia haemolytica (MhA) has been shown to participate in pathogenicity of mannheimiosis due to its capacity to adhere to tracheal epithelial cells and activate the oxidative burst of bovine neutrophils. In this work, we purified the MhA receptor from bovine neutrophils (MhAr) by affinity chromatography on MhA-Sepharose. The MhAr, which corresponded to approximately 2% of the protein from cell lysate, is a glycoprotein mainly composed of Glu, Ala, Ser, Gly, and Asp, without cysteine. The glycan portion, which corresponds to 20% by weight, is composed of GalNAc, GlcNAc, Man, Gal, and NeuAc. The receptor is a 165-kDa glycoprotein, as determined by molecular sieve chromatography under native conditions; SDS-PAGE analysis shows a heterodimer of 83 and 80 kDa subunits. This work suggests that the GlcNAc-containing receptor plays a relevant role by activating bovine neutrophils through non-opsonic mechanisms.

  2. Biological activity of Serratia marcescens cytotoxin

    Directory of Open Access Journals (Sweden)

    G.V. Carbonell

    2003-03-01

    Full Text Available Serratia marcescens cytotoxin was purified to homogeneity by ion-exchange chromatography on a DEAE Sepharose Fast Flow column, followed by gel filtration chromatography on a Sephadex G100 column. The molecular mass of the cytotoxin was estimated to be about 50 kDa. Some biological properties of the cytotoxin were analyzed and compared with well-characterized toxins, such as VT1, VT2 and CNF from Escherichia coli and hemolysin produced by S. marcescens. The sensitivity of the cell lines CHO, HeLa, HEp-2, Vero, BHK-21, MA 104 and J774 to the cytotoxin was determined by the cell viability assay using neutral red. CHO and HEp-2 were highly sensitive, with massive cellular death after 1 h of treatment, followed by BHK-21, HeLa, Vero and J774 cells, while MA 104 was insensitive to the toxin. Cytotoxin induced morphological changes such as cell rounding with cytoplasmic retraction and nuclear compactation which were evident 15 min after the addition of cytotoxin. The cytotoxic assays show that 15 min of treatment with the cytotoxin induced irreversible intoxication of the cells, determined by loss of cell viability. Concentrations of 2 CD50 (0.56 µg/ml of purified cytotoxin did not present any hemolytic activity, showing that the cytotoxin is distinct from S. marcescens hemolysin. Antisera prepared against S. marcescens cytotoxin did not neutralize the cytotoxic activity of VT1, VT2 or CNF toxin, indicating that these toxins do not share antigenic determinants with cytotoxin. Moreover, we did not detect gene sequences for any of these toxins in S. marcescens by PCR assay. These results suggest that S. marcescens cytotoxin is not related to any of these toxins from E. coli.

  3. Purification, crystallization and preliminary X-ray structural studies of a 7.2 kDa cytotoxin isolated from the venom of Daboia russelli russelli of the Viperidae family

    Science.gov (United States)

    Roy Choudhury, Subhasree; Gomes, Aparna; Gomes, Antony; Dattagupta, Jiban K.; Sen, Udayaditya

    2006-01-01

    A cytotoxin (MW 7.2 kDa) from Indian Russell’s viper (Daboia russelli russelli) venom possessing antiproliferative activity, cardiotoxicity, neurotoxicity and myotoxicity has been purified, characterized and crystallized. The crystals belong to the tetragonal space group P41, with unit-cell parameters a = b = 47.94, c = 50.2 Å. Larger crystals, which diffracted to 1.5 Å, were found to be twinned; diffraction data were therefore collected to 2.93 Å resolution using a smaller crystal. Molecular-replacement calculations identified two molecules of the protein in the asymmetric unit, which is in accordance with the calculated V M value. PMID:16511326

  4. Effect of intranasal recombinant Mannheimia haemolytica ...

    African Journals Online (AJOL)

    ADEYEYE

    2015-03-20

    Mar 20, 2015 ... *Correspondence: Tel.: +2348066486080, E-mail: banabis2001@yahoo.com. Abstract. There is dearth of information on the haematological changes associated with Mannheimia haemolytica vaccination in goats, hence this report which describes some haematological changes observed following.

  5. Purification

    DEFF Research Database (Denmark)

    Andersen, Astrid Oberborbeck

    2017-01-01

    of categories can be understood as practices of purification. However, a purely technical grip on water is never possible. Unruly elements, like weather, contamination, urban dwellers, and competing interests, interfere and make processes of intervention unstable. Water is never completely cleaned, and, equally......In Arequipa, Peru’s second largest city, engineers work hard to control water flows and provide different sectors with clean and sufficient water. In 2011, only 10 percent of the totality of water used daily by Arequipa’s then close to 1 million people—in households, tourism, industry, and mining......—was treated before it was returned to the river where it continues its flow downstream towards cultivated fields and, finally, into the Pacific Ocean. It takes specialized knowledge and manifold technologies to manage water and sustain life in Arequipa, and engineers are central actors for making water flow...

  6. Immunogenicity of Pasteurella multocida and Mannheimia haemolytica outer membrane vesicles

    Science.gov (United States)

    Roier, Sandro; Fenninger, Judith C.; Leitner, Deborah R.; Rechberger, Gerald N.; Reidl, Joachim; Schild, Stefan

    2013-01-01

    Pasteurella multocida is able to cause disease in humans and in a wide range of animal hosts, including fowl cholera in birds, atrophic rhinitis in pigs, and snuffles in rabbits. Together with Mannheimia haemolytica, P. multocida also represents a major bacterial causative agent of bovine respiratory disease (BRD), which is one of the most important causes for economic losses for the cattle backgrounding and feedlot industry. Commercially available vaccines only partially prevent infections caused by P. multocida and M. haemolytica. Thus, this study characterized the immunogenicity of P. multocida and M. haemolytica outer membrane vesicles (OMVs) upon intranasal immunization of BALB/c mice. Enzyme-linked immunosorbent assays (ELISA) revealed that OMVs derived from P. multocida or M. haemolytica are able to induce robust humoral and mucosal immune responses against the respective donor strain. In addition, also significant cross-immunogenic potential was observed for both OMV types. Colonization studies showed that a potential protective immune response against P. multocida is not only achieved by immunization with P. multocida OMVs, but also by immunization with OMVs derived from M. haemolytica. Immunoblot and immunoprecipitation analyses demonstrated that M. haemolytica OMVs induce a more complex immune response compared to P. multocida OMVs. The outer membrane proteins OmpA, OmpH, and P6 were identified as the three major immunogenic proteins of P. multocida OMVs. Amongst others, the serotype 1-specific antigen, an uncharacterized outer membrane protein, as well as the outer membrane proteins P2 and OmpA were found to be the most important antigens of M. haemolytica OMVs. These findings are useful for the future development of broad-spectrum OMV based vaccines against BRD and other infections caused by P. multocida or M. haemolytica. PMID:23731905

  7. The cytotoxin of Pseudomonas aeruginosa : Cytotoxicity requires proteolytic activation

    NARCIS (Netherlands)

    Orlik-Eisel, Gabriele; Lutz, Frieder; Henschen, Agnes; Eisel, Ulrich; Struckmeier, Martin; Kräuter, Josef; Niemann, Heiner

    The primary structure of a cytotoxin from Pseudomonas aeruginosa was determined by sequencing of the structural gene. The cytotoxin (31,700 Mr) lacks an N-terminal signal sequence for bacterial secretion but contains a pentapeptide consensus sequence commonly found in prokaryotic proteins which

  8. Cobra venom cytotoxins; apoptotic or necrotic agents?

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    Ebrahim, Karim; Shirazi, Farshad H; Mirakabadi, Abbas Zare; Vatanpour, Hossein

    2015-12-15

    Organs homeostasis is controlled by a dynamic balance between cell proliferation and apoptosis. Failure to induction of apoptosis has been implicated in tumor development. Cytotoxin-I (CTX-I) and cytotoxin-II (CTX-II) are two physiologically active polypeptides found in Caspian cobra venom. Anticancer activity and mechanism of cell death induced by these toxins have been studied. The toxins were purified by different chromatographic steps and their cytotoxicity and pattern of cell death were determined by MTT, LDH release, acridine orange/ethidium bromide (AO/EtBr) double staining, flow cytometric analysis, caspase-3 activity and neutral red assays. The IC50 of CTX-II in MCF-7, HepG2, DU-145 and HL-60 was 4.1 ± 1.3, 21.2 ± 4.4, 9.4 ± 1.8 μg/mL and 16.3 ± 1.9 respectively while the IC50 of this toxin in normal MDCK cell line was 54.5 ± 3.9 μg/mL. LDH release suddenly increase after a specific toxins concentrations in all cell lines. AO/EtBr double staining, flow cytometric analysis and caspase-3 activity assay confirm dose and time-dependent induction of apoptosis by both toxins. CTX-I and CTX-II treated cells lost their lysosomal membrane integrity and couldn't uptake neutral red day. CTX-I and CTX-II showed significant anticancer activity with minimum effects on normal cells and better IC50 compared to current anticancer drug; cisplatin. They induce their apoptotic effect via lysosomal pathways and release of cathepsins to cytosol. These effects were seen in limited rage of toxins concentrations and pattern of cell death rapidly changes to necrosis by increase in toxin's concentration. In conclusion, significant apoptogenic effects of these toxins candidate them as a possible anticancer agent. Copyright © 2015 Elsevier Ltd. All rights reserved.

  9. 9 CFR 113.68 - Pasteurella Haemolytica Vaccine, Bovine.

    Science.gov (United States)

    2010-01-01

    ... established as pure, safe, and immunogenic shall be used for vaccine production. All serials of vaccine shall.... (b) Each lot of Master Seed used for vaccine production shall be tested for immunogenicity. The... 9 Animals and Animal Products 1 2010-01-01 2010-01-01 false Pasteurella Haemolytica Vaccine...

  10. ISOLATION OF Mannheimia haemolytica FROM THE UHT MILK

    OpenAIRE

    YILMAZ, Aysun; DALKILIÇ KAYA, Gözde

    2012-01-01

      ABSTRACT This study describes the isolation and identification of Mannheimia haemolytica from the UHT milk. This is important for public health since human might get infected with this bacteria. Three package of semi-skimmed UHT drinking milk with cacao was obtained from a commercial company and analysed in a private laboratory (Istanbul, Turkey) bacteriologically. The gross apparence of the milk was not normal. It was thickened and clotted. Milk homogenates were analysed by using ...

  11. Proximity-Dependent Inhibition of Growth of Mannheimia haemolytica by Pasteurella multocida

    Science.gov (United States)

    Bavananthasivam, Jegarubee; Dassanayake, Rohana P.; Kugadas, Abirami; Shanthalingam, Sudarvili; Call, Douglas R.; Knowles, Donald P.

    2012-01-01

    Mannheimia haemolytica, Pasteurella multocida, and Bibersteinia trehalosi have been identified in the lungs of pneumonic bighorn sheep (BHS; Ovis canadensis). Of these pathogens, M. haemolytica has been shown to consistently cause fatal pneumonia in BHS under experimental conditions. However, M. haemolytica has been isolated by culture less frequently than the other bacteria. We hypothesized that the growth of M. haemolytica is inhibited by other bacteria in the lungs of BHS. The objective of this study was to determine whether P. multocida inhibits the growth of M. haemolytica. Although in monoculture both bacteria exhibited similar growth characteristics, in coculture with P. multocida there was a clear inhibition of growth of M. haemolytica. The inhibition was detected at mid-log phase and continued through the stationary phase. When cultured in the same medium, the growth of M. haemolytica was inhibited when both bacteria were separated by a membrane that allowed contact (pore size, 8.0 μm) but not when they were separated by a membrane that limited contact (pore size, 0.4 μm). Lytic bacteriophages or bactericidal compounds could not be detected in the culture supernatant fluid from monocultures of P. multocida or from P. multocida-M. haemolytica cocultures. These results indicate that P. multocida inhibits the growth of M. haemolytica by a contact- or proximity-dependent mechanism. If the inhibition of growth of M. haemolytica by P. multocida occurs in vivo as well, it could explain the inconsistent isolation of M. haemolytica from the lungs of pneumonic BHS. PMID:22798357

  12. A multiplex PCR assay for molecular capsular serotyping of Mannheimia haemolytica serotypes 1, 2, and 6

    Science.gov (United States)

    Mannheimia haemolytica is an important respiratory pathogen of ruminants. Of the 12 capsular serovars identified, 1 and 6 are most frequently associated with disease in cattle, while 2 is largely a commensal. Comparative analysis of 24 M. haemolytica genomes was used to identify unique genes associa...

  13. Proximity-dependent inhibition of growth of mannheimia haemolytica by pasteurella multocida.

    Science.gov (United States)

    Mannheimia haemolytica, Pasteurella multocida, and Bibersteinia trehalosi have been identified in the lungs of pneumonic bighorn sheep (BHS; Ovis canadensis). Of these pathogens, M. haemolytica has been shown to consistently cause fatal pneumonia in BHS under experimental conditions. However, M. hae...

  14. Rapid detection of Mannheimia haemolytica in lung tissues of sheep and from bacterial culture.

    Science.gov (United States)

    Kumar, Jyoti; Dixit, Shivendra Kumar; Kumar, Rajiv

    2015-09-01

    This study was aimed to detect Mannheimia haemolytica in lung tissues of sheep and from a bacterial culture. M. haemolytica is one of the most important and well-established etiological agents of pneumonia in sheep and other ruminants throughout the world. Accurate diagnosis of M. haemolytica primarily relies on bacteriological examination, biochemical characteristics and, biotyping and serotyping of the isolates. In an effort to facilitate rapid M. haemolytica detection, polymerase chain reaction assay targeting Pasteurella haemolytica serotype-1 specific antigens (PHSSA), Rpt2 and 12S ribosomal RNA (rRNA) genes were used to detect M. haemolytica directly from lung tissues and from bacterial culture. A total of 12 archived lung tissues from sheep that died of pneumonia on an organized farm were used. A multiplex polymerase chain reaction (mPCR) based on two-amplicons targeted PHSSA and Rpt2 genes of M. haemolytica were used for identification of M. haemolytica isolates in culture from the lung samples. All the 12 lung tissue samples were tested for the presence M. haemolytica by PHSSA and Rpt2 genes based PCR and its confirmation by sequencing of the amplicons. All the 12 lung tissue samples tested for the presence of PHSSA and Rpt2 genes of M. haemolytica by mPCR were found to be positive. Amplification of 12S rRNA gene fragment as internal amplification control was obtained with each mPCR reaction performed from DNA extracted directly from lung tissue samples. All the M. haemolytica were also positive for mPCR. No amplified DNA bands were observed for negative control reactions. All the three nucleotide sequences were deposited in NCBI GenBank (Accession No. KJ534629, KJ534630 and KJ534631). Sequencing of the amplified products revealed the identity of 99-100%, with published sequence of PHSSA and Rpt2 genes of M. haemolytica available in the NCBI database. Sheep specific mitochondrial 12S rRNA gene sequence also revealed the identity of 98% with published

  15. Rapid detection of Mannheimia haemolytica in lung tissues of sheep and from bacterial culture

    Directory of Open Access Journals (Sweden)

    Jyoti Kumar

    2015-09-01

    Full Text Available Aim: This study was aimed to detect Mannheimia haemolytica in lung tissues of sheep and from a bacterial culture. Introduction: M. haemolytica is one of the most important and well-established etiological agents of pneumonia in sheep and other ruminants throughout the world. Accurate diagnosis of M. haemolytica primarily relies on bacteriological examination, biochemical characteristics and, biotyping and serotyping of the isolates. In an effort to facilitate rapid M. haemolytica detection, polymerase chain reaction assay targeting Pasteurella haemolytica serotype-1 specific antigens (PHSSA, Rpt2 and 12S ribosomal RNA (rRNA genes were used to detect M. haemolytica directly from lung tissues and from bacterial culture. Materials and Methods: A total of 12 archived lung tissues from sheep that died of pneumonia on an organized farm were used. A multiplex polymerase chain reaction (mPCR based on two-amplicons targeted PHSSA and Rpt2 genes of M. haemolytica were used for identification of M. haemolytica isolates in culture from the lung samples. All the 12 lung tissue samples were tested for the presence M. haemolytica by PHSSA and Rpt2 genes based PCR and its confirmation by sequencing of the amplicons. Results: All the 12 lung tissue samples tested for the presence of PHSSA and Rpt2 genes of M. haemolytica by mPCR were found to be positive. Amplification of 12S rRNA gene fragment as internal amplification control was obtained with each mPCR reaction performed from DNA extracted directly from lung tissue samples. All the M. haemolytica were also positive for mPCR. No amplified DNA bands were observed for negative control reactions. All the three nucleotide sequences were deposited in NCBI GenBank (Accession No. KJ534629, KJ534630 and KJ534631. Sequencing of the amplified products revealed the identity of 99-100%, with published sequence of PHSSA and Rpt2 genes of M. haemolytica available in the NCBI database. Sheep specific mitochondrial 12S r

  16. Naja naja oxiana Cobra Venom Cytotoxins CTI and CTII Disrupt Mitochondrial Membrane Integrity: Implications for Basic Three-Fingered Cytotoxins.

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    Sardar E Gasanov

    Full Text Available Cobra venom cytotoxins are basic three-fingered, amphipathic, non-enzymatic proteins that constitute a major fraction of cobra venom. While cytotoxins cause mitochondrial dysfunction in different cell types, the mechanisms by which cytotoxins bind to mitochondria remain unknown. We analyzed the abilities of CTI and CTII, S-type and P-type cytotoxins from Naja naja oxiana respectively, to associate with isolated mitochondrial fractions or with model membranes that simulate the mitochondrial lipid environment by using a myriad of biophysical techniques. Phosphorus-31 nuclear magnetic resonance (31P-NMR spectroscopy data suggest that both cytotoxins bind to isolated mitochondrial fractions and promote the formation of aberrant non-bilayer structures. We then hypothesized that CTI and CTII bind to cardiolipin (CL to disrupt mitochondrial membranes. Collectively, 31P-NMR, electron paramagnetic resonance (EPR, proton NMR (1H-NMR, deuterium NMR (2H-NMR spectroscopy, differential scanning calorimetry, and erythrosine phosphorescence assays suggest that CTI and CTII bind to CL to generate non-bilayer structures and promote the permeabilization, dehydration and fusion of large unilamellar phosphatidylcholine (PC liposomes enriched with CL. On the other hand, CTII but not CTI caused biophysical alterations of large unilamellar PC liposomes enriched with phosphatidylserine (PS. Mechanistically, single molecule docking simulations identified putative CL, PS and PC binding sites in CTI and CTII. While the predicted binding sites for PS and PC share a high number of interactive amino acid residues in CTI and CTII, the CL biding sites in CTII and CTI are more divergent as it contains additional interactive amino acid residues. Overall, our data suggest that cytotoxins physically associate with mitochondrial membranes by binding to CL to disrupt mitochondrial structural integrity.

  17. Naja naja oxiana Cobra Venom Cytotoxins CTI and CTII Disrupt Mitochondrial Membrane Integrity: Implications for Basic Three-Fingered Cytotoxins.

    Science.gov (United States)

    Gasanov, Sardar E; Shrivastava, Indira H; Israilov, Firuz S; Kim, Aleksandr A; Rylova, Kamila A; Zhang, Boris; Dagda, Ruben K

    2015-01-01

    Cobra venom cytotoxins are basic three-fingered, amphipathic, non-enzymatic proteins that constitute a major fraction of cobra venom. While cytotoxins cause mitochondrial dysfunction in different cell types, the mechanisms by which cytotoxins bind to mitochondria remain unknown. We analyzed the abilities of CTI and CTII, S-type and P-type cytotoxins from Naja naja oxiana respectively, to associate with isolated mitochondrial fractions or with model membranes that simulate the mitochondrial lipid environment by using a myriad of biophysical techniques. Phosphorus-31 nuclear magnetic resonance (31P-NMR) spectroscopy data suggest that both cytotoxins bind to isolated mitochondrial fractions and promote the formation of aberrant non-bilayer structures. We then hypothesized that CTI and CTII bind to cardiolipin (CL) to disrupt mitochondrial membranes. Collectively, 31P-NMR, electron paramagnetic resonance (EPR), proton NMR (1H-NMR), deuterium NMR (2H-NMR) spectroscopy, differential scanning calorimetry, and erythrosine phosphorescence assays suggest that CTI and CTII bind to CL to generate non-bilayer structures and promote the permeabilization, dehydration and fusion of large unilamellar phosphatidylcholine (PC) liposomes enriched with CL. On the other hand, CTII but not CTI caused biophysical alterations of large unilamellar PC liposomes enriched with phosphatidylserine (PS). Mechanistically, single molecule docking simulations identified putative CL, PS and PC binding sites in CTI and CTII. While the predicted binding sites for PS and PC share a high number of interactive amino acid residues in CTI and CTII, the CL biding sites in CTII and CTI are more divergent as it contains additional interactive amino acid residues. Overall, our data suggest that cytotoxins physically associate with mitochondrial membranes by binding to CL to disrupt mitochondrial structural integrity.

  18. Naja naja oxiana Cobra Venom Cytotoxins CTI and CTII Disrupt Mitochondrial Membrane Integrity: Implications for Basic Three-Fingered Cytotoxins

    Science.gov (United States)

    Gasanov, Sardar E.; Shrivastava, Indira H.; Israilov, Firuz S.; Kim, Aleksandr A.; Rylova, Kamila A.; Zhang, Boris; Dagda, Ruben K.

    2015-01-01

    Cobra venom cytotoxins are basic three-fingered, amphipathic, non-enzymatic proteins that constitute a major fraction of cobra venom. While cytotoxins cause mitochondrial dysfunction in different cell types, the mechanisms by which cytotoxins bind to mitochondria remain unknown. We analyzed the abilities of CTI and CTII, S-type and P-type cytotoxins from Naja naja oxiana respectively, to associate with isolated mitochondrial fractions or with model membranes that simulate the mitochondrial lipid environment by using a myriad of biophysical techniques. Phosphorus-31 nuclear magnetic resonance (31P-NMR) spectroscopy data suggest that both cytotoxins bind to isolated mitochondrial fractions and promote the formation of aberrant non-bilayer structures. We then hypothesized that CTI and CTII bind to cardiolipin (CL) to disrupt mitochondrial membranes. Collectively, 31P-NMR, electron paramagnetic resonance (EPR), proton NMR (1H-NMR), deuterium NMR (2H-NMR) spectroscopy, differential scanning calorimetry, and erythrosine phosphorescence assays suggest that CTI and CTII bind to CL to generate non-bilayer structures and promote the permeabilization, dehydration and fusion of large unilamellar phosphatidylcholine (PC) liposomes enriched with CL. On the other hand, CTII but not CTI caused biophysical alterations of large unilamellar PC liposomes enriched with phosphatidylserine (PS). Mechanistically, single molecule docking simulations identified putative CL, PS and PC binding sites in CTI and CTII. While the predicted binding sites for PS and PC share a high number of interactive amino acid residues in CTI and CTII, the CL biding sites in CTII and CTI are more divergent as it contains additional interactive amino acid residues. Overall, our data suggest that cytotoxins physically associate with mitochondrial membranes by binding to CL to disrupt mitochondrial structural integrity. PMID:26091109

  19. Impact of Cytotoxin-Associated Gene Product-A Positive ...

    African Journals Online (AJOL)

    Introduction: Available data on the possible association between Helicobacter Pylori (H. pylori) infection and diabetes mellitus (DM) are contradictory. The prevalence of cytotoxin associated gene product A (cagA) positive H. pylori is high in Egypt. This study aims to examine its association with type 2 DM, and its effect on ...

  20. Two outer membrane proteins are bovine lactoferrin-binding proteins in Mannheimia haemolytica A1.

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    Samaniego-Barrón, Luisa; Luna-Castro, Sarahí; Piña-Vázquez, Carolina; Suárez-Güemes, Francisco; de la Garza, Mireya

    2016-09-06

    Mannheimia haemolytica is a Gram negative bacterium that is part of the bovine respiratory disease, which causes important economic losses in the livestock industry. In the present work, the interaction between M. haemolytica A1 and bovine lactoferrin (BLf) was studied. This iron-chelating glycoprotein is part of the mammalian innate-immune system and is present in milk and mucosal secretions; Lf is also contained in neutrophils secondary granules, which release this glycoprotein at infection sites. It was evidenced that M. haemolytica was not able to use iron-charged BLf (BholoLf) as a sole iron source; nevertheless, iron-lacked BLf (BapoLf) showed a bactericidal effect against M. haemolytica with MIC of 4.88 ± 1.88 and 7.31 ± 1.62 μM for M. haemolytica strain F (field isolate) and M. haemolytica strain R (reference strain), respectively. Through overlay assays and 2-D electrophoresis, two OMP of 32.9 and 34.2 kDa with estimated IP of 8.18 and 9.35, respectively, were observed to bind both BapoLf and BholoLf; these OMP were identified by Maldi-Tof as OmpA (heat-modifiable OMP) and a membrane protein (porin). These M. haemolytica BLf binding proteins could be interacting in vivo with both forms of BLf depending on the iron state of the bovine.

  1. LKTA and PlpE small fragments fusion protein protect against Mannheimia haemolytica challenge.

    Science.gov (United States)

    Guzmán-Brambila, Carolina; Quintero-Fabián, Saray; González-Castillo, Celia; de Obeso-Fernández del Valle, Álvaro; Flores-Samaniego, Beatriz; de la Mora, Germán; Rojas-Mayorquín, Argelia E; Ortuño-Sahagún, Daniel

    2012-12-01

    Bovine respiratory disease (BRD) complex is a major cause of economic losses for the cattle backgrounding and feedlot industries. Mannheimia haemolytica is considered the most important pathogen associated with this disease. Vaccines against M. haemolytica have been prepared and used for many decades, but traditional bacterins have failed to demonstrate effective protection and their use has often exacerbated disease in vaccinated animals. Thus, the BRD complex continues to exert a strong adverse effect on the health and wellbeing of stocker and feeder cattle. Therefore, generation of recombinant proteins has been helpful in formulating enhanced vaccines against M. haemolytica, which could confer better protection against BRD. In the present study, we formulated a vaccine preparation enriched with recombinant small fragments of leukotoxin A (LKTA) and outer-membrane lipoprotein (PlpE) proteins, and demonstrated its ability to generate high antibody titers in rabbits and sheep, which protected against M. haemolytica bacterial challenge in mice. Copyright © 2012 Elsevier Ltd. All rights reserved.

  2. Bibersteinia trehalosi inhibits the growth of mannheimia haemolytica by a proximity-dependent mechanism

    Science.gov (United States)

    Mannheimia (Pasteurella) haemolytica is the only pathogen that consistently causes severe bronchopneumonia and rapid death of bighorn sheep (BHS; Ovis canadensis) under experimental conditions. Paradoxically, Bibersteinia (Pasteurella) trehalosi and Pasteurella multocida have been isolated from BHS ...

  3. Bighorn sheep × domestic sheep hybrids survive Mannheimia haemolytica challenge in the absence of vaccination.

    Science.gov (United States)

    Subramaniam, R; Shanthalingam, S; Bavananthasivam, J; Kugadas, A; Raghavan, B; Batra, S A; Herndon, C N; Rodriguez, J; Tibary, A; Nelson, D; Potter, K A; Foreyt, W J; Srikumaran, S

    2014-06-04

    Bighorn sheep (BHS, Ovis canadensis) are much more susceptible than domestic sheep (DS, Ovis aries) to pneumonia caused by leukotoxin (Lkt)-producing members of the Family Pasteurellaceae, particularly Mannheimia haemolytica and Bibersteinia trehalosi. Leukotoxin is widely accepted as the critical virulence factor of these bacteria since Lkt-negative mutants do not cause death of BHS. Typically, DS carry Lkt-positive M. haemolytica and/or B. trehalosi as commensal bacteria in their nasopharynx. In contrast, most BHS do not carry Lkt-positive M. haemolytica or B. trehalosi, or carry Lkt-negative strains in their nasopharynx. In previous studies, we demonstrated that unimmunized DS resist M. haemolytica challenge while BHS succumb to it. We hypothesized that Lkt-neutralizing antibodies, induced by Lkt-positive M. haemolytica and/or B. trehalosi innately carried by DS in their nasopharynx, render them less susceptible to infection by these bacteria. In this study we developed BHS×DS F1 hybrids by artificial insemination of domestic ewes with BHS semen. F1 hybrids were fertile, and produced F2 hybrids and back-crosses. The F1, F2, and back-crosses were raised together with domestic ewes. All these animals acquired Lkt-positive M. haemolytica and/or B. trehalosi, and developed high titers of Lkt-neutralizing antibodies in the absence of vaccination. Furthermore, all of these animals resisted challenge with lethal dose of M. haemolytica. These results suggest that lack of previous exposure to Lkt is at least partially responsible for fatal pneumonia in BHS when they acquire Lkt-positive M. haemolytica and/or B. trehalosi from DS when the two species commingle. Copyright © 2014 Elsevier B.V. All rights reserved.

  4. Cancer cell injury by cytotoxins from cobra venom is mediated through lysosomal damage.

    Science.gov (United States)

    Feofanov, Alexei V; Sharonov, George V; Astapova, Maria V; Rodionov, Dmitriy I; Utkin, Yuriy N; Arseniev, Alexander S

    2005-08-15

    Cytotoxins from cobra venom are known to manifest cytotoxicity in various cell types. It is widely accepted that the plasma membrane is a target of cytotoxins, but the mechanism of their action remains obscure. Using the confocal spectral imaging technique, we show for the first time that cytotoxins from cobra venom penetrate readily into living cancer cells and accumulate markedly in lysosomes. Cytotoxins CT1 and CT2 from Naja oxiana, CT3 from Naja kaouthia and CT1 from Naja haje are demonstrated to possess this property with respect to human lung adenocarcinoma A549 and promyelocytic leukaemia HL60 cells. Immobilized plasma membrane binding accompanies the internalization of CT3 from Naja kaouthia in the HL60 cells, but it is very weak for other cytotoxins. Detectable membrane binding is not a property of any of the cytotoxins tested in A549 cells. The kinetics and concentration-dependence of cytotoxin accumulation in lysosomes correlate well with their cytotoxic effects. On the basis of the results obtained, we propose that lysosomes are a primary target of the lytic action of cytotoxins. Plasma membrane permeabilization seems to be a downstream event relative to lysosome rupture. Direct damage to the plasma membrane may be a complementary mechanism, but its relative contribution to the cytotoxic action depends on the cytotoxin structure and cell type.

  5. Cancer cell injury by cytotoxins from cobra venom is mediated through lysosomal damage

    Science.gov (United States)

    2005-01-01

    Cytotoxins from cobra venom are known to manifest cytotoxicity in various cell types. It is widely accepted that the plasma membrane is a target of cytotoxins, but the mechanism of their action remains obscure. Using the confocal spectral imaging technique, we show for the first time that cytotoxins from cobra venom penetrate readily into living cancer cells and accumulate markedly in lysosomes. Cytotoxins CT1 and CT2 from Naja oxiana, CT3 from Naja kaouthia and CT1 from Naja haje are demonstrated to possess this property with respect to human lung adenocarcinoma A549 and promyelocytic leukaemia HL60 cells. Immobilized plasma membrane binding accompanies the internalization of CT3 from Naja kaouthia in the HL60 cells, but it is very weak for other cytotoxins. Detectable membrane binding is not a property of any of the cytotoxins tested in A549 cells. The kinetics and concentration-dependence of cytotoxin accumulation in lysosomes correlate well with their cytotoxic effects. On the basis of the results obtained, we propose that lysosomes are a primary target of the lytic action of cytotoxins. Plasma membrane permeabilization seems to be a downstream event relative to lysosome rupture. Direct damage to the plasma membrane may be a complementary mechanism, but its relative contribution to the cytotoxic action depends on the cytotoxin structure and cell type. PMID:15847607

  6. Differential susceptibility of bighorn sheep and domestic sheep neutrophils to Mannheimia haemolytica leukotoxin is not due to differential expression of cell surface CD18

    Science.gov (United States)

    Bighorn sheep (BHS) are more susceptible to pneumonia caused by Mannheimia haemolytica (M. haemolytica) than are domestic sheep (DS). Leukotoxin produced by M. haemolytica is accepted as the critical virulence factor for BHS, based on the fact that Lkt-deletion mutants do not cause death of BHS. Al...

  7. Beta(2) integrin Mac-1 is a receptor for Mannheimia haemolytica leukotoxin on bovine and ovine leukocytes

    Science.gov (United States)

    Pneumonia caused by Mannheimia haemolytica is an important disease of cattle (BO), domestic sheep (DS, Ovis aries) and bighorn sheep (BHS, Ovis canadensis). Leukotoxin (Lkt) produced by M. haemolytica is cytolytic to all leukocyte subsets of these three species. Although it is certain that CD18, the...

  8. Phenotypic and genotypic characterization of Mannheimia (Pasteurella) haemolytica-like strains isolated from diseased animals in Denmark

    DEFF Research Database (Denmark)

    Angen, Øystein; Ahrens, Peter; Bisgaard, M.

    2002-01-01

    Trehalose-negative strains of the Pasteurella haemolytica complex have recently been transferred to a new genus, Mannheimia. This genus presently consists of five named species: M. haemolytica, M. glucosida, M. granulomatis, M. ruminalis and M. varigena. The purpose of this study was to investigate...

  9. Pathogen variation across time and space: sequencing to characterize Mannheimia haemolytica diversity.

    Science.gov (United States)

    Clawson, Michael L; Murray, Robert W

    2014-12-01

    Bovine respiratory disease complex (BRDC) is a major animal health and economic issue that affects cattle industries worldwide. Within the USA, the beef cattle industry loses up to an estimated 1 billion dollars a year due to BRDC. There are many contributors to BRDC, including environmental stressors and viral and/or bacterial infections. One species of bacteria in particular, Mannheimia haemolytica, is recognized as the major cause of severe fibrinonecrotic pneumonia in cattle. M. haemolytica is an opportunistic pathogen that normally populates the upper respiratory tract of cattle, and invades the lower respiratory tract in stressed and/or virally infected cattle by mechanisms that are not completely understood. However, not all M. haemolytica appear to be equally pathogenic to cattle. Thus, a test could be developed to distinguish M. haemolytica genetic subtypes by their propensity to cause respiratory disease, allowing isolation and/or treatment of cattle harboring strains with an increased propensity to cause disease. To that end, the genomes of over 300 M. haemolytica strains are being sequenced.

  10. Mannheimia haemolytica A2 secretes different proteases into the culture medium and in outer membrane vesicles.

    Science.gov (United States)

    Ramírez Rico, Gerardo; Martínez-Castillo, Moisés; González-Ruíz, Cynthia; Luna-Castro, Sarahí; de la Garza, Mireya

    2017-12-01

    Respiratory diseases in ruminants have a significantly negative impact on the worldwide economy. The bacterium Mannheimia haemolytica is involved in pneumonic infections in bovine and ovine. In gram-negative bacteria, six secretion systems related to the colonization process and host tissue damage have been reported. In addition, in the last two decades, the production of outer membrane vesicles has been studied as a different bacterial strategy to release virulence factors, such as exotoxins, lipopolysaccharides, and proteases. However, in M. haemolytica serotype A2, protease secretion and release in vesicles have not been reported as virulence mechanisms. The aim of this work was to identify proteases released into the culture supernatant and in vesicles of M. haemolytica A2. Our results showed evident differences in the molecular mass and activity of proteases present in culture supernatants and outer membrane vesicles based on zymography assays. The biochemical characterization of M. haemolytica proteases revealed that the main types were cysteine and metalloproteases. A specific metalloprotease of 100 kDa was active in the culture supernatants, but it was not active and was found in low quantities in vesicles. Proteases could be an important virulence factor during the infectious pneumonic process led by M. haemolytica. Copyright © 2017 Elsevier Ltd. All rights reserved.

  11. Nasal isolation of Mannheimia haemolytica and Pasteurella multocida as predictors of respiratory disease in shipped calves.

    Science.gov (United States)

    Taylor, J D; Holland, B P; Step, D L; Payton, M E; Confer, A W

    2015-04-01

    Three hundred ninety five calves were purchased from sale barns and delivered to the Willard Sparks Beef Research Center. Nasal swabs were collected to determine if presence of Mannheimia haemolytica and Pasteurella multocida in the upper respiratory tract (URT) can facilitate diagnosis of bovine respiratory disease (BRD). Samples were collected at arrival and at treatment for BRD. Clinically healthy control calves were sampled at time of treatment of sick calves. M. haemolytica was more commonly isolated from calves at treatment than at time of arrival or from control calves. M. haemolytica was more common in calves requiring treatment than in those never treated. Need for treatment and number of treatments were negatively associated with average daily gain, supporting the accuracy of diagnosis. These results suggest that URT sampling, when combined with clinical diagnosis, may assist in providing greater diagnostic accuracy, improving ability to evaluate risk factors, interventions, and treatments. Copyright © 2014 Elsevier Ltd. All rights reserved.

  12. Susceptibility to tulathromycin in Mannheimia haemolytica isolated from feedlot cattle over a three-year period

    Directory of Open Access Journals (Sweden)

    Trevor W. Alexander

    2013-10-01

    Full Text Available Mannheimia haemolytica isolated from feedlot cattle were tested for tulathromycin resistance. Cattle were sampled over a three-year period, starting 12 months after approval of tulathromycin for prevention and treatment of bovine respiratory disease. Nasopharyngeal samples from approximately 5,814 cattle were collected when cattle entered feedlots (N = 4 and again from the same cattle after ≥ 60 d on feed. The antimicrobial use history for each animal was recorded. Mannheimia haemolytica was isolated from 796 (13.7% entry samples and 1,038 (20.6% ≥ 60 d samples. Of the cattle positive for M. haemolytica, 18.5%, 2.9%, and 2.4% were administered therapeutic concentrations of tulathromycin, tilmicosin, or tylosin tartrate, respectively. In addition, 13.2% were administered subtherapeutic concentrations of tylosin phosphate in feed. In years one and two, no tulathromycin-resistant M. haemolytica were detected, whereas 5 isolates (0.4% were resistant in year three. These resistant isolates were collected from three cattle originating from a single pen, were all serotype 1, and were genetically related (≥ 89% similarity according to pulsed-field gel electrophoreses patterns. The five tulathromycin-resistant isolates were multi-drug resistant also exhibiting resistance to oxytetracycline, tilmicosin, ampicillin, or penicillin. The macrolide resistance genes erm(42, erm(A, erm(B, erm(F, erm(X and msr(E-mph(E, were not detected in the tulathromycin-resistant M. haemolytica. This study showed that tulathromycin resistance in M. haemolytica from a general population of feedlot cattle in western Canada was low and did not change over a three-year period after tulathromycin was approved for use in cattle.

  13. Effects of Actinobacillus pleuropneumoniae cytotoxins on generation of oxygen radicals by porcine neutrophils

    Directory of Open Access Journals (Sweden)

    Simson Tarigan

    1999-03-01

    Full Text Available Cytotoxins produced by Actinobacillus pleuropneumoniae (App suggested to be the most important pathogenic and virulent factors for this organism. However, the mechanisms on how the cytotoxins contribute to the disease process remain unclear. The purpose of this study is to investigate the effect of the cytotoxins on the oxidative-burst metabolism of porcine neutrophils. In this study, neutrophils were firstly loaded with an oxidative probe dichlorofluorescin diacetate (DCFHDA then expose to cytotoxins. Cells producing oxygen radicals emitted fluorescence and its intensity was measured with a FACScan flow cytometer. All cytotoxins derived from either App serotypes producing ApxI and ApxII, App serotypes producing ApxII only, or App serotypes producing ApxII and ApxIII were capable of stimulating neutrophils for oxygen-radical generation. However, compared with phorbol myristate acetate (PMA, App cytotoxins were much weaker as stimulants for oxygen radicals. In addition, Apx preparation stimulated an oxidative-burst metabolism of neutrophils at a low, narrow range of Apx doses. At higher doses, the toxins inhibit the oxidative burst metabolism. The effects of cytotoxins produced by App during infection on recruited neutrophils into the lungs are assumed to be comparable to those observed in this in vitro study. Neutrophils, and other host cells, adjacent to the bacteria become lysis due to high toxin concentration, whereas those at some distance to the bacteria produce oxygen radicals which in turn cause tissue damage or necrosis.

  14. Rational design and evaluation of mammalian ribonuclease cytotoxins.

    Science.gov (United States)

    Lomax, Jo E; Eller, Chelcie H; Raines, Ronald T

    2012-01-01

    Mammalian pancreatic-type ribonucleases (ptRNases) comprise an enzyme family that is remarkably well suited for therapeutic exploitation. ptRNases are robust and prodigious catalysts of RNA cleavage that can naturally access the cytosol. Instilling cytotoxic activity requires endowing them with the ability to evade a cytosolic inhibitor protein while retaining other key attributes. These efforts have informed our understanding of ptRNase-based cytotoxins, as well as the action of protein-based drugs with cytosolic targets. Here, we address the most pressing problems encountered in the design of cytotoxic ptRNases, along with potential solutions. In addition, we describe assays that can be used to evaluate a successful design in vitro, in cellulo, and in vivo. The emerging information validates the continuing development of ptRNases as chemotherapeutic agents. Copyright © 2012 Elsevier Inc. All rights reserved.

  15. Multiplex PCR To Identify Macrolide Resistance Determinants in Mannheimia haemolytica and Pasteurella multocida

    DEFF Research Database (Denmark)

    Rose, Simon; Desmolaize, Benoit; Jaju, Puneet

    2012-01-01

    isolates of P. multocida and M. haemolytica and correlated with MICs for the veterinary macrolides tulathromycin and tilmicosin, and the newer compounds gamithromycin and tildipirosin. The multiplex PCR system gives a rapid and robustly accurate determination of macrolide resistance genotypes and bacterial...

  16. Genome sequences of serotype A6 Mannheimia haemolytica isolates D174 and D38 recovered from bovine pneumonia

    Science.gov (United States)

    Here we report two genomes, one complete and one draft, from virulent bovine strains of Mannheimia haemolytica(strains D174 and D38)serotype A2 recovered prior to the field usage of modern antimicrobial drugs....

  17. Genome sequences of Mannheimia haemolytica serotype A1 strains D153 and D193 from bovine pneumonia

    Science.gov (United States)

    Here we report two genomes, one complete and one draft, from virulent bovine strains of Mannheimia haemolytica(strains D171 and D35)serotype A2 recovered prior to the field usage of modern antimicrobial drugs....

  18. Ulcerative colitis and Crohn's disease tissue cytotoxins

    Energy Technology Data Exchange (ETDEWEB)

    McLaren, L.C.; Gitnick, G.

    1982-06-01

    Bowel-wall tissue filtrates from patients with inflammatory bowel disease produce cytopathic effects in tissue culture. The cytopathic effects inducers have been reported to have the characteristics of a small RNA virus. Clostridium difficile toxin also produces cytopathic effects and has been found in the stools of patients with Crohn's disease and ulcerative colitis. The present study concerns the further characterization of the cytopathic inducers in tissues of inflammatory bowel disease patients. It was found that they are nonsedimentable at 148,000 g for 2 h and resistant to inactivation by UV light. They are proteins that are distinct from C. difficile toxin and are unique cytotoxins which are associated with the early cytopathic effects observed in Riff-free chick embryo and rabbit ileum cell cultures. These results suggest that the early cytopathic effects previously described are not produced by a virus. They do not explain the delayed cytopathic effects seen in rabbit ileum or WI-38 cells.

  19. Is Cytotoxin K from Bacillus cereus a bona fide enterotoxin?

    Science.gov (United States)

    Castiaux, Virginie; Liu, Xiaojin; Delbrassinne, Laurence; Mahillon, Jacques

    2015-10-15

    Cytotoxin K (CytK) produced by Bacillus cereus s.l. has generally been considered to be associated with the foodborne diarrhoeal syndrome. Two distinct variants of CytK have been reported: CytK-1 from Bacillus cytotoxicus and CytK-2 from B. cereus. In order to determine whether CytK plays a significant role in the diarrhoeal disease, the occurrence of cytK genes was assessed among 390 B. cereus isolates with different origins including clinical and food poisoning samples and was found to be 46%. Interestingly, the cytK occurrence was slightly lower in food poisoning and clinical isolates than in environmental samples. Seventy cytK-2 positive strains (including 28 isolates from foodborne outbreaks) were then selected in order to assess their genetic diversity. A genetic dendrogram based on the cytK-2 sequences of these 70 strains and on two cytK-1 sequences from strains NVH 391-98 and 883-00 showed an important diversity. However, no strain clustering according to the origin or source of isolation was observed. These observations were confirmed by Multi-Locus Sequences Typing (MLST) based on five different loci of housekeeping genes (ccpA, recF, sucC, purF and gdpD) for which no grouping of foodborne outbreak strains could be identified. Therefore, the choice of cytK as virulence factor for the diarrhoeal pathotype does not seem to be relevant per se, even though the involvement of CytK in the diarrhoeal syndrome cannot be fully excluded. Potential synergistic effects between CytK and other virulence factors, together with their potential variable expression levels should be further investigated. Copyright © 2015 Elsevier B.V. All rights reserved.

  20. Multiplex PCR To Identify Macrolide Resistance Determinants in Mannheimia haemolytica and Pasteurella multocida

    OpenAIRE

    Rose, Simon; Desmolaize, Benoit; Jaju, Puneet; Wilhelm, Cornelia; Warrass, Ralf; Douthwaite, Stephen

    2012-01-01

    The bacterial pathogens Mannheimia haemolytica and Pasteurella multocida are major etiological agents in respiratory tract infections of cattle. Although these infections can generally be successfully treated with veterinary macrolide antibiotics, a few recent isolates have shown resistance to these drugs. Macrolide resistance in members of the family Pasteurellaceae is conferred by combinations of at least three genes: erm(42), which encodes a monomethyltransferase and confers a type I MLSB ...

  1. PCR assay detects Mannheimia haemolytica in culture-negative pneumonic lung tissues of bighorn sheep (Ovis canadensis) from outbreaks in the western USA, 2009-2010.

    Science.gov (United States)

    Shanthalingam, Sudarvili; Goldy, Andrea; Bavananthasivam, Jegarubee; Subramaniam, Renuka; Batra, Sai Arun; Kugadas, Abirami; Raghavan, Bindu; Dassanayake, Rohana P; Jennings-Gaines, Jessica E; Killion, Halcyon J; Edwards, William H; Ramsey, Jennifer M; Anderson, Neil J; Wolff, Peregrine L; Mansfield, Kristin; Bruning, Darren; Srikumaran, Subramaniam

    2014-01-01

    Mannheimia haemolytica consistently causes severe bronchopneumonia and rapid death of bighorn sheep (Ovis canadensis) under experimental conditions. However, Bibersteinia trehalosi and Pasteurella multocida have been isolated from pneumonic bighorn lung tissues more frequently than M. haemolytica by culture-based methods. We hypothesized that assays more sensitive than culture would detect M. haemolytica in pneumonic lung tissues more accurately. Therefore, our first objective was to develop a PCR assay specific for M. haemolytica and use it to determine if this organism was present in the pneumonic lungs of bighorns during the 2009-2010 outbreaks in Montana, Nevada, and Washington, USA. Mannheimia haemolytica was detected by the species-specific PCR assay in 77% of archived pneumonic lung tissues that were negative by culture. Leukotoxin-negative M. haemolytica does not cause fatal pneumonia in bighorns. Therefore, our second objective was to determine if the leukotoxin gene was also present in the lung tissues as a means of determining the leukotoxicity of M. haemolytica that were present in the lungs. The leukotoxin-specific PCR assay detected leukotoxin gene in 91% of lung tissues that were negative for M. haemolytica by culture. Mycoplasma ovipneumoniae, an organism associated with bighorn pneumonia, was detected in 65% of pneumonic bighorn lung tissues by PCR or culture. A PCR assessment of distribution of these pathogens in the nasopharynx of healthy bighorns from populations that did not experience an all-age die-off in the past 20 yr revealed that M. ovipneumoniae was present in 31% of the animals whereas leukotoxin-positive M. haemolytica was present in only 4%. Taken together, these results indicate that culture-based methods are not reliable for detection of M. haemolytica and that leukotoxin-positive M. haemolytica was a predominant etiologic agent of the pneumonia outbreaks of 2009-2010.

  2. Multiplex PCR to identify macrolide resistance determinants in Mannheimia haemolytica and Pasteurella multocida.

    Science.gov (United States)

    Rose, Simon; Desmolaize, Benoit; Jaju, Puneet; Wilhelm, Cornelia; Warrass, Ralf; Douthwaite, Stephen

    2012-07-01

    The bacterial pathogens Mannheimia haemolytica and Pasteurella multocida are major etiological agents in respiratory tract infections of cattle. Although these infections can generally be successfully treated with veterinary macrolide antibiotics, a few recent isolates have shown resistance to these drugs. Macrolide resistance in members of the family Pasteurellaceae is conferred by combinations of at least three genes: erm(42), which encodes a monomethyltransferase and confers a type I MLS(B) (macrolide, lincosamide, and streptogramin B) phenotype; msr(E), which encodes a macrolide efflux pump; and mph(E), which encodes a macrolide-inactivating phosphotransferase. Here, we describe a multiplex PCR assay that detects the presence of erm(42), msr(E), and mph(E) and differentiates between these genes. In addition, the assay distinguishes P. multocida from M. haemolytica by amplifying distinctive fragments of the 23S rRNA (rrl) genes. One rrl fragment acts as a general indicator of gammaproteobacterial species and confirms whether the PCR assay has functioned as intended on strains that are negative for erm(42), msr(E), and mph(E). The multiplex system has been tested on more than 40 selected isolates of P. multocida and M. haemolytica and correlated with MICs for the veterinary macrolides tulathromycin and tilmicosin, and the newer compounds gamithromycin and tildipirosin. The multiplex PCR system gives a rapid and robustly accurate determination of macrolide resistance genotypes and bacterial genus, matching results from microbiological methods and whole-genome sequencing.

  3. Molecular epidemiology of an outbreak of clinical mastitis in sheep caused by Mannheimia haemolytica.

    Science.gov (United States)

    Omaleki, Lida; Browning, Glenn F; Allen, Joanne L; Markham, Philip F; Barber, Stuart R

    2016-08-15

    The aetiology and epidemiology of outbreaks of clinical mastitis in sheep under extensive pastoral conditions are incompletely understood. The objective of this study was to conduct a detailed investigation of a clinical mastitis outbreak that affected more than 10% of 230 at-risk ewes on a sheep and grain producing property in south east Australia during drought conditions in 2009. Milk samples were collected aseptically from all affected ewes and plated on sheep blood agar for bacterial identification. M. haemolytica was isolated from 80% of the samples that yielded cultivable microorganisms and thus was the main microorganism responsible for the outbreak. Analysis of the restriction endonuclease cleavage patterns of the isolates using pulsed field gel electrophoresis revealed some evidence of clonality, suggesting the possibility of horizontal transmission, but there was also considerable diversity between the clusters of closely related isolates. Multilocus sequence typing of the M. haemolytica isolates revealed most of the isolates belonged to ST1 with no association between the PFGE and MLST fingerprints of the isolates. Resistance to neomycin, streptomycin and sulphafurazole was detected in some of the isolates, but they were all susceptible to penicillin, ampicillin, ceftiofur, amoxycillin/clavulanic acid, ciprofloxacin, tetracycline, erythromycin and trimethoprim. This is the first published record of a comparison of the strains of M. haemolytica involved in a clinical mastitis outbreak in sheep and demonstrates the importance of this pathogen in sheep production systems, particularly during adverse climatic conditions and increased stocking rate. Copyright © 2016 Elsevier B.V. All rights reserved.

  4. Multiplex PCR To Identify Macrolide Resistance Determinants in Mannheimia haemolytica and Pasteurella multocida

    Science.gov (United States)

    Rose, Simon; Desmolaize, Benoit; Jaju, Puneet; Wilhelm, Cornelia; Warrass, Ralf

    2012-01-01

    The bacterial pathogens Mannheimia haemolytica and Pasteurella multocida are major etiological agents in respiratory tract infections of cattle. Although these infections can generally be successfully treated with veterinary macrolide antibiotics, a few recent isolates have shown resistance to these drugs. Macrolide resistance in members of the family Pasteurellaceae is conferred by combinations of at least three genes: erm(42), which encodes a monomethyltransferase and confers a type I MLSB (macrolide, lincosamide, and streptogramin B) phenotype; msr(E), which encodes a macrolide efflux pump; and mph(E), which encodes a macrolide-inactivating phosphotransferase. Here, we describe a multiplex PCR assay that detects the presence of erm(42), msr(E), and mph(E) and differentiates between these genes. In addition, the assay distinguishes P. multocida from M. haemolytica by amplifying distinctive fragments of the 23S rRNA (rrl) genes. One rrl fragment acts as a general indicator of gammaproteobacterial species and confirms whether the PCR assay has functioned as intended on strains that are negative for erm(42), msr(E), and mph(E). The multiplex system has been tested on more than 40 selected isolates of P. multocida and M. haemolytica and correlated with MICs for the veterinary macrolides tulathromycin and tilmicosin, and the newer compounds gamithromycin and tildipirosin. The multiplex PCR system gives a rapid and robustly accurate determination of macrolide resistance genotypes and bacterial genus, matching results from microbiological methods and whole-genome sequencing. PMID:22564832

  5. Transfection of non-susceptible cells with Ovis aries recombinant lymphocyte function-associated antigen 1 renders susceptibility to Mannheimia haemolytica leukotoxin

    Science.gov (United States)

    Mannheimia haemolytica is an important etiological agent of pneumonia in domestic sheep (DS, Ovis aries). Leukotoxin (Lkt) produced by this organism is the principal virulence factor responsible for the acute inflammation and lung injury characteristic of M. haemolytica caused disease. Previously, w...

  6. New interpretive criteria for danofloxacin antibacterial susceptibility testing against Mannheimia haemolytica and Pasteurella multocida associated with bovine respiratory disease.

    Science.gov (United States)

    Sweeney, Michael T; Papich, Mark G; Watts, Jeffrey L

    2017-03-01

    Danofloxacin is a fluoroquinolone antibacterial agent approved for use in veterinary medicine to treat and control bovine respiratory disease caused by Mannheimia haemolytica or Pasteurella multocida. Susceptible minimal inhibitory concentration (MIC) breakpoint (≤0.25 µg/mL) and disk diffusion interpretive criteria (≥22 mm) values for danofloxacin against M. haemolytica and P. multocida were first approved by the Clinical and Laboratory Standards Institute (CLSI) in 2003. However, intermediate and resistant breakpoint values were not established because only susceptible wild-type populations were evident at the time of breakpoint approvals. Since then, nonsusceptible isolates of M. haemolytica and P. multocida have been identified. We report danofloxacin intermediate MIC breakpoint (0.5 µg/mL) and disk diffusion interpretive criteria (18-21 mm), as well as danofloxacin-resistant MIC breakpoint (≥1 µg/mL) and disk diffusion interpretive criteria (≤17 mm), based on scattergram plots of MIC values versus disk zone diameters and calculated error-bound rates using M. haemolytica and P. multocida isolates recovered from bovine respiratory disease in North America in 2004-2014. These newly established intermediate and resistant clinical breakpoint values have been endorsed by CLSI and can be used for interpreting results from antibacterial susceptibility testing of danofloxacin against M. haemolytica and P. multocida isolated from bovine respiratory disease.

  7. BIÓPSIA PULMONAR EM BEZERROS COM BRONCOPNEUMONIA INDUZIDA PELA Mannheimia haemolytica PULMONAR BIOPSY IN CALVES WITH BRONCHOPNEUMONIA INDUCED BY Mannheimia haemolytica

    Directory of Open Access Journals (Sweden)

    Daniel Pessoa Gomes da Silva

    2009-09-01

    Full Text Available Com o propósito de avaliar a segurança, a eficácia diagnóstica da biópsia pulmonar e comparar a possível ocorrência de complicações decorrentes da técnica, entre bezerros sadios e com broncopneumonia induzida, utilizaram-se dez bezerros (G1 sadios e vinte bezerros portadores de broncopneumonia, divididos em quatro grupos de cinco bezerros (G2 a G5, os quais foram biopsiados 12, 24, 48 e 72 horas após a inoculação com Mannheimia haemolytica, respectivamente. A presença de crepitação grossa, som submaciço à percussão e as alterações radiográficas auxiliaram no diagnóstico da broncopneumonia e localizaram a área pulmonar a ser biopsiada nos grupos G2 a G5. As alterações microscópicas, visualizadas nos animais do grupo G2 a G5, foram as de broncopneumonia fibrinopurulenta. Nos bezerros do grupo G1 as alterações relacionadas à técnica foram: tosse, epistaxe, dispneia mista, taquipneia e taquicardia. Cinco (25% bezerros com broncopneumonia desenvolveram hemotórax após a biópsia e as alterações relacionadas à técnica foram: taquipneia, taquicardia, tosse, dispneia mista, apatia, mucosas pálidas e decúbito. Conclui-se que a biópsia pulmonar permite o diagnóstico de broncopneumonia em bezerros, contudo as complicações decorrentes da técnica limitam o seu uso apenas nas situações em que os métodos convencionais não tenham possibilitado o diagnóstico.

    PALAVRAS-CHAVES: Bezerros, biópsia pulmonar, broncopneumonia, Mannheimia haemolytica.

    The purpose of this study was to evaluate the safety and diagnostic efficacy of lung biopsy and to compare the possible occurrence of complications due to this technique in healthy calves and in calves with bronchopneumonia. Thirty Holstein calves were divided into a group of ten healthy animals (G1 and into four experimental groups (G2 to G5 of five calves each

  8. A novel Erm monomethyltransferase in antibiotic-resistant isolates of Mannheimia haemolytica and Pasteurella multocida

    DEFF Research Database (Denmark)

    Desmolaize, Benoit; Rose, Simon; Warrass, Ralf

    2011-01-01

    Mannheimia haemolytica and Pasteurella multocida are aetiological agents commonly associated with respiratory tract infections in cattle. Recent isolates of these pathogens have been shown to be resistant to macrolides and other ribosome-targeting antibiotics. Direct analysis of the 23S r......RNAs by mass spectrometry revealed that nucleotide A2058 is monomethylated, consistent with a Type I erm phenotype conferring macrolide-lincosamide resistance. The erm resistance determinant was identified by full genome sequencing of isolates. The sequence of this resistance determinant, now termed erm(42...

  9. Macrolide resistance conferred by rRNA mutations in field isolates of Mannheimia haemolytica and Pasteurella multocida

    DEFF Research Database (Denmark)

    Olsen, Anders S; Warrass, Ralf; Douthwaite, Stephen Roger

    2014-01-01

    . haemolytica identified as being highly resistant (MICs >64 mg/L) to the macrolides erythromycin, gamithromycin, tilmicosin, tildipirosin and tulathromycin were screened by multiplex PCR for the previously identified resistance genes erm(42), msr(E) and mph(E). Strains lacking these determinants were analysed...

  10. Macrolide resistance conferred by rRNA mutations in field isolates of Mannheimia haemolytica and Pasteurella multocida.

    Science.gov (United States)

    Olsen, Anders S; Warrass, Ralf; Douthwaite, Stephen

    2015-02-01

    To determine how resistance to macrolides is conferred in field isolates of Pasteurella multocida and Mannheimia haemolytica that lack previously identified resistance determinants for rRNA methylation, efflux and macrolide-modifying enzymes. Isolates of P. multocida and M. haemolytica identified as being highly resistant (MICs >64 mg/L) to the macrolides erythromycin, gamithromycin, tilmicosin, tildipirosin and tulathromycin were screened by multiplex PCR for the previously identified resistance genes erm(42), msr(E) and mph(E). Strains lacking these determinants were analysed by genome sequencing and primer extension on the rRNAs. Macrolide resistance in one M. haemolytica isolate was conferred by the 23S rRNA mutation A2058G; resistance in three P. multocida isolates were caused by mutations at the neighbouring nucleotide A2059G. In each strain, all six copies of the rrn operons encoded the respective mutations. There were no mutations in the ribosomal protein genes rplD or rplV, and no other macrolide resistance mechanism was evident. High-level macrolide resistance can arise from 23S rRNA mutations in P. multocida and M. haemolytica despite their multiple copies of rrn. Selective pressures from exposure to different macrolide or lincosamide drugs presumably resulted in consolidation of either the A2058G or the A2059G mutation. © The Author 2014. Published by Oxford University Press on behalf of the British Society for Antimicrobial Chemotherapy. All rights reserved. For Permissions, please e-mail: journals.permissions@oup.com.

  11. Effect of subtherapeutic vs therapeutic administration of macrolides on antimicrobial resistance in Mannheimia haemolytica and enterococci isolated from beef cattle

    Directory of Open Access Journals (Sweden)

    Rahat eZaheer

    2013-05-01

    Full Text Available Macrolides are the first-line treatment against bovine respiratory disease, and are also used to treat infections in humans. The macrolide, tylosin phosphate, is often included in the diet of cattle as a preventative for liver abscesses in many regions of the world outside of Europe. This study investigated the effects of administering macrolides to beef cattle either systemically through a single subcutaneous injection (therapeutic or continuously in-feed (subtherapeutic, on the prevalence and antimicrobial resistance of Mannheimia haemolytica and Enterococcus spp. isolated from the nasopharynx and faeces, respectively. Nasopharyngeal and faecal samples were collected weekly over 28 days from untreated beef steers and from steers injected once with tilmicosin or tulathromycin or continuously fed tylosin phosphate at dosages recommended by manufacturers. Tilmicosin and tulathromycin were effective in lowering (P < 0.05 the prevalence of M. haemolytica, whereas subtherpeutic tylosin had no effect. M. haemolytica isolated from control- and macrolide-treated animals were susceptible to macrolides as well as to other antibiotics. Major bacteria co-isolated with M. haemolytica included Pasteurella multocida, Staphylococcus spp., Acinetobacter spp., E. coli and Bacillus spp. With the exception of M. haemolytica and P. multocida, erythromycin resistance was frequently found in other isolated species. Both methods of macrolide administration increased (P < 0.05 the levels erythromycin-resistance enterococci in faeces. Development of resistance to injectable macrolides in bacteria isolated from the nasopharynx was species dependent. Therapeutic administration of tilmicosin and tularthromycin selected for macrolide resistant bacteria within both the respiratory and intestinal tract, whereas suptherapeutic administration of tylosin only selected for macrolide resistance in enteric bacteria.

  12. The Shiga and Shiga-Like Cytotoxins: Gene Regulation and Functional Analysis of the Binding Subunits

    Science.gov (United States)

    1989-05-05

    disease (hemolytic Eshcerichia coli toxemia) in pigs and mice. Am. J. Vet. Res. Jan. issue:88-94. Griffin, D. E., and P. Gemski. 1983. Release of...Shiga-like toxin production by Eshcerichia coli . Infect. Immun. 56: .106-111. Willshaw, G. A., H. R. Smith, s. M. Scotland and B. Rowe. 1985. Cloning...directed by: Alison D. O’Brien, Ph.D., Professor, Department of Microbiology Strains of Escherichia coli produce cytotoxins which are related to the

  13. Hamiltonian purification

    Energy Technology Data Exchange (ETDEWEB)

    Orsucci, Davide [Scuola Normale Superiore, I-56126 Pisa (Italy); Burgarth, Daniel [Department of Mathematics, Aberystwyth University, Aberystwyth SY23 3BZ (United Kingdom); Facchi, Paolo; Pascazio, Saverio [Dipartimento di Fisica and MECENAS, Università di Bari, I-70126 Bari (Italy); INFN, Sezione di Bari, I-70126 Bari (Italy); Nakazato, Hiromichi; Yuasa, Kazuya [Department of Physics, Waseda University, Tokyo 169-8555 (Japan); Giovannetti, Vittorio [NEST, Scuola Normale Superiore and Istituto Nanoscienze-CNR, I-56126 Pisa (Italy)

    2015-12-15

    The problem of Hamiltonian purification introduced by Burgarth et al. [Nat. Commun. 5, 5173 (2014)] is formalized and discussed. Specifically, given a set of non-commuting Hamiltonians (h{sub 1}, …, h{sub m}) operating on a d-dimensional quantum system ℋ{sub d}, the problem consists in identifying a set of commuting Hamiltonians (H{sub 1}, …, H{sub m}) operating on a larger d{sub E}-dimensional system ℋ{sub d{sub E}} which embeds ℋ{sub d} as a proper subspace, such that h{sub j} = PH{sub j}P with P being the projection which allows one to recover ℋ{sub d} from ℋ{sub d{sub E}}. The notions of spanning-set purification and generator purification of an algebra are also introduced and optimal solutions for u(d) are provided.

  14. Pharmacokinetic/pharmacodynamic integration and modelling of amoxicillin for the calf pathogens Mannheimia haemolytica and Pasteurella multocida.

    Science.gov (United States)

    Lees, P; Pelligand, L; Illambas, J; Potter, T; Lacroix, M; Rycroft, A; Toutain, P-L

    2015-10-01

    The antimicrobial properties of amoxicillin were determined for the bovine respiratory tract pathogens, Mannheima haemolytica and Pasteurella multocida. Minimum inhibitory concentration (MIC), minimum bactericidal concentration (MBC) and time-kill curves were established. Pharmacokinetic (PK)/pharmacodynamic (PD) modelling of the time-kill data, based on the sigmoidal Emax equation, generated parameters for three levels of efficacy, namely bacteriostatic, bactericidal (3log10 reduction) and 4log10 reduction in bacterial counts. For these levels, mean AUC(0-24 h) /MIC serum values for M. haemolytica were 29.1, 57.3 and 71.5 h, respectively, and corresponding values for P. multocida were 28.1, 44.9 and 59.5 h. Amoxicillin PK was determined in calf serum, inflamed (exudate) and noninflamed (transudate) tissue cage fluids, after intramuscular administration of a depot formulation at a dosage of 15 mg/kg. Mean residence times were 16.5 (serum), 29.6 (exudate) and 29.0 h (transudate). Based on serum MICs, integration of in vivo PK and in vitro PD data established maximum concentration (Cmax )/MIC ratios of 13.9:1 and 25.2:1, area under concentration-time curve (AUC0-∞ )/MIC ratios of 179 and 325 h and T>MIC of 40.3 and 57.6 h for P. multocida and M. haemolytica, respectively. Monte Carlo simulations for a 90% target attainment rate predicted single dose to achieve bacteriostatic and bactericidal actions over 48 h of 17.7 and 28.3 mg/kg (M. haemolytica) and 17.7 and 34.9 mg/kg (P. multocida). © 2015 John Wiley & Sons Ltd.

  15. Polonium purification

    Energy Technology Data Exchange (ETDEWEB)

    Baker, J.D.

    1996-09-01

    Three processes for the purification of {sup 210}Po from irradiated bismuth targets are described. Safety equipment includes shielded hotcells for the initial separation from other activation products, gloveboxes for handling the volatile and highly toxic materials, and provisions for ventilation. All chemical separations must be performed under vacuum or in inerted systems. Two of the processes require large amounts of electricity; the third requires vessels made from exotic materials.

  16. Prevalence of Helicobacter pylori vacuolating cytotoxin and its allelic mosaicism as a predictive marker for Iranian dyspeptic patients

    DEFF Research Database (Denmark)

    Mohammadi, M; Oghalaie, A; Mohajerani, N

    2003-01-01

    can serve as screening markers for such a population, H. pylori strains were isolated from one hundred and thirty two dyspeptic patients. H. pylori genomic DNA was extracted and underwent PCR-amplification for the cytotoxin alleles. Genotyping of the signal sequence region of the vacA gene identified......Helicobacter pylori infects the majority of the population in the developing countries. However, the rate of gastrointestinal complications such as peptic ulcers and gastric malignancies has no parallel with the infection. In order to determine whether cytotoxin (vacA) and its allelic polymorphism...

  17. Staphylococcus epidermidis and Staphylococcus haemolyticus: Molecular Detection of Cytotoxin and Enterotoxin Genes.

    Science.gov (United States)

    Pinheiro, Luiza; Brito, Carla Ivo; de Oliveira, Adilson; Martins, Patrícia Yoshida Faccioli; Pereira, Valéria Cataneli; da Cunha, Maria de Lourdes Ribeiro de Souza

    2015-09-14

    Although opportunistic pathogens, coagulase-negative staphylococci (CoNS), including Staphylococcus epidermidis and Staphylococcus haemolyticus, have long been regarded as avirulent organisms. The role of toxins in the development of infections caused by CoNS is still controversial. The objective of this study was to characterize the presence of enterotoxin and cytotoxin genes in S. epidermidis and S. haemolyticus isolates obtained from blood cultures. Cytotoxin genes were detected by PCR using novel species-specific primers. Among the 85 S. epidermidis and 84 S. haemolyticus isolates, 95.3% and 79.8%, respectively, carried at least one enterotoxin gene. The most frequent enterotoxin genes were sea (53.3%), seg (64.5%) and sei (67.5%). The seg gene was positively associated with S. epidermidis (p = 0.02), and this species was more toxigenic than S. haemolyticus. The hla/yidD gene was detected in 92.9% of S. epidermidis and the hla gene in 91.7% of S. haemolyticus isolates; hlb was detected in 92.9% of the S. epidermidis isolates and hld in 95.3%. Nosocomial Staphylococcus epidermidis and S. haemolyticus isolates exhibited a high toxigenic potential, mainly producing the non-classical enterotoxins seg and sei. The previously unreported detection of hla/yidD and hlb in S. epidermidis and S. haemolyticus using species-specific primers showed that these hemolysin genes differ between CoNS species and that they are highly frequent in blood culture isolates.

  18. Biochemical characterization of a polysialyltransferase from Mannheimia haemolytica A2 and comparison to other bacterial polysialyltransferases.

    Directory of Open Access Journals (Sweden)

    Theresa Lindhout

    Full Text Available Polysialic acids are bioactive carbohydrates found in eukaryotes and some bacterial pathogens. The bacterial polysialyltransferases (PSTs, which catalyze the synthesis of polysialic acid capsules, have previously been identified in select strains of Escherichia coli and Neisseria meningitidis and are classified in the Carbohydrate-Active enZYmes Database as glycosyltransferase family GT-38. In this study using DNA sequence analysis and functional characterization we have identified a novel polysialyltransferase from the bovine/ovine pathogen Mannheimia haemolytica A2 (PSTMh. The enzyme was expressed in recombinant form as a soluble maltose-binding-protein fusion in parallel with the related PSTs from E. coli K1 and N. meningitidis group B in order to perform a side-by-side comparison. Biochemical properties including solubility, acceptor preference, reaction pH optima, thermostability, kinetics, and product chain length for the enzymes were compared using a synthetic fluorescent acceptor molecule. PSTMh exhibited biochemical properties that make it an attractive candidate for chemi-enzymatic synthesis applications of polysialic acid. The activity of PSTMh was examined on a model glycoprotein and the surface of a neuroprogenitor cell line where the results supported its development for use in applications to therapeutic protein modification and cell surface glycan remodelling to enable cell migration at implantation sites to promote wound healing. The three PSTs examined here demonstrated different properties that would each be useful to therapeutic applications.

  19. Susceptibilidade in vitro a antimicrobianos da Mannheimia haemolytica e da Pasteurella multocida isoladas de ovinos sadios e com doenças respiratórias

    Directory of Open Access Journals (Sweden)

    Leomar Viana

    2007-12-01

    Full Text Available Pasteurella multocida e Mannheimia haemolytica (P. haemolytica estão associadas a enfermidades no sistema respiratório de ovinos. Com o objetivo de avaliar a susceptibilidade in vitro destes microrganismos frente aos antimicrobianos, foram colhidas amostras de nasofaringe (n=180 e orofaringe (n=82 de ovinos com e sem enfermidade respiratória. Dentre os antimicrobianos testados, a sensibilidade foi maior para enrofloxacina (100% e florfenicol (100%, considerando-se ambas as espécies bacterianas. Observou-se resistência de M. haemolytica e P. multocida à tetraciclina (15,64% e 17,65%, respectivamente e penicilina (1.82% e 4.2%, respectivamente.

  20. The occurrence of subtilase-cytotoxin-encoding genes in environmental Escherichia coli isolated from a Northern California estuary.

    Science.gov (United States)

    Pereira, Maria das Graças C; Byrne, Barbara A; Nguyen, Trân B H; Lewis, David J; Atwill, E Robert

    2013-06-01

    The presence of subtilase-cytotoxin-encoding genes was determined in 397 environmental Escherichia coli strains isolated from water, suspended solids, and sediments sampled from different hydrological and environmental conditions in a California estuary. A total of 7 strains (1.76%) were found to harbor subtilase-cytotoxin-encoding genes. Using primers targeting subA only, we generated PCR amplicons from 2 strains; while using primers targeting the 3' end of SubA downstream to the 5' end of SubB, amplicons of 232 bp were generated from 5 additional strains. The 556 bp subA sequences were almost identical to that in the subtilase-cytotoxin-positive strain ED 591 (98%), while subAB sequences of 2 non-Shiga-toxigenic strains revealed 100% similarity with the Shiga-toxigenic E. coli O113:H21 strain 98NK2 that was isolated from an outbreak of hemolytic uremic syndrome. Additionally, the serogroup O113:H21 was present in this collection of environmental E. coli, and it was found to harbor stx2d, hra1 that encodes the heat resistant agglutinin 1, and a subAB sequence similar to that in the non-Shiga-toxigenic E. coli subtilase cytotoxin strain ED 591. To further understand potential health risks posed by strains encoding SubAB, future epidemiological studies should consider screening isolates for subAB regardless of the presence of Shiga-toxin-encoding genes.

  1. Ocorrência de pneumonia associada à infecção por Mannheimia haemolytica em ovinos de Minas Gerais Occurrence of pneumonia associated to infection by Mannheimia haemolytica in sheep of Minas Gerais

    Directory of Open Access Journals (Sweden)

    Marina Rios de Araújo

    2009-09-01

    Full Text Available O trabalho descreve um surto de pneumonia em ovinos em uma propriedade na região central de Minas Gerais. Clinicamente os animais apresentavam apatia, mostravam dificuldade respiratória durante dois ou três dias ou morriam subitamente. À necropsia as alterações pulmonares eram similares em todos os ovinos. Havia consolidação dos lobos craniais e da parte ventral dos lobos caudais e ao corte fluía exsudato mucopurulento da traquéia e dos brônquios. No parênquima dos lobos craniais havia áreas brancas multifocais a coalescentes com 0,2-0,5cm de diâmetro, levemente proeminentes e intercaladas por áreas vermelho-escuras. Pleurite fibrinosa foi observada nos Ovinos 1, 2 e 3. As lesões de consolidação ocupavam cerca de 70-80% da extensão pulmonar. Microscopicamente, as alterações eram de broncopneumonia fibrinopurulenta com intensa hiperemia, áreas com hemorragia intra-alveolar e espessamento dos septos interlobulares por inúmeros neutrófilos, restos celulares e intensa exsudação de fibrina. Áreas multifocais com necrose de liquefação contendo numerosas colônias bacterianas foram observadas no Ovino 3. Nos lobos craniais dos Ovinos 1, 2 e 3, haviam áreas com neutrófilos degenerados formando aglomerados de células alongadas com formato de "grãos de aveia" associados a colônias bacterianas. As alterações histológicas foram características de pneumonia causada por Mannheimia (M. haemolytica. Amostras dos lobos craniais de todos os ovinos foram encaminhadas para cultivo bacteriológico e M. haemolytica foi isolada e identificada em todos os animais. Este é o primeiro relato correlacionando os achados patológicos e o isolamento de M. haemolytica como causa de broncopneumonia em ovinos no Brasil.This paper describes an outbreak of pneumonia in a sheep herd in the central region of Minas Gerais, Brazil. Clinically, the animals presented apathy, exhibited respiratory difficulty during 2 to 3 days or sudden death. The

  2. An ecologic study comparing distribution of Pasteurella trehalosi and Mannheimia haemolytica between Sierra Nevada bighorn sheep, White Mountain bighorn sheep, and domestic sheep.

    Science.gov (United States)

    Tomassini, Letizia; Gonzales, Ben; Weiser, Glen C; Sischo, William

    2009-10-01

    The prevalence and phenotypic variability of Pasteurella and Mannheimia isolates from Sierra Nevada bighorn sheep (Ovis canadensis sierrae), White Mountain bighorn sheep (Ovis canadensis nelsoni), and domestic sheep (Ovis aries) from California, USA, were compared. The White Mountain bighorn sheep population had a recent history of pneumonia-associated mortality, whereas the Sierra Nevada bighorn sheep population had no recent history of pneumonia-associated mortality. The domestic sheep flocks were pastured in areas geographically near both populations but were not known to have direct contact with either bighorn sheep population. Oropharyngeal swab samples were collected from healthy domestic and bighorn sheep and cultured to characterize bacterial species, hemolysis, biogroups, and biovariants. Pasteurella trehalosi and Mannheimia haemolytica were detected in all of the study populations, but the relative proportion of each bacterial species differed among sheep populations. Pasteurella trehalosi was more common than M. haemolytica in the bighorn sheep populations, whereas the opposite was true in domestic sheep. Mannheimia haemolytica was separated into 11 biogroups, and P. trehalosi was characterized into two biogroups. Biogroup distributions for M. haemolytica and P. trehalosi differed among the three populations; however, no difference was detected for the distribution of P. trehalosi biogroups between the Sierra Nevada bighorn sheep and domestic sheep. The prevalence odds ratios (pOR) for the distribution of M. haemolytica biogroups suggested little difference between White Mountain bighorn sheep and domestic sheep compared with Sierra Nevada bighorn sheep and domestic sheep, although these comparisons had relatively large confidence intervals for the point estimates. Hemolytic activity of the isolates was not different among the sheep populations for M. haemolytica but was different for P. trehalosi. No clear evidence of association was found in the

  3. An outbreak of Vero cytotoxin producing Escherichia coli O157 infection associated with takeaway sandwiches.

    LENUS (Irish Health Repository)

    McDonnell, R J

    1997-12-12

    An outbreak of food poisoning due to Escherichia coli O157 phage type 2 Vero cytotoxin 2 affected 26 people in southern counties of England in May and June 1995. The organism was isolated from faecal specimens from 23 patients, 16 of whom lived in Dorset and seven in Hampshire. Isolates were indistinguishable by phage typing, Vero cytotoxin gene typing, restriction fragment length polymorphism, and pulsed field gel electrophoresis. Three associated cases, linked epidemiologically to the outbreak, were confirmed serologically by detection of antibodies to E. coli O157 lipopolysaccharide. Twenty-two of the 26 patients were adults: four were admitted to hospital with haemorrhagic colitis. Four cases were children: two were admitted to hospital with haemolytic uraemic syndrome (HUS). There were no deaths. Although E. coli O157 was not isolated from any food samples, illness was associated with having eaten cold meats in sandwiches bought from two sandwich producers, in Weymouth and in Portsmouth. Both shops were supplied by the same wholesaler, who kept no records and obtained cooked meats from several sources in packs that did not carry adequate identification marks. It was, therefore, impossible to trace back to the original producer or to investigate further to determine the origin of contamination with E. coli O157. To protect the public health it is essential that all wholesale packs of ready-to-eat food carry date codes and the producer\\'s identification mark. Detailed record keeping should be part of hazard analysis critical control point (HACCP) systems and should be maintained throughout the chain of distribution from the producer to retail outlets.

  4. Complexities of the pathogenesis of Mannheimia haemolytica and Haemophilus somnus infections: challenges and potential opportunities for prevention?

    Science.gov (United States)

    Czuprynski, Charles J; Leite, Fabio; Sylte, Matt; Kuckleburg, C; Schultz, Ron; Inzana, Tom; Behling-Kelly, Erica; Corbeil, Lynette

    2004-12-01

    Progress in producing improved vaccines against bacterial diseases of cattle is limited by an incomplete understanding of the pathogenesis of these agents. Our group has been involved in investigations of two members of the family Pasteurellaceae, Mannheimia haemolytica and Haemophilus somnus, which illustrate some of the complexities that must be confronted. Susceptibility to M. haemolytica is greatly increased during active viral respiratory infection, resulting in rapid onset of a severe and even lethal pleuropneumonia. Despite years of investigation, understanding of the mechanisms underlying this viral-bacterial synergism is incomplete. We have investigated the hypothesis that active viral infection increases the susceptibility of bovine leukocytes to the M. haemolytica leukotoxin by increasing the expression of or activating the beta2 integrin CD11a/CD18 (LFA-1) on the leukocyte surface. In vitro exposure to proinflammatory cytokines (i.e. interleukin-1beta, tumor necrosis factor-alpha and interferon-gamma) increases LFA-1 expression on bovine leukocytes, which in turn correlates with increased binding and responsiveness to the leukotoxin. Alveolar macrophages and peripheral blood leukocytes from cattle with active bovine herpesvirus-1 (BVH-1) infection are more susceptible to the lethal effects of the leukotoxin ex vivo than leukocytes from uninfected cattle. Likewise, in vitro incubation of bovine leukocytes with bovine herpesvirus 1 (BHV-1) potentiates LFA-1 expression and makes the cells more responsive to leukotoxin. A striking characteristic of H. somnus infection is its propensity to cause vasculitis. We have shown that H. somnus and its lipo-oligosaccharide (LOS) trigger caspase activation and apoptosis in bovine endothelial cells in vitro. This effect is associated with the production of reactive oxygen and nitrogen intermediates, and is amplified in the presence of platelets. The adverse effects of H. somnus LOS are mediated in part by activation of

  5. Untersuchungen zur Wirksamkeit einer Mannheimia-haemolytica-Adsorbatvakzine in Beziehung zur Immunitätslage von Mastkälbern

    OpenAIRE

    Scheller, Regina

    2004-01-01

    Untersuchungen zur Wirksamkeit einer Mannheimia-haemolytica-Adsorbatvakzine in Beziehung zur Immunitätslage von Mastkälbern aus dem Institut für Bakteriologie und Mykologie der Veterinärmedizinischen Fakultät der Universität Leipzig (140 S., 15 Abb., 18 Tab., 407 Lit.) Die Enzootische Bronchopneumonie des Rindes stellt einen der zentralen und wirtschaftlich bedeutsamsten Krankheitskomplexe in der Rinderaufzucht und –mast dar. Unter den Bedingungen der konzentrierten Stallhaltung größerer Tier...

  6. Genomic signatures of Mannheimia haemolytica that associate with the lungs of cattle with respiratory disease, an integrative conjugative element, and antibiotic resistance genes

    Science.gov (United States)

    Background: Mannheimia haemolytica typically resides in cattle as a commensal member of the upper respiratory tract microbiome. However, some strains can invade their lungs and cause respiratory disease and death, including those with multi-drug resistance. A nucleotide polymorphism typing system ...

  7. Transmission of Mannheimia haemolytica from domestic sheep (Ovis aries) to bighorn sheep (Ovis canadensis): unequivocal demonstration with green fluorescent protein-tagged organisms.

    Science.gov (United States)

    Lawrence, Paulraj K; Shanthalingam, Sudarvili; Dassanayake, Rohana P; Subramaniam, Renuka; Herndon, Caroline N; Knowles, Donald P; Rurangirwa, Fred R; Foreyt, William J; Wayman, Gary; Marciel, Ann Marie; Highlander, Sarah K; Srikumaran, Subramaniam

    2010-07-01

    Previous studies demonstrated that bighorn sheep (Ovis canadensis) died of pneumonia when commingled with domestic sheep (Ovis aries) but did not conclusively prove that the responsible pathogens were transmitted from domestic to bighorn sheep. The objective of this study was to determine, unambiguously, whether Mannheimia haemolytica can be transmitted from domestic to bighorn sheep when they commingle. Four isolates of M. haemolytica were obtained from the pharynx of two of four domestic sheep and tagged with a plasmid carrying the genes for green fluorescent protein (GFP) and ampicillin resistance (AP(R)). Four domestic sheep, colonized with the tagged bacteria, were kept about 10 m apart from four bighorn sheep for 1 mo with no clinical signs of pneumonia observed in the bighorn sheep during that period. The domestic and bighorn sheep were then allowed to have fence-line contact for 2 mo. During that period, three bighorn sheep acquired the tagged bacteria from the domestic sheep. At the end of the 2 mo of fence-line contact, the animals were allowed to commingle. All four bighorn sheep died 2 days to 9 days following commingling. The lungs from all four bighorn sheep showed gross and histopathologic lesions characteristic of M. haemolytica pneumonia. Tagged M. haemolytica were isolated from all four bighorn sheep, as confirmed by growth in ampicillin-containing culture medium, PCR-amplification of genes encoding GFP and Ap(R), and immunofluorescent staining of GFP. These results unequivocally demonstrate transmission of M. haemolytica from domestic to bighorn sheep, resulting in pneumonia and death of bighorn sheep.

  8. Lindbladian purification

    Science.gov (United States)

    Arenz, Christian; Burgarth, Daniel; Giovannetti, Vittorio; Nakazato, Hiromichi; Yuasa, Kazuya

    2017-06-01

    In a recent work (Burgarth et al 2014, Nat. Commun. 5 5173), it was shown that a series of frequent measurements can project the dynamics of a quantum system onto a subspace in which the dynamics can be more complex. In this subspace, even full controllability can be achieved, although the controllability over the system before the projection is very poor since the control Hamiltonians commute with each other. We can also think of the opposite: any Hamiltonians of a quantum system, which are in general noncommutative with each other, can be made commutative by embedding them in an extended Hilbert space, thus the dynamics in the extended space becomes trivial and simple. This idea of making noncommutative Hamiltonians commutative is called ‘Hamiltonian purification.’ The original noncommutative Hamiltonians are recovered by projecting the system back onto the original Hilbert space through frequent measurements. Here, we generalise this idea to open-system dynamics by presenting a simple construction to make Lindbladians, as well as Hamiltonians, commutative on a larger space with an auxiliary system. We show that the original dynamics can be recovered through frequently measuring the auxiliary system in a non-selective way. Moreover, we provide a universal pair of Lindbladians that describe an ‘accessible’ open quantum system for generic system sizes. This allows us to conclude that through a series of frequent non-selective measurements a nonaccessible open quantum system generally becomes accessible. This sheds further light on the role of measurement backaction on the control of quantum systems.

  9. Comparative study of structure and activity of cytotoxins from venom of the cobras Naja oxiana, Naja kaouthia, and Naja haje.

    Science.gov (United States)

    Feofanov, A V; Sharonov, G V; Dubinnyi, M A; Astapova, M V; Kudelina, I A; Dubovskii, P V; Rodionov, D I; Utkin, Yu N; Arseniev, A S

    2004-10-01

    Cytotoxins are positively charged polypeptides that constitute about 60% of all proteins in cobra venom; they have a wide spectrum of biological activities. By CD spectroscopy, cytotoxins CT1 and CT2 Naja oxiana, CT3 Naja kaouthia, and CT1 and CT2 Naja haje were shown to have similar secondary structure in an aqueous environment, with dominating beta-sheet structure, and to vary in the twisting angle of the beta-sheet and the conformation of disulfide groups. Using dodecylphosphocholine micelles and liposomes, CT1 and CT2 Naja oxiana were shown to incorporate into lipid structures without changes in the secondary structure of the peptides. The binding of CT1 and CT2 Naja oxiana with liposomes was associated with an increase in the beta-sheet twisting and a sign change of the dihedral angle of one disulfide group. The cytotoxins were considerably different in cytotoxicity and cooperativity of the effect on human promyelocytic leukemia cells HL60, mouse myelomonocytic cells WEHI-3, and human erythroleukemic cells K562. The most toxic CT2 Naja oxiana and CT3 Naja kaouthia possessed low cooperativity of interaction (Hill coefficient h = 0.6-0.8), unlike 10-20-fold less toxic CT1 and CT2 Naja haje (h = 1.2-1.7). CT1 Naja oxiana has an intermediate position on the cytotoxicity scale and is characterized by h = 0.5-0.8. The cytotoxins under study induced necrosis of HL60 cells and failed to activate apoptosis. The differences in cytotoxicity are supposed to be related not with features of the secondary structure of the peptides, but with interactions of side chains of variable amino acid residues with lipids and/or membrane proteins.

  10. Estimating risk of C. difficile transmission from PCR positive but cytotoxin negative cases.

    Directory of Open Access Journals (Sweden)

    Mini Kamboj

    Full Text Available The use of molecular methods to diagnose Clostridium difficile infection (CDI has improved diagnostic yield compared to conventional methods. However, PCR testing can detect colonization and has introduced several practical challenges pertaining to need for treatment and isolation of cases.For all new cases detected by real-time PCR, concurrent cytotoxin assay was performed and genetic characterization with MLVA (multi-locus variable number tandem repeat analysis was done to determine relatedness. We used PCR cycle threshold (Ct of detection as surrogate marker for bacterial burden in stool.Overall, 54 cases of CDI were detected during the study period. 42 were concurrently tested by CYT and characterized by MLVA .MLVA analysis revealed marked genetic diversity with no ongoing outbreaks; four cases were due to NAP1 strain. CYT -/PCR + cases had a higher median Ct value of detection compared to CYT+/PCR + cases (28.2 vs 22.5; p = 0.01. Among 25 strains that were genetically related, 9/11 isolates in this dominant cluster were positive by CYT compared to 4/14 in non-dominant clusters (p = 0.02.CYT-/PCR+ cases contribute to hospital based transmission. However, the risk of transmission of C. difficile from CYT +/PCR+ cases may be higher than those that are CYT-/PCR+.

  11. Melanoma-specific ferrocene esters of the fungal cytotoxin illudin M.

    Science.gov (United States)

    Knauer, Sebastian; Biersack, Bernhard; Zoldakova, Miroslava; Effenberger, Katharina; Milius, Wolfgang; Schobert, Rainer

    2009-09-01

    The unfavorable therapeutic index of the fungal cytotoxin illudin M was to be improved by covalent attachment of the redox modulator and phenyl isobiostere ferrocene. Esters of illudin M with ferrocenoic and 1,1'-ferrocenedioic acid were prepared, structurally characterised (X-ray), and tested for cytotoxicity [MTT assay, 3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyltetrazolium bromide], induction of apoptosis (TUNEL assay; western blotting for caspase-9), and tumor specificity in cells of human HL-60 leukemia, human 518A2 melanoma, and in nonmalignant human foreskin fibroblasts. The diester of illudin M with 1,1'-ferrocenedioic acid was distinctly more antiproliferative and apoptosis inducing in the melanoma cells [half maximal inhibitory concentration, IC50(48 h) = 0.4+/-0.1 micromol/l] than in the HL-60 cells [IC50(48 h) = 3.0+/-1.6 micromol/l] and in the nonmalignant fibroblasts [IC50(48 h) = 3.7+/-1.9 micromol/l]. This corresponds to a doubling of the therapeutic index with respect to illudin M. The monoester of illudin M with ferrocenoic acid was nine times less efficacious in the cancer cells, when compared with the diester. In conclusion, the ferrocene diminishes the general toxicity of the illudin M moiety and increases its cell line specificity. The bis(illudinyl M) 1,1'-ferrocenedioate presumably operates by a synergistic, two-pronged attack on its molecular targets.

  12. DXD Motif-Dependent and -Independent Effects of the Chlamydia trachomatis Cytotoxin CT166

    Directory of Open Access Journals (Sweden)

    Miriam Bothe

    2015-02-01

    Full Text Available The Gram-negative, intracellular bacterium Chlamydia trachomatis causes acute and chronic urogenital tract infection, potentially leading to infertility and ectopic pregnancy. The only partially characterized cytotoxin CT166 of serovar D exhibits a DXD motif, which is important for the enzymatic activity of many bacterial and mammalian type A glycosyltransferases, leading to the hypothesis that CT166 possess glycosyltransferase activity. CT166-expressing HeLa cells exhibit actin reorganization, including cell rounding, which has been attributed to the inhibition of the Rho-GTPases Rac/Cdc42. Exploiting the glycosylation-sensitive Ras(27H5 antibody, we here show that CT166 induces an epitope change in Ras, resulting in inhibited ERK and PI3K signaling and delayed cell cycle progression. Consistent with the hypothesis that these effects strictly depend on the DXD motif, CT166 with the mutated DXD motif causes neither Ras-ERK inhibition nor delayed cell cycle progression. In contrast, CT166 with the mutated DXD motif is still capable of inhibiting cell migration, suggesting that CT166 with the mutated DXD motif cannot be regarded as inactive in any case. Taken together, CT166 affects various fundamental cellular processes, strongly suggesting its importance for the intracellular survival of chlamydia.

  13. Snake cytotoxins bind to membranes via interactions with phosphatidylserine head groups of lipids.

    Directory of Open Access Journals (Sweden)

    Anastasia G Konshina

    Full Text Available The major representatives of Elapidae snake venom, cytotoxins (CTs, share similar three-fingered fold and exert diverse range of biological activities against various cell types. CT-induced cell death starts from the membrane recognition process, whose molecular details remain unclear. It is known, however, that the presence of anionic lipids in cell membranes is one of the important factors determining CT-membrane binding. In this work, we therefore investigated specific interactions between one of the most abundant of such lipids, phosphatidylserine (PS, and CT 4 of Naja kaouthia using a combined, experimental and modeling, approach. It was shown that incorporation of PS into zwitterionic liposomes greatly increased the membrane-damaging activity of CT 4 measured by the release of the liposome-entrapped calcein fluorescent dye. The CT-induced leakage rate depends on the PS concentration with a maximum at approximately 20% PS. Interestingly, the effects observed for PS were much more pronounced than those measured for another anionic lipid, sulfatide. To delineate the potential PS binding sites on CT 4 and estimate their relative affinities, a series of computer simulations was performed for the systems containing the head group of PS and different spatial models of CT 4 in aqueous solution and in an implicit membrane. This was done using an original hybrid computational protocol implementing docking, Monte Carlo and molecular dynamics simulations. As a result, at least three putative PS-binding sites with different affinities to PS molecule were delineated. Being located in different parts of the CT molecule, these anion-binding sites can potentially facilitate and modulate the multi-step process of the toxin insertion into lipid bilayers. This feature together with the diverse binding affinities of the sites to a wide variety of anionic targets on the membrane surface appears to be functionally meaningful and may adjust CT action against

  14. Helicobacter pylori γ-glutamyl transpeptidase and vacuolating cytotoxin promote gastric persistence and immune tolerance.

    Science.gov (United States)

    Oertli, Mathias; Noben, Manuel; Engler, Daniela B; Semper, Raphaela P; Reuter, Sebastian; Maxeiner, Joachim; Gerhard, Markus; Taube, Christian; Müller, Anne

    2013-02-19

    Infection with the gastric bacterial pathogen Helicobacter pylori is typically contracted in early childhood and often persists for decades. The immunomodulatory properties of H. pylori that allow it to colonize humans persistently are believed to also account for H. pylori's protective effects against allergic and chronic inflammatory diseases. H. pylori infection efficiently reprograms dendritic cells (DCs) toward a tolerogenic phenotype and induces regulatory T cells (Tregs) with highly suppressive activity in models of allergen-induced asthma. We show here that two H. pylori virulence determinants, the γ-glutamyl transpeptidase GGT and the vacuolating cytotoxin VacA, contribute critically and nonredundantly to H. pylori's tolerizing effects on murine DCs in vitro and in vivo. The tolerance-promoting effects of both factors are independent of their described suppressive activity on T cells. Isogenic H. pylori mutants lacking either GGT or VacA are incapable of preventing LPS-induced DC maturation and fail to drive DC tolerization as assessed by induction of Treg properties in cocultured naive T cells. The Δggt and ΔvacA mutants colonize mice at significantly reduced levels, induce stronger T-helper 1 (Th1) and T-helper 17 (Th17) responses, and/or trigger more severe gastric pathology. Both factors promote the efficient induction of Tregs in vivo, and VacA is required to prevent allergen-induced asthma. The defects of the Δggt mutant in vitro and in vivo are phenocopied by pharmacological inhibition of the transpeptidase activity of GGT in all readouts. In conclusion, our results reveal the molecular players and mechanistic basis for H. pylori-induced immunomodulation, promoting persistent infection and conferring protection against allergic asthma.

  15. Action of shiga toxin type-2 and subtilase cytotoxin on human microvascular endothelial cells.

    Directory of Open Access Journals (Sweden)

    María M Amaral

    Full Text Available The hemolytic uremic syndrome (HUS associated with diarrhea is a complication of Shiga toxin (Stx-producing Escherichia coli (STEC infection. In Argentina, HUS is endemic and responsible for acute and chronic renal failure in children younger than 5 years old. The human kidney is the most affected organ due to the presence of very Stx-sensitive cells, such as microvascular endothelial cells. Recently, Subtilase cytotoxin (SubAB was proposed as a new toxin that may contribute to HUS pathogenesis, although its action on human glomerular endothelial cells (HGEC has not been described yet. In this study, we compared the effects of SubAB with those caused by Stx2 on primary cultures of HGEC isolated from fragments of human pediatric renal cortex. HGEC were characterized as endothelial since they expressed von Willebrand factor (VWF and platelet/endothelial cell adhesion molecule 1 (PECAM-1. HGEC also expressed the globotriaosylceramide (Gb3 receptor for Stx2. Both, Stx2 and SubAB induced swelling and detachment of HGEC and the consequent decrease in cell viability in a time-dependent manner. Preincubation of HGEC with C-9 -a competitive inhibitor of Gb3 synthesis-protected HGEC from Stx2 but not from SubAB cytotoxic effects. Stx2 increased apoptosis in a time-dependent manner while SubAB increased apoptosis at 4 and 6 h but decreased at 24 h. The apoptosis induced by SubAB relative to Stx2 was higher at 4 and 6 h, but lower at 24 h. Furthermore, necrosis caused by Stx2 was significantly higher than that induced by SubAB at all the time points evaluated. Our data provide evidence for the first time how SubAB could cooperate with the development of endothelial damage characteristic of HUS pathogenesis.

  16. Bacillus cereus cytotoxins Hbl, Nhe and CytK are secreted via the Sec translocation pathway

    Directory of Open Access Journals (Sweden)

    Lindbäck Toril

    2010-11-01

    Full Text Available Abstract Background Bacillus cereus and the closely related Bacillus thuringiensis are Gram positive opportunistic pathogens that may cause food poisoning, and the three secreted pore-forming cytotoxins Hbl, Nhe and CytK have been implicated as the causative agents of diarrhoeal disease. It has been proposed that the Hbl toxin is secreted using the flagellar export apparatus (FEA despite the presence of Sec-type signal peptides. As protein secretion is of key importance in virulence of a microorganism, the mechanisms by which these toxins are secreted were further investigated. Results Sec-type signal peptides were identified in all toxin components, and secretion of Hbl component B was shown to be dependent on an intact Sec-type signal peptide sequence. Further indication that secretion of Hbl, Nhe and CytK is dependent on the Sec translocation pathway, the main pathway on which bacterial secretion relies, was suggested by the observed intracellular accumulation and reduced secretion of the toxins in cultures supplemented with the SecA inhibitor sodium azide. Although a FEA deficient strain (a flhA mutant showed reduced toxin expression and reduced cytotoxicity, it readily secreted overexpressed Hbl B, showing that the FEA is not required for Hbl secretion. Thus, the concurrent lack of flagella and reduced toxin secretion in the FEA deficient strain may point towards the presence of a regulatory link between motility and virulence genes, rather than FEA-dependent toxin secretion. Conclusions The Hbl, Nhe and CytK toxins appear to be secreted using the Sec pathway, and the reduced Hbl expression of a FEA deficient strain was shown not to be due to a secretion defect.

  17. Impact of timing and dosage of a fluoroquinolone treatment on the microbiological, pathological and clinical outcomes of calves challenged with Mannheimia haemolytica

    Directory of Open Access Journals (Sweden)

    Guillaume eLhermie

    2016-03-01

    Full Text Available The efficacy of an early and low inoculum-adjusted marbofloxacin treatment was evaluated on microbiological and clinical outcomes in calves infected with 4.107 CFU of Mannheimia haemolytica A1. Twenty-two calves were included based on their rectal temperature rise in the 10 h after challenge and allocated in 4 groups, receiving a single intramuscular injection of saline (CON, 2 mg/kg marbofloxacin 2-4 h after inclusion (early treatment, E2, 2 or 10 mg/kg marbofloxacin 35-39 h after inclusion (late treatments, L2, L10. In CON calves, M. haemolytica DNA loads in bronchoalveolar lavages continuously increased from inclusion to day 4, and were associated with persistent respiratory clinical signs and lung lesions. At times of early and late treatments, M. haemolytica loads ranged within 3.5-4 and 5.5-6 log10 DNA copies/mL, respectively. Early 2 mg/kg marbofloxacin treatment led to rapid and total elimination of bacteria in all calves. The late treatments induced a reduction of bacterial loads, but 3 of 6 L2 and 1 of 6 L10 calves were still positive for M. haemolytica at day 4. Except for CON calves, all animals exhibited clinical improvement within 24h after treatment. However, early 2 mg/kg treatment was more efficacious to prevent pulmonary lesions, as indicated by the reduction of the extension and severity of gross lesions and by the histopathological scores. These results demonstrated for the first time that a reduced antibiotic regimen given at an early stage of the disease and targeting a low bacterial load could be efficacious in a natural bovine model of pneumonia.

  18. Comparison of standardised versus non-standardised methods for testing the in vitro potency of oxytetracycline against Mannheimia haemolytica and Pasteurella multocida.

    Science.gov (United States)

    Lees, P; Illambas, J; Pelligand, L; Toutain, P-L

    2016-12-01

    The in vitro pharmacodynamics of oxytetracycline was established for six isolates of each of the calf pneumonia pathogens Mannheimia haemolytica and Pasteurella multocida. Minimum inhibitory concentration (MIC), minimum bactericidal concentration (MBC) and bacterial time-kill curves were determined in two matrices, Mueller Hinton broth (MHB) and calf serum. Geometric mean MIC ratios, serum:MHB, were 25.2:1 (M. haemolytica) and 27.4:1 (P. multocida). The degree of binding of oxytetracycline to serum protein was 52.4%. Differences between serum and broth MICs could not be accounted for by oxytetracycline binding to serum protein. In vitro time-kill data suggested a co-dependent killing action of oxytetracycline. The in vitro data indicate inhibition of the killing action of oxytetracycline by serum factor(s). The nature of the inhibition requires further study. The outcome of treatment with oxytetracycline of respiratory tract infections in calves caused by M. haemolytica and P. multocida may not be related solely to a direct killing action. Copyright © 2016 Elsevier Ltd. All rights reserved.

  19. Snake Venom Cytotoxins, Phospholipase A2s, and Zn2+-dependent Metalloproteinases: Mechanisms of Action and Pharmacological Relevance

    Science.gov (United States)

    Gasanov, Sardar E; Dagda, Ruben K; Rael, Eppie D

    2014-01-01

    Snake venom toxins are responsible for causing severe pathology and toxicity following envenomation including necrosis, apoptosis, neurotoxicity, myotoxicity, cardiotoxicity, profuse hemorrhage, and disruption of blood homeostasis. Clinically, snake venom toxins therefore represent a significant hazard to snakebite victims which underscores the need to produce more efficient anti-venom. Some snake venom toxins, however, have great potential as drugs for treating human diseases. In this review, we discuss the biochemistry, structure/function, and pathology induced by snake venom toxins on human tissue. We provide a broad overview of cobra venom cytotoxins, catalytically active and inactive phospholipase A2s (PLA2s), and Zn2+-dependent metalloproteinases. We also propose biomedical applications whereby snake venom toxins can be employed for treating human diseases. Cobra venom cytotoxins, for example, may be utilized as anti-cancer agents since they are efficient at destroying certain types of cancer cells including leukemia. Additionally, increasing our understanding of the molecular mechanism(s) by which snake venom PLA2s promote hydrolysis of cell membrane phospholipids can give insight into the underlying biomedical implications for treating autoimmune disorders that are caused by dysregulated endogenous PLA2 activity. Lastly, we provide an exhaustive overview of snake venom Zn2+-dependent metalloproteinases and suggest ways by which these enzymes can be engineered for treating deep vein thrombosis and neurodegenerative disorders. PMID:24949227

  20. Snake Venom Cytotoxins, Phospholipase A2s, and Zn(2+)-dependent Metalloproteinases: Mechanisms of Action and Pharmacological Relevance.

    Science.gov (United States)

    Gasanov, Sardar E; Dagda, Ruben K; Rael, Eppie D

    2014-01-25

    Snake venom toxins are responsible for causing severe pathology and toxicity following envenomation including necrosis, apoptosis, neurotoxicity, myotoxicity, cardiotoxicity, profuse hemorrhage, and disruption of blood homeostasis. Clinically, snake venom toxins therefore represent a significant hazard to snakebite victims which underscores the need to produce more efficient anti-venom. Some snake venom toxins, however, have great potential as drugs for treating human diseases. In this review, we discuss the biochemistry, structure/function, and pathology induced by snake venom toxins on human tissue. We provide a broad overview of cobra venom cytotoxins, catalytically active and inactive phospholipase A2s (PLA2s), and Zn(2+)-dependent metalloproteinases. We also propose biomedical applications whereby snake venom toxins can be employed for treating human diseases. Cobra venom cytotoxins, for example, may be utilized as anti-cancer agents since they are efficient at destroying certain types of cancer cells including leukemia. Additionally, increasing our understanding of the molecular mechanism(s) by which snake venom PLA2s promote hydrolysis of cell membrane phospholipids can give insight into the underlying biomedical implications for treating autoimmune disorders that are caused by dysregulated endogenous PLA2 activity. Lastly, we provide an exhaustive overview of snake venom Zn(2+)-dependent metalloproteinases and suggest ways by which these enzymes can be engineered for treating deep vein thrombosis and neurodegenerative disorders.

  1. The Borexino purification system

    Science.gov (United States)

    Benziger, Jay

    2014-05-01

    Purification of 278 tons of liquid scintillator and 889 tons of buffer shielding for the Borexino solar neutrino detector is performed with a system of combined distillation, water extraction, gas stripping and filtration. The purification system removed K, U and Th by distillation of the pseudocumene solvent and the PPO fluor. Noble gases, Rn, Kr and Ar were removed by gas stripping. Distillation was also employed to remove optical impurities and reduce the attenuation of scintillation light. The success of the purification system has facilitated the first time real time detection of low energy solar neutrinos.

  2. General outbreaks of vero cytotoxin producing Escherichia coli O157 in England and Wales from 1992 to 1994.

    LENUS (Irish Health Repository)

    Wall, P G

    1996-02-02

    We have reviewed all general outbreaks of infection due to Vero cytotoxin producing Escherichia coli (VTEC) O157 reported in England and Wales from 1992 to 1994. One hundred and seventy-three people were affected in 18 outbreaks, compared with 76 people in seven outbreaks in the preceding three years (1989 to 1991). Outbreaks occurred throughout England and Wales. Thirty-eight per cent of cases were admitted to hospital, 21% developed haemolytic uraemic syndrome, and 3% died. VTEC O157 infection causes particular concern because of its serious complications--haemorrhagic colitis and haemolytic uraemic syndrome, its capacity to spread from person to person as well as by food and water, and its reservoir in dairy and beef cattle.

  3. The amino acid sequence of toxin V II 2, a cytotoxin homologue from banded Egyptian cobra (Naja haje annulifera) venom.

    Science.gov (United States)

    Joubert, F J

    1975-12-01

    Toxin V II 2 comprises 60 amino acid residues and is cross-linked by four disulphide bridges. The complete amino acid sequence of this toxin was elucidated. The reduced and S-carboxymethylated toxin was digested with trypsin and chymotrypsin and the peptides were purified by ion-exchange chromatography and chromatography or electrophoresis on paper. The Edman procedure, either through the use of the automatic sequenator or by manual manipulation, was employed to obtain the sequence of the intact toxin and the pure peptides. The chymotryptic digest provided the necessary overlapping peptides which allowed the alignment of tryptic peptides. The amino acid sequence of Naja haje annulifera toxin V II 2 shows a high degree of homology with cytotoxin V II 1 of the same venom.

  4. Targeting the tumour vasculature: exploitation of low oxygenation and sensitivity to NOS inhibition by treatment with a hypoxic cytotoxin.

    Directory of Open Access Journals (Sweden)

    Jennifer H E Baker

    Full Text Available Many cancer research efforts focus on exploiting genetic-level features that may be targeted for therapy. Tissue-level features of the tumour microenvironment also represent useful therapeutic targets. Here we investigate the presence of low oxygen tension and sensitivity to NOS inhibition of tumour vasculature as potential tumour-specific features that may be targeted by hypoxic cytotoxins, a class of therapeutics currently under investigation. We have previously demonstrated that tirapazamine (TPZ mediates central vascular dysfunction in tumours. TPZ is a hypoxic cytotoxin that is also a competitive inhibitor of NOS. Here we further investigated the vascular-targeting activity of TPZ by combining it with NOS inhibitor L-NNA, or with low oxygen content gas breathing. Tumours were analyzed via multiplex immunohistochemical staining that revealed irreversible loss of perfusion and enhanced tumour cell death when TPZ was combined with either low oxygen or a NOS inhibitor. Tumour growth rate was reduced by TPZ + NOS inhibition, and tumours previously resistant to TPZ-mediated vascular dysfunction were sensitized by low oxygen breathing. Additional mapping analysis suggests that tumours with reduced vascular-associated stroma may have greater sensitivity to these effects. These results indicate that poorly oxygenated tumour vessels, also being abnormally organized and with inadequate smooth muscle, may be successfully targeted for significant anti-cancer effects by inhibition of NOS and hypoxia-activated prodrug toxicity. This strategy illustrates a novel use of hypoxia-activated cytotoxic prodrugs as vascular targeting agents, and also represents a novel mechanism for targeting tumour vessels.

  5. Pulmonary lesions and clinical disease response to Mannheimia haemolytica challenge 10 days following administration of tildipirosin or tulathromycin.

    Science.gov (United States)

    Amrine, D E; White, B J; Larson, R L; Mosier, D A

    2014-01-01

    This clinical trial evaluated the impact of metaphylactic antimicrobial administration 10 d before experimental inoculation with Mannheimia haemolytica (MH) to mitigate pulmonary lesions. Thirty-three crossbreed heifers were procured as a single group and were randomly allocated to 1 of 3 blocks and to treatment, tildipirosin (ZUP; 4 mg/kg) or tulathromycin (DRX; 2.5 mg/kg) or saline (SAL; 1 mL/45.5 kg), within block on arrival at Kansas State University. All trial procedures were staggered by 7-d intervals for each block, resulting in all animals within a block receiving treatment, challenge, and necropsy on the same dates. Heifers within each block received an endoscopic MH challenge 10 d following treatment administration (d 0) and were housed in individual indoor stalls for 3 d postchallenge. Clinical illness scores (CIS), respiration quality scores, appetite scores, and injection site reactions were recorded on all animals from d 0 through d 13. Rectal temperatures were measured once daily on all animals from d 8 through d 13. Heifers were necropsied, and lung lesions were evaluated on d 13. Lung lesion data were evaluated using nonparametric methods (Kruskall-Wallis), and standard least squares models were used to evaluate the remaining variables. The pulmonary lesion scores (percentage of affected lung) ranged from 3.3% to 39.8% for all heifers with 92% (11/12) of ZUP-treated heifers having Tildipirosin-treated heifers had lower (P tildipirosin 10 d before MH challenge have less pulmonary damage and fewer clinical signs of illness compared to heifers treated with DRX or SAL.

  6. Water purification in Borexino

    Energy Technology Data Exchange (ETDEWEB)

    Giammarchi, M. [Infn Milano (Italy); Balata, M.; Ioannucci, L.; Nisi, S. [Laboratori Nazionali del Gran Sasso (Italy); Goretti, A.; Ianni, A. [Princeton University (United States); Miramonti, L. [Dip. di Fisica dell' Università di Milano e Infn (Italy)

    2013-08-08

    Astroparticle Physics and Underground experiments searching for rare nuclear events, need high purity materials to act as detectors or detector shielding. Water has the advantage of being cheap, dense and easily available. Most of all, water can be purified to the goal of obatining a high level of radiopurity. Water Purification can be achieved by means of a combination of processes, including filtration, reverse osmosis, deionization and gas stripping. The Water Purification System for the Borexino experiment, will be described together with its main performances.

  7. Water purification in Borexino

    Science.gov (United States)

    Giammarchi, M.; Balata, M.; Goretti, A.; Ianni, A.; Ioannucci, L.; Miramonti, L.; Nisi, S.

    2013-08-01

    Astroparticle Physics and Underground experiments searching for rare nuclear events, need high purity materials to act as detectors or detector shielding. Water has the advantage of being cheap, dense and easily available. Most of all, water can be purified to the goal of obatining a high level of radiopurity. Water Purification can be achieved by means of a combination of processes, including filtration, reverse osmosis, deionization and gas stripping. The Water Purification System for the Borexino experiment, will be described together with its main performances.

  8. Differential Susceptibility of Bighorn Sheep (Ovis canadensis) and Domestic Sheep (Ovis aries) Neutrophils to Mannheimia haemolytica Leukotoxin is not due to Differential Expression of Cell Surface CD18.

    Science.gov (United States)

    Dassanayake, Rohana P; Shanthalingam, Sudarvili; Liu, Weiguo; Casas, Eduardo; Srikumaran, Subramaniam

    2017-07-01

    Bighornsheep ( Ovis canadensis ) are more susceptible to pneumonia caused by Mannheimia haemolytica than are domestic sheep ( Ovis aries ). Leukotoxin produced by M. haemolytica is the principal virulence factor involved in pneumonia pathogenesis. Although leukotoxin is cytolytic to all subsets of ruminant leukocytes, neutrophils are the most susceptible subset. Bighorn sheep neutrophils are four- to eightfold more susceptible to leukotoxin-induced cytolysis than are domestic sheep neutrophils. We hypothesized that the higher susceptibility of bighorn sheep neutrophils, in comparison to domestic sheep neutrophils, is due to higher expression of CD18, the receptor for leukotoxin on leukocytes. Our objective was to quantify CD18 expression on neutrophils of bighorn sheep and domestic sheep. Cell-surface CD18 expression on bighorn sheep and domestic sheep neutrophils was measured as antibody binding capacity of cells by flow cytometric analysis with two fluorochrome-conjugated anti-CD18 monoclonal antibodies (BAQ30A and HUH82A) and microspheres. Contrary to our expectations, CD18 expression was higher (Psheep neutrophils in comparison to bighorn sheep neutrophils. These findings suggest that the higher in vitro susceptibility to leukotoxin of bighorn sheep neutrophils compared to domestic sheep neutrophils is not due to higher expression of the leukotoxin receptor CD18 on bighorn sheep neutrophils.

  9. Coarse-grained molecular dynamics simulations of cobra cytotoxin A3 interactions with a lipid bilayer: penetration of loops into membranes.

    Science.gov (United States)

    Su, Zhi-Yuan; Wang, Yeng-Tseng

    2011-02-10

    Cobra cytotoxins, which are small three-looped proteins composed of approximately 60 amino acid residues, primarily act by destroying the bilayer membranes of cells and artificial vesicles. However, the molecular mechanism governing this process is not yet completely understood. We used coarse-grained molecular dynamics (CGMD) simulations to study the mechanism underlying the penetration of cardiotoxin A3 (CTX A3), the major toxic component of Naja atra (Chinese cobra) venom, into a hydrated 1-palmitoyl-2-oleoyl-1-sn-3-phosphatidylcholine (POPC) lipid bilayer. We performed CGMD simulations for three different conformations of the cobra cytotoxin-the tail, lying, and harrow conformations. The results of our simulations indicate that two of these, the tail and lying conformations, did not penetrate the bilayer system. Further, for the harrow conformation, loops 2 and 3 played important roles in penetration of CTX A3 into the bilayer system.

  10. Expression, Purification and Characterization of Ricin vectors used for exogenous antigen delivery into the MHC Class I presentation pathway

    Directory of Open Access Journals (Sweden)

    Smith Daniel C.

    2003-01-01

    Full Text Available Disarmed versions of the cytotoxin ricin can deliver fused peptides into target cells leading to MHC class I-restricted antigen presentation [Smith et al. J Immunol 2002; 169:99-107]. The ricin delivery vector must contain an attenuated catalytic domain to prevent target cell death, and the fused peptide epitope must remain intact for delivery and functional loading to MHC class I molecules. Expression in E. coli and purification by cation exchange chromatography of the fusion protein is described. Before used for delivery, the activity of the vector must be characterized in vitro, via an N-glycosidase assay, and in vivo, by a cytotoxicity assay. The presence of an intact epitope must be confirmed using mass spectrometry by comparing the actual mass with the predicted mass.

  11. Sat, the secreted autotransporter toxin of uropathogenic Escherichia coli, is a vacuolating cytotoxin for bladder and kidney epithelial cells.

    Science.gov (United States)

    Guyer, Debra M; Radulovic, Suzana; Jones, Faye-Ellen; Mobley, Harry L T

    2002-08-01

    The secreted autotransporter toxin (Sat) of uropathogenic Escherichia coli exhibits cytopathic activity upon incubation with HEp-2 cells. We further investigated the effects of Sat on cell lines more relevant to the urinary tract, namely, those derived from bladder and kidney epithelium. Sat elicited elongation of cells and apparent loosening of cellular junctions upon incubation with Vero kidney cells. Additionally, incubation with Sat triggered significant vacuolation within the cytoplasm of both human bladder (CRL-1749) and kidney (CRL-1573) cell lines. This activity has been associated with only a few other known toxins. Following transurethral infection of CBA mice with a sat mutant, no reduction of CFU in urine, bladder, or kidney tissue was seen compared to that in mice infected with wild-type E. coli CFT073. However, significant histological changes were observed within the kidneys of mice infected with wild-type E. coli CFT073, including dissolution of the glomerular membrane and vacuolation of proximal tubule cells. Such damage was not observed in kidney sections of mice infected with a Sat-deficient mutant. These results indicate that Sat, a vacuolating cytotoxin expressed by uropathogenic E. coli CFT073, elicits defined damage to kidney epithelium during upper urinary tract infection and thus contributes to pathogenesis of urinary tract infection.

  12. Ouabain Protects Human Renal Cells against the Cytotoxic Effects of Shiga Toxin Type 2 and Subtilase Cytotoxin

    Directory of Open Access Journals (Sweden)

    María M. Amaral

    2017-07-01

    Full Text Available Hemolytic uremic syndrome (HUS is one of the most common causes of acute renal failure in children. The majority of cases are associated with Shiga toxin (Stx-producing Escherichia coli (STEC. In Argentina, HUS is endemic and presents the highest incidence rate in the world. STEC strains expressing Stx type 2 (Stx2 are responsible for the most severe cases of this pathology. Subtilase cytotoxin (SubAB is another STEC virulence factor that may contribute to HUS pathogenesis. To date, neither a licensed vaccine nor effective therapy for HUS is available for humans. Considering that Ouabain (OUA may prevent the apoptosis process, in this study we evaluated if OUA is able to avoid the damage caused by Stx2 and SubAB on human glomerular endothelial cells (HGEC and the human proximal tubule epithelial cell (HK-2 line. HGEC and HK-2 were pretreated with OUA and then incubated with the toxins. OUA protected the HGEC viability from Stx2 and SubAB cytotoxic effects, and also prevented the HK-2 viability from Stx2 effects. The protective action of OUA on HGEC and HK-2 was associated with a decrease in apoptosis and an increase in cell proliferation. Our data provide evidence that OUA could be considered as a therapeutic strategy to avoid the renal damage that precedes HUS.

  13. Structural and Dynamic "Portraits" of Recombinant and Native Cytotoxin I from Naja oxiana: How Close Are They?

    Science.gov (United States)

    Dubovskii, Peter V; Dubinnyi, Maxim A; Konshina, Anastasia G; Kazakova, Ekaterina D; Sorokoumova, Galina M; Ilyasova, Tatyana M; Shulepko, Mikhail A; Chertkova, Rita V; Lyukmanova, Ekaterina N; Dolgikh, Dmitry A; Arseniev, Alexander S; Efremov, Roman G

    2017-08-29

    Today, recombinant proteins are quite widely used in biomedical and biotechnological applications. At the same time, the question about their full equivalence to the native analogues remains unanswered. To gain additional insight into this problem, intimate atomistic details of a relatively simple protein, small and structurally rigid recombinant cardiotoxin I (CTI) from cobra Naja oxiana venom, were characterized using nuclear magnetic resonance (NMR) spectroscopy and atomistic molecular dynamics (MD) simulations in water. Compared to the natural protein, it contains an additional Met residue at the N-terminus. In this work, the NMR-derived spatial structure of uniformly 13 C- and 15 N-labeled CTI and its dynamic behavior were investigated and subjected to comparative analysis with the corresponding data for the native toxin. The differences were found in dihedral angles of only a single residue, adjacent to the N-terminal methionine. Microsecond-long MD traces of the toxins reveal an increased flexibility in the residues spatially close to the N-Met. As the detected structural and dynamic changes of the two CTI models do not result in substantial differences in their cytotoxicities, we assume that the recombinant protein can be used for many purposes as a reasonable surrogate of the native one. In addition, we discuss general features of the spatial organization of cytotoxins, implied by the results of the current combined NMR and MD study.

  14. Portable neon purification system

    Energy Technology Data Exchange (ETDEWEB)

    Richardson, R.A.; Schmitt, R.L.

    1995-08-01

    This paper describes the principle design features of a portable neon purification system and the results of the system performance testing. Neon gas replaces air in the Ring Imaging Cherenkov detector without using vacuum, in experiment E781(SELEX) at Fermilab. The portable neon purification system purifies neon gas by, first purging air with CO{sub 2}, freezing the CO{sub 2}, then cryoadsorbing the remaining contaminants. The freezer removes carbon dioxide from a neon gas mixture down to a maximum concentration of 500 parts-per-million (ppm). The charcoal bed adsorber removes nitrogen from neon gas down to a maximum concentration of 100 ppm. The original RICH vessel was designed to hold vacuum but its photomultiplier tube plates were not.

  15. Electron beam silicon purification

    Energy Technology Data Exchange (ETDEWEB)

    Kravtsov, Anatoly [SIA ' ' KEPP EU' ' , Riga (Latvia); Kravtsov, Alexey [' ' KEPP-service' ' Ltd., Moscow (Russian Federation)

    2014-11-15

    Purification of heavily doped electronic grade silicon by evaporation of N-type impurities with electron beam heating was investigated in process with a batch weight up to 50 kilos. Effective temperature of the melt, an indicative parameter suitable for purification process characterization was calculated and appeared to be stable for different load weight processes. Purified material was successfully approbated in standard CZ processes of three different companies. Each company used its standard process and obtained CZ monocrystals applicable for photovoltaic application. These facts enable process to be successfully scaled up to commercial volumes (150-300 kg) and yield solar grade silicon. (copyright 2014 WILEY-VCH Verlag GmbH and Co. KGaA, Weinheim) (orig.)

  16. Protection against osteoarthritis in experimental animals by nanogold conjugated snake venom protein toxin gold nanoparticle-Naja kaouthia cytotoxin 1.

    Science.gov (United States)

    Gomes, Antony; Saha, Partha Pratim; Bhowmik, Tanmoy; Dasgupta, Anjan Kumar; Dasgupta, Subir Chandra

    2016-12-01

    Increased severity of osteoarthritis (OA) and adverse side effects of its treatment led to the search for alternative therapies. It was previously reported that snake venom protein toxin Naja kaouthia cytotoxin 1 (NKCT1) and gold nanoparticle (GNP) individually have potential against excremental arthritis. In this study, we analyzed the protective activity of GNP conjugated protein toxin NKCT1 (GNP-NKCT1) against experimental OA. Gold nanoparticle conjugation with NKCT1 (GNP-NKCT1) was done and its physiochemical properties were studied. OA was induced in male albino rats by intra-articular injection of bacterial collagenase and treatment was done with NKCT1/GNP-NKCT1/standard drug (indomethacin). Physical parameter (ankle diameter), urinary markers (hydroxyproline, glucosamine, pyridinoline, deoxypyridinoline), serum and synovial membrane pro-inflammatory markers [tumour necrosis factor-alpha (TNF-α), interleukin-1β (IL-1β), IL-17, vascular endothelial growth factor (VEGF)] and matrix metalloproteinase 1 (MMP1) were measured. Joint histopathology and scanning electron microscopy imaging of articular cartilage surface were also done. Physical parameters, urinary markers, serum and synovial membrane pro-inflammatory makers and MMP1 were increased in arthritic rats and significantly restored after GNP-NKCT1/NKCT1 treatment. Joint histopathology and scanning electron microscopy imaging of articular cartilage surface also indicated the protective effect of GNP-NKCT1 against inflammatory response and cartilage degradation in osteoarthritic rats. In this study restoration of the arthritic markers and bone degradation by GNP-NKCT1 treatment indicated the anti-osteoarthritic property of GNP-NKCT1. Further studies need to be done to confirm these findings.

  17. Effects of Escherichia coli subtilase cytotoxin and Shiga toxin 2 on primary cultures of human renal tubular epithelial cells.

    Science.gov (United States)

    Márquez, Laura B; Velázquez, Natalia; Repetto, Horacio A; Paton, Adrienne W; Paton, James C; Ibarra, Cristina; Silberstein, Claudia

    2014-01-01

    Shiga toxin (Stx)-producing Escherichia coli (STEC) cause post-diarrhea Hemolytic Uremic Syndrome (HUS), which is the most common cause of acute renal failure in children in many parts of the world. Several non-O157 STEC strains also produce Subtilase cytotoxin (SubAB) that may contribute to HUS pathogenesis. The aim of the present work was to examine the cytotoxic effects of SubAB on primary cultures of human cortical renal tubular epithelial cells (HRTEC) and compare its effects with those produced by Shiga toxin type 2 (Stx2), in order to evaluate their contribution to renal injury in HUS. For this purpose, cell viability, proliferation rate, and apoptosis were assayed on HRTEC incubated with SubAB and/or Stx2 toxins. SubAB significantly reduced cell viability and cell proliferation rate, as well as stimulating cell apoptosis in HRTEC cultures in a time dependent manner. However, HRTEC cultures were significantly more sensitive to the cytotoxic effects of Stx2 than those produced by SubAB. No synergism was observed when HRTEC were co-incubated with both SubAB and Stx2. When HRTEC were incubated with the inactive SubAA272B toxin, results were similar to those in untreated control cells. Similar stimulation of apoptosis was observed in Vero cells incubated with SubAB or/and Stx2, compared to HRTEC. In conclusion, primary cultures of HRTEC are significantly sensitive to the cytotoxic effects of SubAB, although, in a lesser extent compared to Stx2.

  18. Effects of Escherichia coli subtilase cytotoxin and Shiga toxin 2 on primary cultures of human renal tubular epithelial cells.

    Directory of Open Access Journals (Sweden)

    Laura B Márquez

    Full Text Available Shiga toxin (Stx-producing Escherichia coli (STEC cause post-diarrhea Hemolytic Uremic Syndrome (HUS, which is the most common cause of acute renal failure in children in many parts of the world. Several non-O157 STEC strains also produce Subtilase cytotoxin (SubAB that may contribute to HUS pathogenesis. The aim of the present work was to examine the cytotoxic effects of SubAB on primary cultures of human cortical renal tubular epithelial cells (HRTEC and compare its effects with those produced by Shiga toxin type 2 (Stx2, in order to evaluate their contribution to renal injury in HUS. For this purpose, cell viability, proliferation rate, and apoptosis were assayed on HRTEC incubated with SubAB and/or Stx2 toxins. SubAB significantly reduced cell viability and cell proliferation rate, as well as stimulating cell apoptosis in HRTEC cultures in a time dependent manner. However, HRTEC cultures were significantly more sensitive to the cytotoxic effects of Stx2 than those produced by SubAB. No synergism was observed when HRTEC were co-incubated with both SubAB and Stx2. When HRTEC were incubated with the inactive SubAA272B toxin, results were similar to those in untreated control cells. Similar stimulation of apoptosis was observed in Vero cells incubated with SubAB or/and Stx2, compared to HRTEC. In conclusion, primary cultures of HRTEC are significantly sensitive to the cytotoxic effects of SubAB, although, in a lesser extent compared to Stx2.

  19. Variations in Helicobacter pylori cytotoxin-associated genes and their influence in progression to gastric cancer: implications for prevention.

    Directory of Open Access Journals (Sweden)

    Cosmeri Rizzato

    Full Text Available Helicobacter pylori (HP is a bacterium that colonizes the human stomach and can establish a long-term infection of the gastric mucosa. Persistent Hp infection often induces gastritis and is associated with the development of peptic ulcer disease, atrophic gastritis, and gastric adenocarcinoma. Virulent HP isolates harbor the cag (cytotoxin-associated genes pathogenicity island (cagPAI, a 40 kb stretch of DNA that encodes components of a type IV secretion system (T4SS. This T4SS forms a pilus for the injection of virulence factors into host target cells, such as the CagA oncoprotein. We analyzed the genetic variability in cagA and other selected genes of the HP cagPAI (cagC, cagE, cagL, cagT, cagV and cag Gamma using DNA extracted from frozen gastric biopsies or from clinical isolates. Study subjects were 95 cagA+ patients that were histologically diagnosed with chronic gastritis or gastric cancer in Venezuela and Mexico, areas with high prevalence of Hp infection. Sequencing reactions were carried out by both Sanger and next-generation pyrosequencing (454 Roche methods. We found a total of 381 variants with unambiguous calls observed in at least 10% of the originally tested samples and reference strains. We compared the frequencies of these genetic variants between gastric cancer and chronic gastritis cases. Twenty-six SNPs (11 non-synonymous and 14 synonymous showed statistically significant differences (P<0.05, and two SNPs, in position 1039 and 1041 of cagE, showed a highly significant association with cancer (p-value = 2.07×10⁻⁶, and the variant codon was located in the VirB3 homology domain of Agrobacterium. The results of this study may provide preliminary information to target antibiotic treatment to high-risk individuals, if effects of these variants are confirmed in further investigations.

  20. Protection against osteoarthritis in experimental animals by nanogold conjugated snake venom protein toxin gold nanoparticle-Naja kaouthia cytotoxin 1

    Science.gov (United States)

    Gomes, Antony; Saha, Partha Pratim; Bhowmik, Tanmoy; Dasgupta, Anjan Kumar; Dasgupta, Subir Chandra

    2016-01-01

    Background & objectives: Increased severity of osteoarthritis (OA) and adverse side effects of its treatment led to the search for alternative therapies. It was previously reported that snake venom protein toxin Naja kaouthia cytotoxin 1 (NKCT1) and gold nanoparticle (GNP) individually have potential against excremental arthritis. In this study, we analyzed the protective activity of GNP conjugated protein toxin NKCT1 (GNP-NKCT1) against experimental OA. Methods: Gold nanoparticle conjugation with NKCT1 (GNP-NKCT1) was done and its physiochemical properties were studied. OA was induced in male albino rats by intra-articular injection of bacterial collagenase and treatment was done with NKCT1/GNP-NKCT1/standard drug (indomethacin). Physical parameter (ankle diameter), urinary markers (hydroxyproline, glucosamine, pyridinoline, deoxypyridinoline), serum and synovial membrane pro-inflammatory markers [tumour necrosis factor-alpha (TNF-α), interleukin-1β (IL-1β), IL-17, vascular endothelial growth factor (VEGF)] and matrix metalloproteinase 1 (MMP1) were measured. Joint histopathology and scanning electron microscopy imaging of articular cartilage surface were also done. Results: Physical parameters, urinary markers, serum and synovial membrane pro-inflammatory makers and MMP1 were increased in arthritic rats and significantly restored after GNP-NKCT1/NKCT1 treatment. Joint histopathology and scanning electron microscopy imaging of articular cartilage surface also indicated the protective effect of GNP-NKCT1 against inflammatory response and cartilage degradation in osteoarthritic rats. Interpretation & conclusions: In this study restoration of the arthritic markers and bone degradation by GNP-NKCT1 treatment indicated the anti-osteoarthritic property of GNP-NKCT1. Further studies need to be done to confirm these findings. PMID:28474628

  1. THE PURIFICATION OF HYPERTENSIN II

    Science.gov (United States)

    Skeggs, Leonard T.; Kahn, Joseph R.; Shumway, Norman P.

    1956-01-01

    The enzymatic conversion of hypertensin I to hypertensin II is described together with the subsequent purification of the product by means of counter-current distribution. Improved methods are also presented for the preparation of renin and its substrate, as well as in methods for the reaction of these materials and the purification of the resulting hypertensin I. PMID:13295488

  2. PURIFICATION AND CHARACTERISATION OF ALKALINE ...

    African Journals Online (AJOL)

    Prof. Adipala Ekwamu

    osmotic shock, indicating that the enzyme is located either in the periplasmic space or is loosely bound to the cell wall. Initially, a DEAE column was used leading to 28% yield and 77 times fold purification, followed by. Sephacryl gel filtration column giving 25% yield and 72 times fold purification; indicating loss of enzyme in ...

  3. Air/Water Purification

    Science.gov (United States)

    1992-01-01

    After 18 years of research into air/water pollution at Stennis Space Center, Dr. B. C. Wolverton formed his own company, Wolverton Environmental Services, Inc., to provide technology and consultation in air and water treatment. Common houseplants are used to absorb potentially harmful materials from bathrooms and kitchens. The plants are fertilized, air is purified, and wastewater is converted to clean water. More than 100 U.S. communities have adopted Wolverton's earlier water hyacinth and artificial marsh applications. Catfish farmers are currently evaluating the artificial marsh technology as a purification system.

  4. Water Purification Product

    Science.gov (United States)

    2004-01-01

    Ecomaster, an affiliate of BioServe Space Technologies, this PentaPure technology has been used to purify water for our nation's Space Shuttle missions since 1981. WTC-Ecomaster of Mirneapolis, Minnesota manufactures water purification systems under the brand name PentaPure (TM). BioServe researcher Dr. George Marchin, of Kansas State University, first demonstrated the superiority of this technology and licensed it to WTC. Marchin continues to perform microgravity research in the development of new technologies for the benefit of life on Earth.

  5. Protein Characterization of Extracellular Microvesicles/Exosomes Released from Cytotoxin-Challenged Rat Cerebrocortical Mixed Culture and Mouse N2a Cells.

    Science.gov (United States)

    Kumar, Dhwani; Manek, Rachna; Raghavan, Vijaya; Wang, Kevin K

    2017-03-10

    A number of neuronal and glial proteins were previously found to be released in free-standing soluble form from cultured brain cells into cell-conditioned media. Here, we sought to examine if similar proteins are also contained in neural and astroglial cell-released extracellular microvesicles/exosomes (MV/E). In this study, MV/E were isolated from cell-conditioned media from control and cytotoxin-challenged rat cerebrocortical mixed culture (CTX) and mouse neuroblastoma N2a cells. Cytotoxin challenges included pro-necrosis calcium ionophore A23187, pro-apoptosis staurosporine (STS), and excitotoxin N-methyl-D-aspartate. Based on established nanoparticle characterization method (dynamic light scattering, NanoTracker, and transmission electron microscopy), we confirmed that these released vesicles are in fact characteristic representation of MV/E by morphology (lipid bilayered vesicles) and by particle size (132-142 nm for CTX and 49-77 nm for N2a cells). We indeed identified neural cell body protein UCH-L1, axonal injury marker αII-spectrin and its breakdown products (SBDPs), astroglial markers GFAP and its breakdown products (GFAP-BDP), dendritic protein BIII-tubulin, synaptic protein synaptophysin, and exosome marker Alix in microvesicles from CTX and/or N2a cells. Furthermore, SBDPs, GFAP-BDP, UCH-L1, and synaptophysin are especially dominant in MV/E isolated from cytotoxin-treated CTX cells. Similarly, SBDPs, βIII-tubulin, and UCH-L1 are more prominently observed in cytotoxin-challenged N2a cells. Lastly, when isolated MV/E from A23187- or STS-challenged N2a cells were introduced to healthy N2a culture, they are capable of evoking cytotoxicity in the latter. Taken together, our study identified that microvesicles/exosomes isolated form healthy and injured brain cells contain certain neural and astroglial proteins, as well as possibly other cytotoxic factors that are capable of propagating cytotoxic effects.

  6. Treatment history and antimicrobial susceptibility results for Mannheimia haemolytica, Pasteurella multocida, and Histophilus somni isolates from bovine respiratory disease cases submitted to the Iowa State University Veterinary Diagnostic Laboratory from 2013 to 2015.

    Science.gov (United States)

    Magstadt, Drew R; Schuler, Adlai M; Coetzee, Johann F; Krull, Adam C; O'Connor, Annette M; Cooper, Vickie L; Engelken, Terry J

    2017-10-01

    Bovine respiratory disease is the most costly disease facing the cattle industry. Increasing resistance to antimicrobial treatment has been presented as a significant contributing factor, often through summarized susceptibility testing data. We assessed the relationship between previous antimicrobial treatment and antimicrobial susceptibility results from isolates of Mannheimia haemolytica, Pasteurella multocida, and Histophilus somni cultured from bovine respiratory cases submitted to the Iowa State University Veterinary Diagnostic Laboratory from 2013 to 2015. Antimicrobial susceptibility data from 1,251 bacterial isolates were included for analysis. More bacterial isolates from cattle that received antimicrobial treatment showed resistance compared to isolates from untreated cattle, and the percentage of resistant isolates increased as the number of antimicrobial treatments increased. Resistance to enrofloxacin, spectinomycin, tilmicosin, and tulathromycin was present in >75% of M. haemolytica isolates from cattle that had received 3 or more antimicrobial treatments; resistance to each of those 4 antimicrobials was present in ≤10% of M. haemolytica isolates from untreated cattle. Similar but less dramatic trends were apparent for isolates of P. multocida and H. somni. The percentage of multi-drug resistant bacterial isolates also increased with the number of treatments. Results of our study suggest that previous antimicrobial treatment may have a profound effect on antimicrobial susceptibility testing. Summarized susceptibility results from diagnostic laboratories should not be used to make generalized statements regarding trends in antimicrobial resistance without providing context regarding antimicrobial treatment history.

  7. Effect of charcoal on water purification

    OpenAIRE

    Suzuki, Hirotaka; Kawahigashi, Tatsuo

    2014-01-01

    [Abstract] A natural basin system purifies water through self-purification, but the water pollution load of a river might exceed its self-purification capacity. Charcoal, which is used for other uses aside from heating, such as air purification, was evaluated experimentally for water quality purification. The experiment described herein is based on simple water quality measurements. Some experimentally obtained results are discussed.

  8. Recovery and purification of ethylene

    Science.gov (United States)

    Reyneke, Rian [Katy, TX; Foral, Michael J [Aurora, IL; Lee, Guang-Chung [Houston, TX; Eng, Wayne W. Y. [League City, TX; Sinclair, Iain [Warrington, GB; Lodgson, Jeffery S [Naperville, IL

    2008-10-21

    A process for the recovery and purification of ethylene and optionally propylene from a stream containing lighter and heavier components that employs an ethylene distributor column and a partially thermally coupled distributed distillation system.

  9. Nanomechanical Water Purification Device Project

    Data.gov (United States)

    National Aeronautics and Space Administration — Seldon Laboratories, LLC, proposes a lightweight, low-pressure water purification device that harnesses the unique properties of carbon nanotubes and will operate...

  10. Bivalent vaccination against pneumonic pasteurellosis in domestic sheep and goats with modified-live in-frame lktA deletion mutants of Mannheimia haemolytica.

    Science.gov (United States)

    Briggs, Robert E; Hauglund, Melissa J; Maheswaran, Samuel K; Tatum, Fred M

    2013-11-01

    A temperature-sensitive shuttle vector, pBB80C, was utilized to generate in-frame deletion mutants of the leukotoxin structural gene (lktA) of Mannheimia haemolytica serotypes 1, 2, 5, 6, 7, 8, 9, and 12. Culture supernatants from the mutants contained a truncated protein with an approximate molecular weight of 66 kDa which was reactive to anti-leukotoxin monoclonal antibody. No protein reactive to anti-LktA monoclonal antibody was detected at the molecular weight 100-105 kDa of native LktA. Sheep and goats vaccinated intramuscularly with a mixture of serotypes 5 and 6 mutants were resistant to virulent challenge with a mixture of the wild-type parent strains. These vaccinates responded serologically to both vaccine serotypes and exhibited markedly-reduced lung lesion volume and pulmonary infectious load compared to control animals. Control animals yielded a mixture of serotypes from lung lobes, but the proportion even within an individual animal varied widely from 95% serotype 5-95% serotype 6. Cultures recovered from liver were homogeneous, but two animals yielded serotype 5 and the other two yielded serotype 6 in pure culture. Published by Elsevier Ltd.

  11. Water Purification Systems

    Science.gov (United States)

    1994-01-01

    Clearwater Pool Technologies employs NASA-developed silver/copper ionization to purify turtle and dolphin tanks, cooling towers, spas, water recycling systems, etc. The pool purifier consists of a microcomputer to monitor water conditions, a pair of metallic electrodes, and a rheostat controller. Ions are generated by passing a low voltage current through the electrodes; the silver ions kill the bacteria, and the copper ions kill algae. This technology has found broad application because it offers an alternative to chemical disinfectants. It was originally developed to purify water on Apollo spacecraft. Caribbean Clear has been using NASA's silver ionization technology for water purification for more than a decade. Two new products incorporate advancements of the basic technology. One is the AquaKing, a system designed for areas with no source of acceptable drinking water. Another is the Caribbean Clear Controller, designed for commercial pool and water park applications where sanitizing is combined with feedback control of pH and an oxidizer, chlorine or bromine. The technology was originally developed to purify water on Apollo spacecraft.

  12. Rapid purification of recombinant histones.

    Directory of Open Access Journals (Sweden)

    Henrike Klinker

    Full Text Available The development of methods to assemble nucleosomes from recombinant histones decades ago has transformed chromatin research. Nevertheless, nucleosome reconstitution remains time consuming to this day, not least because the four individual histones must be purified first. Here, we present a streamlined purification protocol of recombinant histones from bacteria. We termed this method "rapid histone purification" (RHP as it circumvents isolation of inclusion bodies and thereby cuts out the most time-consuming step of traditional purification protocols. Instead of inclusion body isolation, whole cell extracts are prepared under strongly denaturing conditions that directly solubilize inclusion bodies. By ion exchange chromatography, the histones are purified from the extracts. The protocol has been successfully applied to all four canonical Drosophila and human histones. RHP histones and histones that were purified from isolated inclusion bodies had similar purities. The different purification strategies also did not impact the quality of octamers reconstituted from these histones. We expect that the RHP protocol can be readily applied to the purification of canonical histones from other species as well as the numerous histone variants.

  13. RELIGION AND PURIFICATION OF SOUL

    Directory of Open Access Journals (Sweden)

    Azam Khodashenas Pelko

    2010-11-01

    Full Text Available The Jainism emphasizes three major teachings about the purification of the soul (jiva, Ahimsa, Aparigrapha and anekantwad. Jainism, The focus of this religion has been purification of the soul by means of right conduct, right faith and right knowledge. The ultimate goal of Hinduism is Moksha or liberation (total freedom. In Hinduism, purification of the soul is a goal that one must work to attain. The Buddhism is the science of pursuing the aim of making the human mind perfect, and of purifying the human soul. The knowledge of purifying of the soul and softening of the hearts is as essential for human. They having the correct motivations means purifying our souls from hypocrisy, caprice, and heedlessness. The primary goal of Taoism may be described as the mystical intuition of the Tao, which is the way, the undivided unity, and the ultimate Reality. According to the Christianity access to truth cannot be conceived without purity of the soul

  14. Purification of Water by Aquatic Plants

    OpenAIRE

    Morimitsu, Katsuhito; Kawahigashi, Tatsuo

    2013-01-01

    [Abstract] Water quality purification of many water systems including those occurring in rivers depends to a great degree on water quality purification activities of aquatic plants and microbes. This paper presents a discussion of results, based on laboratory experiments, of purification by aquatic plants.

  15. Estudo clínico e laboratorial da pneumonia de bezerros induzida pela inoculação intrabronquial de Mannheimia haemolytica

    Directory of Open Access Journals (Sweden)

    Fagliari J.J.

    2003-01-01

    Full Text Available Foram utilizados 14 bezerros da raça Holandesa entre 15 e 30 dias de idade, sete inoculados com 5ml de solução salina fosfato-tamponada Dulbecco (DPBS e sete inoculados com 5x10(9 UFC de Mannheimia haemolytica/5 ml de DPBS, via intrabronquial. O exame clínico e as colheitas de sangue e de lavado broncoalveolar foram realizados antes e 4, 6, 12 e 24 horas após a inoculação. O exame histopatológico dos pulmões revelou lesões de pneumonia fibrinosa hemorrágica. Os sinais clínicos surgiram quatro horas após a inoculação das bactérias. Os sete bezerros enfermos apresentavam apatia, respiração abdominal, alteração dos ruídos respiratórios, hipertermia, elevação das freqüências cardíaca e respiratória e arritmia cardíaca. Os exames de laboratório mostraram aumento na contagem de hemácias e do volume globular, indicando hemoconcentração. A diminuição de PO2 e a elevação de PCO2 sugerem inadequada ventilação pulmonar. Observaram-se neutrofilia e alto número de neutrófilos e monócitos no fluido broncoalveolar. O pH do sangue e as concentrações plasmáticas de HCO3, Na e K não apresentaram alterações significativas.

  16. Purification and properties of dialkylfluorophosphatase

    NARCIS (Netherlands)

    Cohen, J.A.; Warringa, M.G.P.J.

    1957-01-01

    1. 1. Zone electrophoresis on starch columns of purified preparations of fluorophosphatase resulted in a further purification. The preparations thus obtained differed in various respects from the cruder ones so far described. 2. 2. In the course of this electrophoresis fractions were obtained,

  17. Bioinspired Materials for Water Purification

    Directory of Open Access Journals (Sweden)

    Alfredo Gonzalez-Perez

    2016-06-01

    Full Text Available Water scarcity issues associated with inadequate access to clean water and sanitation is a ubiquitous problem occurring globally. Addressing future challenges will require a combination of new technological development in water purification and environmental remediation technology with suitable conservation policies. In this scenario, new bioinspired materials will play a pivotal role in the development of more efficient and environmentally friendly solutions. The role of amphiphilic self-assembly on the fabrication of new biomimetic membranes for membrane separation like reverse osmosis is emphasized. Mesoporous support materials for semiconductor growth in the photocatalytic degradation of pollutants and new carriers for immobilization of bacteria in bioreactors are used in the removal and processing of different kind of water pollutants like heavy metals. Obstacles to improve and optimize the fabrication as well as a better understanding of their performance in small-scale and pilot purification systems need to be addressed. However, it is expected that these new biomimetic materials will find their way into the current water purification technologies to improve their purification/removal performance in a cost-effective and environmentally friendly way.

  18. Statistical and Judgmental Criteria for Scale Purification

    DEFF Research Database (Denmark)

    Wieland, Andreas; Durach, Christian F.; Kembro, Joakim

    2017-01-01

    of scale purification, to critically analyze the current state of scale purification in supply chain management (SCM) research and to provide suggestions for advancing the scale-purification process. Design/methodology/approach A framework for making scale-purification decisions is developed and used...... of methodological rigor and coherence is identified when it comes to current purification practices in empirical SCM research. Suggestions for methodological improvements are provided. Research limitations/implications The framework and additional suggestions will help to advance the knowledge about scale...... to analyze and critically reflect on the application of scale purification in leading SCM journals. Findings This research highlights the need for rigorous scale-purification decisions based on both statistical and judgmental criteria. By applying the proposed framework to the SCM discipline, a lack...

  19. Sodium purification in Rapsodie; La purification du sodium a Rapsodie

    Energy Technology Data Exchange (ETDEWEB)

    Giraud, B. [Commissariat a l' Energie Atomique, Dir. des Piles Atomiques, Cadarache (France). Centre d' Etudes Nucleaires

    1968-07-01

    This report is one of a series of publications presenting the main results of tests carried out during the start-up of the first french fast neutron reactor: Rapsodie. The article presents the sodium purification techniques used in the reactor cooling circuits both from the constructional point of view and with respect to results obtained during the first years working. (author) [French] Ce rapport fait partie d'une serie de publications presentant l'essentiel des resultats des essais effectues a l'occasion du demarrage du premier reacteur francais a neutrons rapides: RAPSODIE. Cet article expose les techniques de la purification du sodium utilise dans les circuits de refroidissement du reacteur tant au point de vue de leur realisation technologique, que des resultats obtenus pendant la premiere annee de fonctionnement. (auteur)

  20. Water purification using organic salts

    Science.gov (United States)

    Currier, Robert P.

    2004-11-23

    Water purification using organic salts. Feed water is mixed with at least one organic salt at a temperature sufficiently low to form organic salt hydrate crystals and brine. The crystals are separated from the brine, rinsed, and melted to form an aqueous solution of organic salt. Some of the water is removed from the aqueous organic salt solution. The purified water is collected, and the remaining more concentrated aqueous organic salt solution is reused.

  1. SNO+ Scintillator Purification and Assay

    Science.gov (United States)

    Ford, R.; Chen, M.; Chkvorets, O.; Hallman, D.; Vázquez-Jáuregui, E.

    2011-04-01

    We describe the R&D on the scintillator purification and assay methods and technology for the SNO+ neutrino and double-beta decay experiment. The SNO+ experiment is a replacement of the SNO heavy water with liquid scintillator comprised of 2 g/L PPO in linear alkylbenzene (LAB). During filling the LAB will be transported underground by rail car and purified by multi-stage distillation and steam stripping at a flow rate of 19 LPM. While the detector is operational the scintillator can be recirculated at 150 LPM (full detector volume in 4 days) to provide repurification as necessary by either water extraction (for Ra, K, Bi) or by functional metal scavenger columns (for Pb, Ra, Bi, Ac, Th) followed by steam stripping to remove noble gases and oxygen (Rn, O2, Kr, Ar). The metal scavenger columns also provide a method for scintillator assay for ex-situ measurement of the U and Th chain radioactivity. We have developed "natural" radioactive spikes of Pb and Ra in LAB and use these for purification testing. Lastly, we present the planned operating modes and purification strategies and the plant specifications and design.

  2. Technological assumptions for biogas purification.

    Science.gov (United States)

    Makareviciene, Violeta; Sendzikiene, Egle

    2015-01-01

    Biogas can be used in the engines of transport vehicles and blended into natural gas networks, but it also requires the removal of carbon dioxide, hydrogen sulphide, and moisture. Biogas purification process flow diagrams have been developed for a process enabling the use of a dolomite suspension, as well as for solutions obtained by the filtration of the suspension, to obtain biogas free of hydrogen sulphide and with a carbon dioxide content that does not exceed 2%. The cost of biogas purification was evaluated on the basis of data on biogas production capacity and biogas production cost obtained from local water treatment facilities. It has been found that, with the use of dolomite suspension, the cost of biogas purification is approximately six times lower than that in the case of using a chemical sorbent such as monoethanolamine. The results showed travelling costs using biogas purified by dolomite suspension are nearly 1.5 time lower than travelling costs using gasoline and slightly lower than travelling costs using mineral diesel fuel.

  3. The impact of meteorology on the occurrence of waterborne outbreaks of vero cytotoxin-producing Escherichia coli (VTEC): a logistic regression approach.

    Science.gov (United States)

    O'Dwyer, Jean; Morris Downes, Margaret; Adley, Catherine C

    2016-02-01

    This study analyses the relationship between meteorological phenomena and outbreaks of waterborne-transmitted vero cytotoxin-producing Escherichia coli (VTEC) in the Republic of Ireland over an 8-year period (2005-2012). Data pertaining to the notification of waterborne VTEC outbreaks were extracted from the Computerised Infectious Disease Reporting system, which is administered through the national Health Protection Surveillance Centre as part of the Health Service Executive. Rainfall and temperature data were obtained from the national meteorological office and categorised as cumulative rainfall, heavy rainfall events in the previous 7 days, and mean temperature. Regression analysis was performed using logistic regression (LR) analysis. The LR model was significant (p < 0.001), with all independent variables: cumulative rainfall, heavy rainfall and mean temperature making a statistically significant contribution to the model. The study has found that rainfall, particularly heavy rainfall in the preceding 7 days of an outbreak, is a strong statistical indicator of a waterborne outbreak and that temperature also impacts waterborne VTEC outbreak occurrence.

  4. The diagnosis of Clostridium difficile-associated diarrhea: comparison of Triage C. difficile panel, EIA for Tox A/B and cytotoxin assays.

    Science.gov (United States)

    Alfa, M J; Swan, B; VanDekerkhove, B; Pang, P; Harding, G K M

    2002-08-01

    This study prospectively compared; Triage(R) C. difficile test (TCT), TechLab C. difficile toxin A-B enzyme immuno-assay (EIA), and cell-culture cytotoxin test (CT). Of the 400 stools tested, 99 were positive by any test with 92, 41 and 58 detected by TCT, EIA and CT, respectively. Culture of discordant samples indicated that 52 contained C. difficile (42 toxigenic, 10 non-toxigenic), 10 contained Clostridium species and 2 had no detectable clostridium isolates. There were 21/42 toxigenic C. difficile isolates from 17 patients whose stools were negative when originally tested by CT. Review of available patient charts indicated that 12/14 did not previously or currently have C. difficile associated diarrhea, whereas 2 patients developed disease within a few days. Compared to CT as the gold standard, the sensitivity and specificity were; 93%, 89% and 66%, 99% for TCT and EIA respectively. The 8 stool samples with Toxin A(-) Toxin B(+) isolates were detected in 8, 4, and 6 samples by TCT, EIA and CT, respectively. In summary, TCT as a screening test allowed reliable reporting for 85% of stools on the day of receipt. For the 15% of stools requiring further testing we recommend the use of CT.

  5. Effects of central administration of oxytocin-saporin cytotoxin on chronic inflammation and feeding/drinking behaviors in adjuvant arthritic rats.

    Science.gov (United States)

    Matsuura, Takanori; Kawasaki, Makoto; Hashimoto, Hirofumi; Yoshimura, Mitsuhiro; Motojima, Yasuhito; Saito, Reiko; Ueno, Hiromichi; Maruyama, Takashi; Sabanai, Ken; Mori, Toshiharu; Ohnishi, Hideo; Sakai, Akinori; Ueta, Yoichi

    2016-05-16

    An increase in the arthritis index as a marker of chronic inflammation and suppression of food intake are observed in adjuvant arthritic (AA) rats. Our previous study demonstrated that central oxytocin (OXT)-ergic pathways were activated potently in AA rats. In the present study, OXT-saporin (SAP) cytotoxin, which chemically disrupts OXT signaling was administered centrally to determine whether central OXT may be involved in the developments of chronic inflammation and alteration of feeding/drinking behavior in AA rats. The arthritis index was significantly enhanced in AA rats pretreated with OXT-SAP administered intrathecally (i.t.) but not intracerebroventricularly (i.c.v.). Suppression of food intake was significantly attenuated transiently in AA rats pretreated with OXT-SAP administered i.c.v. but not i.t. Suppression of drinking behavior was not affected by i.t. or i.c.v. administration of OXT-SAP in AA rats. In addition, intraperitoneal administration of an OXT receptor antagonist did not change the arthritis index or feeding/drinking behavior in AA rats. These results suggest that central OXT-ergic pathways may be involved in anti-inflammation at the spinal level and suppression of feeding behavior at the forebrain-brainstem level in AA rats. Copyright © 2016 Elsevier Ireland Ltd. All rights reserved.

  6. Nanostructured Catalytic Reactors for Air Purification Project

    Data.gov (United States)

    National Aeronautics and Space Administration — This SBIR Phase I project proposes the development of lightweight compact nanostructured catalytic reactors for air purification from toxic gaseous organic...

  7. Nanostructured Catalytic Reactors for Air Purification Project

    Data.gov (United States)

    National Aeronautics and Space Administration — This SBIR Phase II project proposes the development of lightweight compact nanostructured catalytic reactors for air purification from toxic gaseous organic...

  8. Comparing Russian and Finnish standards of water purification

    OpenAIRE

    Maria, Pupkova

    2012-01-01

    The subject of this thesis is water purification. The first aim of this thesis is to consider different ways of water purification. The second aim is to compare Finnish and Russian standards of water purification. The third one is to show water purification methods on the pattern of Mikkeli water purification plan. Water purification methods of water intended for human consumption will be described.Combined tables will be done according to the quality requirement of drinking water of both,...

  9. Purification of rhamnolipid using colloidal magnetic nanoparticles ...

    African Journals Online (AJOL)

    Phospholipid-coated colloidal magnetic nanoparticles with mean magnetite core size of 9 nm are shown to be effective ion exchange media for the recovery and purification of Rhaminolipid from culture mixtures. These particles have high adsorption capacity for purification (an order of magnitude larger than the best ...

  10. Water purification in low background experiments

    Science.gov (United States)

    Giammarchi, Marco

    2017-10-01

    Water purification is an important technique in high-mass low radioactivity experiments in modern physics. Water is frequently used both as a shielding and as the sensitive part of a particle detector in underground arrangements, especially in the frame of Astroparticle Physics studies. In this paper, I will describe the main purification techniques and discuss some of its performances.

  11. HOUSEHOLD PURIFICATION OF FLUORIDE CONTAMINATED MAGADI (TRONA)

    DEFF Research Database (Denmark)

    Nielsen, Joan Maj; Dahi, Elian

    1997-01-01

    Purification of fluoride contaminated magadi is studied using bone char sorption and calcium precipitation. The bone char treatment is found to be workable both in columns and in batches where the magadi is dissolved in water prior to treatment. The concentrations in the solutions were 89 g magadi...... treatment method. A procedure for purification of fluoride contaminated magadi at household level is described....

  12. Purification and characterization of amidase from acrylamide ...

    African Journals Online (AJOL)

    An amidase from a newly isolated acrylamide-degrading bacterium Burkholderia sp. strain DR.Y27 was purified to homogeneity by a combination of anion exchange and gel filtration chromatography. The purification strategy achieved 11.15 of purification fold and a yield of 1.55%. The purified amidase consisted of four ...

  13. THE PURIFICATION OF HYPERTENSIN I

    Science.gov (United States)

    Skeggs, Leonard T.; Marsh, Walton H.; Kahn, Joseph R.; Shumway, Norman P.

    1954-01-01

    The purification of hypertensin I has been described. The final product which is four times as powerful a pressor agent as l-arterenol, is obtained with an over-all recovery of 40 per cent. The product consists of a single component in countercurrent distribution, having a nitrogen content of 15.97 per cent and a specific activity of 7050 Goldblatt units per mg. of N or 1125 units per mg. of solid. Acid hydrolysis and paper chromatography indicate in a preliminary fashion that there are about nine amino acids present in the intact polypeptide. PMID:13201713

  14. The fundamentals of RNA purification.

    Science.gov (United States)

    Nilsen, Timothy W

    2013-07-01

    The ability to purify, analyze, and manipulate RNA is now essential for many laboratories working in the life sciences; however, the skills and practices required to work with RNA are not present in every laboratory, and initiating RNA research can be intimidating. In this article, we provide an overview of RNA purification procedures and discuss strategies to prevent RNA degradation, so that any competent researcher can confidently purify RNA and use it to perform meaningful experiments from the most basic to the highly sophisticated.

  15. Comparison of the primary structures, cytotoxicities, and affinities to phospholipids of five kinds of cytotoxins from the venom of Indian cobra, Naja naja.

    Science.gov (United States)

    Suzuki-Matsubara, Mieko; Athauda, Senarath B P; Suzuki, Yoshiyuki; Matsubara, Kazumi; Moriyama, Akihiko

    2016-01-01

    The molecular mechanism underlying the hemolytic and cytolytic processes of cobra cytotoxins (CTXs) is not yet fully elucidated. To examine this, we analyzed the amino acid sequences, hemolytic and cytotoxic activities, and affinities to phospholipids of the five major CTXs purified from the venom of Indian cobra, Naja naja. CTX2, CTX7, and CTX8 belonged to S-type, and CTX9 and CTX10 to P-type. Comparisons of CTX7 with CTX8 and CTX9 with CTX10 revealed similar primary structures and hemolytic and cytolytic activities. CTX2, whose primary structure was rather different from the others, showed several times weaker hemolytic and cytolytic biological activities than the others. The comparison of CTX2 with CTX7 suggested the importance of Lys30 in loop II for the strong hemolytic and cytolytic activities of S-type CTXs. Cloning of 12 CTX cDNAs from the Naja naja venom cDNA library revealed that 18 out of 23 substitutions found in CTX cDNAs were nonsynonymous. This clearly indicated the accelerated evolution of CTX genes. Multiple sequence alignment of 51 kinds of CTX cDNAs and calculations of nonsynonymous and synonymous substitutions indicated that the codons coding the three loops' regions, which may interact with the hydrophobic tails of phospholipids, have undergone an accelerated evolution. In contrast, the codons coding for amino acid residues considered to participate in the recognition and binding of the hydrophilic head groups of phospholipids, eight Cys residues, and those likely stabilizing β core structure, were all conserved. Copyright © 2015 Elsevier Inc. All rights reserved.

  16. Reevaluation of the Premier Clostridium difficile toxin A and B immunoassay with comparison to glutamate dehydrogenase common antigen testing evaluating Bartels cytotoxin and Prodesse ProGastro Cd polymerase chain reaction as confirmatory procedures.

    Science.gov (United States)

    Doing, Kirk M; Hintz, Marilyn S; Keefe, Calvin; Horne, Sarah; LeVasseur, Shelby; Kulikowski, Martha L

    2010-02-01

    Enzyme immunoassays are currently the most common tests used in the clinical laboratory for the detection of Clostridium difficile toxins; however, significant problems with their performance have recently been described. We prospectively reevaluated the Meridian Premier C. difficile toxin A/B assay with direct comparison to a 2-step algorithm that screened for C. difficile common antigen and compared cytotoxin and real-time polymerase chain reaction (PCR) as confirmatory procedures. The Premier assay lacked sufficient sensitivity, missing 25% of true-positive samples. PCR was the most sensitive method and the only procedure that allowed same day testing and reporting. 2010 Elsevier Inc. All rights reserved.

  17. Semiconductor grade, solar silicon purification project

    Science.gov (United States)

    Ingle, W. M.; Rosler, R. R.; Thompson, S. W.; Chaney, R. E.

    1979-01-01

    Experimental apparatus and procedures used in the development of a 3-step SiF2(x) polymer transport purification process are described. Both S.S.M.S. and E.S. analysis demonstrated that major purification had occured and some samples were indistinguishable from semiconductor grade silicon (except possibly for phosphorus). Recent electrical analysis via crystal growth reveals that the product contains compensated phosphorus and boron. The low projected product cost and short energy payback time suggest that the economics of this process will result in a cost less than the goal of $10/Kg(1975 dollars). The process appears to be readily scalable to a major silicon purification facility.

  18. Protein purification by affinity precipitation.

    Science.gov (United States)

    Hilbrig, Frank; Freitag, Ruth

    2003-06-25

    Developing the most efficient strategy for the purification of a (recombinant) protein especially at large scale remains a challenge. A typical problem of the downstream process of mammalian cell products is, for instance, the early capture of the highly diluted product from the complex process stream. Affinity precipitation has been suggested in this context. The technique is known for over 20 years, but has recently received more attention due to the development of new materials for its implementation, but also because it seems ideally suited to specific product capture at large scale. The present review gives a comprehensive overview over this technique. Besides an introduction to the basic principle and a brief summary of the historical development, the main focus is on the current state-of-art of the technique, the available materials, important recent applications, as well as process design strategies and operating procedures. Special consideration is given to affinity precipitation for product recovery at large scale.

  19. Reverse osmosis water purification system

    Science.gov (United States)

    Ahlstrom, H. G.; Hames, P. S.; Menninger, F. J.

    1986-01-01

    A reverse osmosis water purification system, which uses a programmable controller (PC) as the control system, was designed and built to maintain the cleanliness and level of water for various systems of a 64-m antenna. The installation operates with other equipment of the antenna at the Goldstone Deep Space Communication Complex. The reverse osmosis system was designed to be fully automatic; with the PC, many complex sequential and timed logic networks were easily implemented and are modified. The PC monitors water levels, pressures, flows, control panel requests, and set points on analog meters; with this information various processes are initiated, monitored, modified, halted, or eliminated as required by the equipment being supplied pure water.

  20. Phase I and pharmacokinetic study of irofulven, a novel mushroom-derived cytotoxin, administered for five consecutive days every four weeks in patients with advanced solid malignancies.

    Science.gov (United States)

    Eckhardt, S G; Baker, S D; Britten, C D; Hidalgo, M; Siu, L; Hammond, L A; Villalona-Calero, M A; Felton, S; Drengler, R; Kuhn, J G; Clark, G M; Smith, S L; MacDonald, J R; Smith, C; Moczygemba, J; Weitman, S; Von Hoff, D D; Rowinsky, E K

    2000-12-15

    To evaluate the toxicity and pharmacologic behavior of the novel mushroom-derived cytotoxin irofulven administered as a 5-minute intravenous (IV) infusion daily for 5 days every 4 weeks to patients with advanced solid malignancies. In this phase I trial, 46 patients were treated with irofulven doses ranging from 1.0 to 17.69 mg/m(2) as a 5-minute IV infusion (two patients received a 1-hour infusion) daily for 5 days every 4 weeks. The modified continual reassessment method was used for dose escalation. Pharmacokinetic studies were performed on days 1 and 5 to characterize the plasma disposition of irofulven. Forty-six patients were treated with 92 courses of irofulven. The dose-limiting toxicities on this schedule were myelosuppression and renal dysfunction. At the 14.15-mg/m(2) dose level, renal dysfunction resembling renal tubular acidosis occurred in four of 10 patients and was ameliorated by prophylactic IV hydration. The 17.69-mg/m(2) dose level was not tolerated because of grade 4 neutropenia and renal toxicity, whereas the 14.15-mg/m(2) dose level was not tolerable with repetitive dosing because of persistent thrombocytopenia. Other common toxicities included mild to moderate nausea, vomiting, facial erythema, and fatigue. One partial response occurred in a patient with advanced, refractory metastatic pancreatic cancer lasting 7 months. Pharmacokinetic studies of irofulven revealed dose-proportional increases in both maximum plasma concentrations and area under the concentration-time curve, while the agent exhibited a rapid elimination half-life of 2 to 10 minutes. Given the results of this study, the recommended dose of irofulven is 10.64 mg/m(2) as a 5-minute IV infusion daily for 5 days every 4 weeks. The preliminary antitumor activity documented in a patient with advanced pancreatic cancer and the striking preclinical antitumor effects of irofulven observed on intermittent dosing schedules support further disease-directed evaluations of this agent on the

  1. Solid State Air Purification System Project

    Data.gov (United States)

    National Aeronautics and Space Administration — The purpose of this proposed research is to develop a new air purification system based on a liquid membrane, capable of purifying carbon dioxide from air in a far...

  2. Purification of Gaussian maximally mixed states

    Energy Technology Data Exchange (ETDEWEB)

    Jeong, Kabgyun [Center for Macroscopic Quantum Control, Department of Physics and Astronomy, Seoul National University, Seoul 08826 (Korea, Republic of); School of Computational Sciences, Korea Institute for Advanced Study, Seoul 02455 (Korea, Republic of); Lim, Youngrong, E-mail: sshaep@gmail.com [Center for Macroscopic Quantum Control, Department of Physics and Astronomy, Seoul National University, Seoul 08826 (Korea, Republic of)

    2016-10-23

    We find that the purifications of several Gaussian maximally mixed states (GMMSs) correspond to some Gaussian maximally entangled states (GMESs) in the continuous-variable regime. Here, we consider a two-mode squeezed vacuum (TMSV) state as a purification of the thermal state and construct a general formalism of the Gaussian purification process. Moreover, we introduce other kind of GMESs via the process. All of our purified states of the GMMSs exhibit Gaussian profiles; thus, the states show maximal quantum entanglement in the Gaussian regime. - Highlights: • Candidates of Gaussian maximally mixed state are proposed. • Obtaining Gaussian maximally entangled states using the purification process. • The suggested states can be applicable for the test of capacity problem in Gaussian regime.

  3. Purification and characterization of thermostable glucoamylase from ...

    African Journals Online (AJOL)

    Thermostable glucoamylase from Rhizopus oligosporus SK5 mutant was purified in a 3-step purification using Imarsil, activated charcoal and Sephadex-G-100 to achieve a 40-fold purification. The enzyme was optimally active at pH 5.0 and temperature of 80 °C. It exhibited a half-life of 60 minutes at 70 °C. Its stability was ...

  4. Purification processes for coal gasification

    Energy Technology Data Exchange (ETDEWEB)

    Fleming, D.K.; Primack, H.S.

    1977-01-01

    It is apparent from the discussion that many routes can be taken to achieve acid-gas removal and sulfur recovery from coal gas. The selection of the optimum purification system is a major task. The type of coal, type of gasifier and the upstream processing all strongly influence the selection. Several generalizations can be made: (1) The cost of the purification sections of a high-Btu gas plant is significant--perhaps 10 to 30% of the capital cost of the coal conversion facility. (2) The cost of purifying gas produced from high-sulfur coal feed is more expensive than the cost for purifying gas produced from low-sulfur coal. (3) The choice of an acid-gas removal system will often be a function of system pressure. The economical choice will usually be: (a) amine-based systems at atmospheric pressure; (b) hot-carbonate systems at moderate pressure or (c) physical-solvent systems at higher pressure. (4) For a high-Btu, high-sulfur case: (a) A selective acid-gas removal system with a Claus plant is probably more economical than a non-selective acid-gas system with liquid oxidation of the H/sub 2/S in the regenerator off-gas. (b) Even moderately selective systems can produce an H/sub 2/S-rich gas suitable for a Claus plant. The CO/sub 2/-rich gas may or may not require further sulfur removal, depending on the selectivity. (5) For a high-Btu, low-sulfur case: (a) The hot carbonate and tertiary amine systems may not be sufficiently selective to produce a gas suitable for feed to a Claus process while a physical solvent system may be. Therefore, the physical solvent system may be expected to be more economical. (b) The regenerated gas from the bulk CO/sub 2/ removal system following a selective physical solvent system may require further sulfur removal, depending upon the sulfur level in the initial feedstock and the selectivity of the system selected.

  5. Photocatalytic materials and technologies for air purification.

    Science.gov (United States)

    Ren, Hangjuan; Koshy, Pramod; Chen, Wen-Fan; Qi, Shaohua; Sorrell, Charles Christopher

    2017-03-05

    Since there is increasing concern for the impact of air quality on human health, the present work surveys the materials and technologies for air purification using photocatalytic materials. The coverage includes (1) current photocatalytic materials for the decomposition of chemical contaminants and disinfection of pathogens present in air and (2) photocatalytic air purification systems that are used currently and under development. The present work focuses on five main themes. First, the mechanisms of photodegradation and photodisinfection are explained. Second, system designs for photocatalytic air purification are surveyed. Third, the photocatalytic materials used for air purification and their characteristics are considered, including both conventional and more recently developed photocatalysts. Fourth, the methods used to fabricate these materials are discussed. Fifth, the most significant coverage is devoted to materials design strategies aimed at improving the performance of photocatalysts for air purification. The review concludes with a brief consideration of promising future directions for materials research in photocatalysis. Copyright © 2016 Elsevier B.V. All rights reserved.

  6. Simplified riboprobe purification using translucent straws as gel tubes.

    Science.gov (United States)

    Kol, S; Ben-Shlomo, I; Adashi, E Y; Rohan, R M

    1996-01-01

    Gel purification of radioactive riboprobes enhances the quality of the ribonuclease protection assay. A simple and effective method for riboprobe purification is described. The method uses acrylamide gels in plastic tubes to achieve electrophoretic separation of the RNA polymerase products.

  7. Ionic behavior of treated water at a water purification plant

    OpenAIRE

    Yanagida, Kazumi; Kawahigashi, Tatsuo

    2012-01-01

    [Abstract] Water at each processing stage in a water purification plant was extracted and analyzed to investigate changes of water quality. Investigations of water at each processing stage at the water purification plant are discussed herein.

  8. Purification treatment for underground water

    Energy Technology Data Exchange (ETDEWEB)

    Fonbershteyn, V.

    1985-08-01

    In order for underground water to be clean and to taste good, iron can be removed from it right underground, in the water-bearing stratum, before it is brought to the surface. G.M. Kommunar, V.S. Alekseyev, and V.T. Grebennikov, candidates of technical sciences and associates of the Moscow All-Union Hydrogeology Scientific Research Institute, developed the practical application of this beneficial technology, which makes it possible to do away with purification installations. With the new technology (Patent No. 985 214, 1 018 918) water saturated with oxygen is sent through an ejector and then pumped into a well. It passes through rocks that serve as a natural filter, and the filter is loaded with oxygen. The filter now becomes a barrier for mineral impurities contained in the artesian water. The amount of time needed to pump the oxidized water into the well is calculated beforehand, knowing the capacity of the water-bearing stratum, the porosity of the rocks, the expenditure of pumped oxidized water, and the radius of the zone of the filtering rocks. While the water is pumped out of the well, its properties are monitored periodically. If the concentration of iron exceeds the allowable norm-0.3 mg per liter-the extraction is halted, and oxidized water is once again pumped into the well. It is convenient and economical to combine several wells into one system, where each well will pump and accept water according to its own schedule. This new technology can also be used to remove manganese, heavy metals, and hydrogen sulfide from underground water.

  9. Non-Chromatographic Purification of Endohedral Metallofullerenes.

    Science.gov (United States)

    Wang, Zhiyong; Omachi, Haruka; Shinohara, Hisanori

    2017-04-29

    The purification of endohedral metallofullerenes by high performance liquid chromatography is very time-consuming and expensive. A number of rapid and inexpensive non-chromatographic methods have thus been developed for large-scale purification of metallofullerenes. In this review, we summarize recent advances in non-chromatographic purification methods of metallofullerenes. Lewis acid-based complexation is one of the most efficient and powerful methods for separation of metallofullerenes from empty fullerenes. The first oxidation potential of metallofullerenes is a critical factor that affects the separation efficiency of the Lewis acid-based method. Supramolecular methods are effective for separation of fullerenes and metallofullerenes that are different in size and shape. Chemical/electrochemical reduction and exohedral functionalization are also utilized to separate and purify metallofullerenes on a large scale.

  10. Purification of glycocalicin from human plasma.

    Science.gov (United States)

    HadjKacem, Basma; Mkaouar, Héla; Ben Amor, Ikram; Gargouri, Jalel; Gargouri, Ali

    2016-01-01

    Glycocalicin (GC) is a large extracellular proteolytic fragment of glycoprotein Ib, a membrane platelet component playing an essential role in the physiological processes of platelet adhesion and aggregation. GC contains the binding sites for thrombin and von Willebrand factor. GC circulates normally in vivo in significant concentrations and the plasma level of this protein reflects a complex function of factors including platelet count or platelet turnover. It can therefore serve as a good indicator for many diseases like hypoplastic thrombocytopenia and idiopathic thrombocytopenic purpura. For this reason, several purification assays have been previously described. In this work, we describe a novel analytical method for GC purification from human platelets based on preparative HPLC gel filtration followed by immuno-affinity chromatography on NHS activated column conjugated with specific antibody. Pure GC was obtained from tiny amount of starting material. Our protocol of GC purification is simple, fast and provides a pure end product. Copyright © 2015 Elsevier B.V. All rights reserved.

  11. 21 CFR 876.5665 - Water purification system for hemodialysis.

    Science.gov (United States)

    2010-04-01

    ... 21 Food and Drugs 8 2010-04-01 2010-04-01 false Water purification system for hemodialysis. 876... SERVICES (CONTINUED) MEDICAL DEVICES GASTROENTEROLOGY-UROLOGY DEVICES Therapeutic Devices § 876.5665 Water purification system for hemodialysis. (a) Identification. A water purification system for hemodialysis is a...

  12. Purification of contaminated groundwater by membrane technology

    Energy Technology Data Exchange (ETDEWEB)

    Youn, In Soo; Chung, Chin Ki; Kim, Byoung Gon [Korea Institute of Geology Mining and Materials, Taejon (Korea, Republic of)

    1996-12-01

    The objective of this study is to apply the membrane separation technology to the purification of contaminated ground water in Korea. Under this scope, the purification was aimed to the drinking water level. The scale of the membrane system was chosen to a small filtration plant for local clean water supplies and/or heavy purifiers for buildings and public uses. The actual conditions of ground water contamination in Korea was surveyed to determine the major components to remove under the drinking water requirements. To set up a hybrid process with membrane methods, conventional purification methods were also investigated for the comparison purpose. The research results are summarized as follows : 1) Contamination of the groundwater in Korea has been found to be widespread across the country. The major contaminant were nitrate, bacteria, and organic chlorides. Some solvents and heavy metals are also supposed to exist in the ground water of industrial complexes, cities, and abandoned mines. 2) The purification methods currently used in public filtration plants appear not to be enough for new contaminants from recent industrial expanding. The advanced purification technologies generally adopted for this problem have been found to be unsuitable due to their very complicated design and operation, and lack of confidence in the purification performance. 3) The reverse osmosis tested with FilmTec FT30 membrane was found to remove nitrate ions in water with over 90 % efficiency. 4) The suitable membrane process for the contaminated groundwater in Korea has been found to be the treatments composed of activated carbon, microfiltration, reverse osmosis or ultrafiltration, and disinfection. The activated carbon treatment could be omitted for the water of low organic contaminants. The microfiltration and the reverse osmosis treatments stand for the conventional methods of filtration plants and the advanced methods for hardly removable components, respectively. It is recommended

  13. Inhibition of leukemic U937 cell growth by induction of apoptosis, cell cycle arrest and suppression of VEGF, MMP-2 and MMP-9 activities by cytotoxin protein NN-32 purified from Indian spectacled cobra (Naja naja) venom.

    Science.gov (United States)

    Das, Tanaya; Bhattacharya, Shamik; Biswas, Archita; Gupta, Shubho Das; Gomes, Antony; Gomes, Aparna

    2013-04-01

    A cytotoxin NN-32 (6.7 kDa) from Indian cobra (Naja naja) venom inhibited human leukemic U937 cell growth as observed by Trypan blue dye exclusion method and cytotoxicity was confirmed by MTT assay. NN-32 induced apoptosis of U937 cell and cell cycle arrest of sub-G1 phase were revealed by FACS analysis. Increased Bax/Bcl-2 ratio, increased caspase 3 and 9 activities, cleaved PARP, decreased VEGF, MMP-2 and MMP-9 activities were observed after NN-32 treatment of U937 cell. Antileukemic activity of NN-32 on U937 cell may be due to activation of apoptosis, arresting cell cycle and antiangiogenesis activities. Copyright © 2013. Published by Elsevier Ltd.

  14. Chemical looping integration with a carbon dioxide gas purification unit

    Science.gov (United States)

    Andrus, Jr., Herbert E.; Jukkola, Glen D.; Thibeault, Paul R.; Liljedahl, Gregory N.

    2017-01-24

    A chemical looping system that contains an oxidizer and a reducer is in fluid communication with a gas purification unit. The gas purification unit has at least one compressor, at least one dryer; and at least one distillation purification system; where the gas purification unit is operative to separate carbon dioxide from other contaminants present in the flue gas stream; and where the gas purification unit is operative to recycle the contaminants to the chemical looping system in the form of a vent gas that provides lift for reactants in the reducer.

  15. Purification of tantalum by plasma arc melting

    Science.gov (United States)

    Dunn, Paul S.; Korzekwa, Deniece R.

    1999-01-01

    Purification of tantalum by plasma arc melting. The level of oxygen and carbon impurities in tantalum was reduced by plasma arc melting the tantalum using a flowing plasma gas generated from a gas mixture of helium and hydrogen. The flowing plasma gases of the present invention were found to be superior to other known flowing plasma gases used for this purpose.

  16. Purification And Characterization Of Marine Bacillus Thuringiensis ...

    African Journals Online (AJOL)

    Urease was purified to homogeneity from Bacillus thuringiensis N2 using different purification steps namely, 55% acetone precipitation, DEAE-Sephadex A50 anion exchange column and Sephadex G120-200 gel filtration chromatography. The enzyme was purified 95.27 fold and showed a final specific activity of 10.48 ...

  17. Purification and biochemical properties of carboxylesterase from ...

    African Journals Online (AJOL)

    The order of inhibition on EII was β- mercaptoethanol > PMSF > DTNB > PCMB > iodoacetate. While the order of inhibition on EIII was β-mercaptoethanol > iodoacetate > DTNB> PCM > PMSF. Keywords: Carboxylesterase, Fasciola gigantica; Purification; Characterization; p-Nitrophenyl; α-Naphthyl; β-Naphthyl; Esters ...

  18. Isolation, partial purification and characterization of antifungal ...

    African Journals Online (AJOL)

    SAM

    2014-07-16

    Jul 16, 2014 ... Full Length Research Paper. Isolation, partial purification and characterization of .... dried seeds were milled with coffee grinder and stored in air tight bags until required. Fifty grams of the matured ... was washed with 2.5% Triton-X 100 (3 × 20 min) with continual shaking to remove SDS followed by washing ...

  19. Isolation, purification and effects of hypoglycemic functional ...

    African Journals Online (AJOL)

    Isolation, purification and effects of hypoglycemic functional polysaccharides from Inonotus obliquus. Tao Hu, Ping Liu, Yuanying Ni, Chuntao Lu. Abstract. Inonotus obliquus is generally used for the treatment of diseases such as cancers, angiocardiopathy and diabetes. However, few studies are available on its functional ...

  20. Partial purification and biochemical characterization of acid ...

    African Journals Online (AJOL)

    hp

    Mung bean (Vigna radiata) is one of the important crops of the North Eastern Region of India. In the present study, acid phosphatase enzyme was isolated and partially purified from germinated local mung bean seeds. The sequential partial purification process was performed using ammonium sulphate precipitation method.

  1. Expression and Purification of Sperm Whale Myoglobin

    Science.gov (United States)

    Miller, Stephen; Indivero, Virginia; Burkhard, Caroline

    2010-01-01

    We present a multiweek laboratory exercise that exposes students to the fundamental techniques of bacterial expression and protein purification through the preparation of sperm whale myoglobin. Myoglobin, a robust oxygen-binding protein, contains a single heme that gives the protein a reddish color, making it an ideal subject for the teaching…

  2. Partial purification and characterization of polygalacturonase ...

    African Journals Online (AJOL)

    Yomi

    2012-04-03

    Apr 3, 2012 ... purification of pearl millet [Pennisetum glaucum (L.) R.Br.] protein extract by cation exchange chromatography, which ... electrophoresis showed prominent bands between 29 and 43 kDa, consistent with the molecular weights of the ... EPGs (De Lorenzo et al., 2001; Federici et al., 2006), but are shown to be ...

  3. Affinity purification of recombinant human plasminogen activator ...

    African Journals Online (AJOL)

    Purpose: To develop processes for effective isolation and purification of recombinant human plasminogen activator (rhPA) from transgenic rabbit milk. Methods: Immunoaffinity chromatography was selected and improved by a special polyol-responsive monoclonal antibody (PR-mAb). Alteplase was used as immunogen ...

  4. Purification and characterization of xylanase from Aspergillus ...

    African Journals Online (AJOL)

    Xylanase was subjected to a three-step purification scheme involving ammonium sulphate precipitation, gel filtration chromatography and anion exchange chromatography. Purity was verified by running the extracted protein on sodium dodecyl sulfate-polyacrylamide gel electrophoresis (SDS-PAGE) and a single band was ...

  5. Rapid cloning and purification of proteins: gateway vectors for protein purification by self-cleaving tags.

    Science.gov (United States)

    Gillies, Alison R; Hsii, Judy F; Oak, Seachol; Wood, David W

    2008-10-01

    We have combined Invitrogen's Gateway cloning technology with self-cleaving purification tags to generate a new system for rapid production of recombinant protein products. To accomplish this, we engineered our previously reported DeltaI-CM cleaving intein to include a Gateway cloning recognition sequence, and demonstrated that the resulting Gateway-competent intein is unaffected. This intein can therefore be used in several previously reported purification methods, while at the same time being compatible with Gateway cloning. We have incorporated this intein into a set of Gateway vectors, which include self-cleaving elastin-like polypeptide (ELP), chitin binding domain (CBD), phasin (polyhydroxybutyrate-binding), or maltose binding domain (MBD) tags. These vectors were verified by Gateway cloning of TEM-1 beta-lactamase and Escherichia coli catalase genes, and the expressed target proteins were purified using the four methods encoded on the vectors. The purification methods were unaffected by replacing the DeltaI-CM intein with the Gateway intein. It was observed that some purification methods were more appropriate for each target than others, suggesting utility of this technology for rapid process identification and optimization. The modular design of the Gateway system and intein purification method suggests that any tag and promoter can be trivially added to this system for the development of additional expression vectors. This technology could greatly facilitate process optimization, allowing several targets and methods to be tested in a high-throughput manner.

  6. The Viability of Photocatalysis for Air Purification

    Directory of Open Access Journals (Sweden)

    Stephen O. Hay

    2015-01-01

    Full Text Available Photocatalytic oxidation (PCO air purification technology is reviewed based on the decades of research conducted by the United Technologies Research Center (UTRC and their external colleagues. UTRC conducted basic research on the reaction rates of various volatile organic compounds (VOCs. The knowledge gained allowed validation of 1D and 3D prototype reactor models that guided further purifier development. Colleagues worldwide validated purifier prototypes in simulated realistic indoor environments. Prototype products were deployed in office environments both in the United States and France. As a result of these validation studies, it was discovered that both catalyst lifetime and byproduct formation are barriers to implementing this technology. Research is ongoing at the University of Connecticut that is applicable to extending catalyst lifetime, increasing catalyst efficiency and extending activation wavelength from the ultraviolet to the visible wavelengths. It is critical that catalyst lifetime is extended to realize cost effective implementation of PCO air purification.

  7. Nanomaterials and Water Purification: Opportunities and Challenges

    Science.gov (United States)

    Savage, Nora; Diallo, Mamadou S.

    2005-10-01

    Advances in nanoscale science and engineering suggest that many of the current problems involving water quality could be resolved or greatly ameliorated using nanosorbents, nanocatalysts, bioactive nanoparticles, nanostructured catalytic membranes and nanoparticle enhanced filtration among other products and processes resulting from the development of nanotechnology. Innovations in the development of novel technologies to desalinate water are among the most exciting and promising. Additionally, nanotechnology-derived products that reduce the concentrations of toxic compounds to sub-ppb levels can assist in the attainment of water quality standards and health advisories. This article gives an overview of the use of nanomaterials in water purification. We highlight recent advances on the development of novel nanoscale materials and processes for treatment of surface water, groundwater and industrial wastewater contaminated by toxic metal ions, radionuclides, organic and inorganic solutes, bacteria and viruses. In addition, we discuss some challenges associated with the development of cost effective and environmentally acceptable functional nanomaterials for water purification.

  8. Nanotechnology for water treatment and purification

    CERN Document Server

    Apblett, Allen

    2014-01-01

    This book describes the latest progress in the application of nanotechnology for water treatment and purification. Leaders in the field present both the fundamental science and a comprehensive overview of the diverse range of tools and technologies that have been developed in this critical area. Expert chapters present the unique physicochemical and surface properties of nanoparticles and the advantages that these provide for engineering applications that ensure a supply of safe drinking water for our growing population. Application areas include generating fresh water from seawater, preventing contamination of the environment, and creating effective and efficient methods for remediation of polluted waters. The chapter authors are leading world-wide experts in the field with either academic or industrial experience, ensuring that this comprehensive volume presents the state-of-the-art in the integration of nanotechnology with water treatment and purification. Covers both wastewater and drinking water treatmen...

  9. Propagation and Purification of Baculovirus oryctes Huger

    Directory of Open Access Journals (Sweden)

    Susamto Somowiyarjo

    1995-12-01

    Full Text Available An isolate of Baculovirus oryctes, a possible biological control agent for coconut beetle (Oryctes rhinoceros Huger from East Java was propagated and purified. The virus could be transmitted by feeding the imago with 10% sucrose containing virus from homogenate of infected beetles. Effectivity of virus to 9 healthy females by sexual copulation. Virus be succesfully purified by a method of Payne. Key words: Baculovirus oryctes, transmission, purification

  10. Conductive diamond electrodes for water purification

    Directory of Open Access Journals (Sweden)

    Carlos Alberto Martínez-Huitle

    2007-12-01

    Full Text Available Nowadays, synthetic diamond has been studied for its application in wastewater treatment, electroanalysis, organic synthesis and sensor areas; however, its use in the water disinfection/purification is its most relevant application. The new electrochemistry applications of diamond electrodes open new perspectives for an easy, effective, and chemical free water treatment. This article highlights and summarizes the results of a selection of papers dealing with electrochemical disinfection using synthetic diamond films.

  11. Purification of metal-organic framework materials

    Science.gov (United States)

    Farha, Omar K.; Hupp, Joseph T.

    2015-06-30

    A method of purification of a solid mixture of a metal-organic framework (MOF) material and an unwanted second material by disposing the solid mixture in a liquid separation medium having a density that lies between those of the wanted MOF material and the unwanted material, whereby the solid mixture separates by density differences into a fraction of wanted MOF material and another fraction of unwanted material.

  12. Extracorporeal Blood Purification in Burns: A Review

    Science.gov (United States)

    2014-09-01

    Second International Consensus Conference of the Acute Dialysis Quality Initiative (ADQI) Group. Crit Care 2004;8:R204 12. [54] Haase M, Bellomo R...month mortality. While the specific pathogens may differ between centers, Gram negative organisms are the leading cause of life threatening infections in...extended dialysis . Blood Purif 2012;34:246 52. [56] Morgera S, Klonower D, Rocktaschel J, Haase M, Priem F, Ziemer S, et al. TNF elimination with

  13. Purification of crude biodiesel using dry washing and membrane technologies

    Directory of Open Access Journals (Sweden)

    I.M. Atadashi

    2015-12-01

    Full Text Available Purification of crude biodiesel is mandatory for the fuel to meet the strict international standard specifications for biodiesel. Therefore, this paper carefully analyzed recently published literatures which deal with the purification of biodiesel. As such, dry washing technologies and the most recent membrane biodiesel purification process have been thoroughly examined. Although purification of biodiesel using dry washing process involving magnesol and ion exchange resins provides high-quality biodiesel fuel, considerable amount of spent absorbents is recorded, besides the skeletal knowledge on its operating process. Further, recent findings have shown that biodiesel purification using membrane technique could offer high-quality biodiesel fuel with less wastewater discharges. Thus, both researchers and industries are expected to benefit from the development of membrane technique in purifying crude biodiesel. As well biodiesel purification via membranes has been shown to be environmentally friendly. For these reasons, it is important to explore and exploit membrane technology to purify crude biodiesel.

  14. Rotating Reverse-Osmosis for Water Purification

    Science.gov (United States)

    Lueptow, RIchard M.

    2004-01-01

    A new design for a water-filtering device combines rotating filtration with reverse osmosis to create a rotating reverse- osmosis system. Rotating filtration has been used for separating plasma from whole blood, while reverse osmosis has been used in purification of water and in some chemical processes. Reverse- osmosis membranes are vulnerable to concentration polarization a type of fouling in which the chemicals meant not to pass through the reverse-osmosis membranes accumulate very near the surfaces of the membranes. The combination of rotating filtration and reverse osmosis is intended to prevent concentration polarization and thereby increase the desired flux of filtered water while decreasing the likelihood of passage of undesired chemical species through the filter. Devices based on this concept could be useful in a variety of commercial applications, including purification and desalination of drinking water, purification of pharmaceutical process water, treatment of household and industrial wastewater, and treatment of industrial process water. A rotating filter consists of a cylindrical porous microfilter rotating within a stationary concentric cylindrical outer shell (see figure). The aqueous suspension enters one end of the annulus between the inner and outer cylinders. Filtrate passes through the rotating cylindrical microfilter and is removed via a hollow shaft. The concentrated suspension is removed at the end of the annulus opposite the end where the suspension entered.

  15. Compressorless Gas Storage and Regenerative Hydrogen Purification Project

    Data.gov (United States)

    National Aeronautics and Space Administration — Microwave regenerative sorption media gas storage/delivery techniques are proposed to address both compressed gas management and hydrogen purification requirements...

  16. Data of expression and purification of recombinant Taq DNA polymerase

    Directory of Open Access Journals (Sweden)

    Na Fang

    2016-12-01

    Full Text Available Polymerase chain reaction (PCR technique is widely used in many experimental conditions, and Taq DNA polymerase is critical in PCR process. In this article, the Taq DNA polymerase expression plasmid is reconstructed and the protein product is obtained by rapid purification, (“Rapid purification of high-activity Taq DNA polymerase” (Pluthero, 1993 [1], “Single-step purification of a thermostable DNA polymerase expressed in Escherichia coli” (Desai and Pfaffle, 1995 [2]. Here we present the production data from protein expression and provide the analysis results of the production from two different vectors. Meanwhile, the purification data is also provided to show the purity of the protein product.

  17. Dense Medium Plasma Water Purification Reactor (DMP WaPR) Project

    Data.gov (United States)

    National Aeronautics and Space Administration — The Dense Medium Plasma Water Purification Reactor offers significant improvements over existing water purification technologies used in Advanced Life Support...

  18. Ribonucleoprotein purification and characterization using RNA Mango.

    Science.gov (United States)

    Panchapakesan, Shanker Shyam S; Ferguson, Matthew L; Hayden, Eric J; Chen, Xin; Hoskins, Aaron A; Unrau, Peter J

    2017-10-01

    The characterization of RNA-protein complexes (RNPs) is a difficult but increasingly important problem in modern biology. By combining the compact RNA Mango aptamer with a fluorogenic thiazole orange desthiobiotin (TO1-Dtb or TO3-Dtb) ligand, we have created an RNA tagging system that simplifies the purification and subsequent characterization of endogenous RNPs. Mango-tagged RNP complexes can be immobilized on a streptavidin solid support and recovered in their native state by the addition of free biotin. Furthermore, Mango-based RNP purification can be adapted to different scales of RNP isolation ranging from pull-down assays to the isolation of large amounts of biochemically defined cellular RNPs. We have incorporated the Mango aptamer into the S. cerevisiae U1 small nuclear RNA (snRNA), shown that the Mango-snRNA is functional in cells, and used the aptamer to pull down a U1 snRNA-associated protein. To demonstrate large-scale isolation of RNPs, we purified and characterized bacterial RNA polymerase holoenzyme (HE) in complex with a Mango-containing 6S RNA. We were able to use the combination of a red-shifted TO3-Dtb ligand and eGFP-tagged HE to follow the binding and release of the 6S RNA by two-color native gel analysis as well as by single-molecule fluorescence cross-correlation spectroscopy. Together these experiments demonstrate how the Mango aptamer in conjunction with simple derivatives of its flurophore ligands enables the purification and characterization of endogenous cellular RNPs in vitro. © 2017 Panchapakesan et al.; Published by Cold Spring Harbor Laboratory Press for the RNA Society.

  19. Purification of cadmium up to 5N+ by vacuum distillation

    Indian Academy of Sciences (India)

    Unknown

    electric vehicle and remote area storage systems. The problem connected with profound purification and characterization of the materials that are used for the synthesis of electronic materials remains critical. The advantage of methods providing efficient purification using rather simple and cheap equipment is evident. As.

  20. Necessity of purification during bacterial DNA extraction with environmental soils

    Directory of Open Access Journals (Sweden)

    Hyun Jeong Lim

    2017-08-01

    Full Text Available Complexity and heterogeneity of soil samples have often implied the inclusion of purification steps in conventional DNA extraction for polymerase chain reaction (PCR assays. Unfortunately the purification steps are also time and labor intensive. Therefore the necessity of DNA purification was re-visited and investigated for a variety of environmental soil samples that contained various amounts of PCR inhibitors. Bead beating and centrifugation was used as the baseline (without purification method for DNA extraction. Its performance was compared with that of conventional DNA extraction kit (with purification. The necessity criteria for DNA purification were established with environmental soil samples. Using lysis conditions at 3000 rpm for 3 minutes with 0.1 mm glass beads, centrifugation time of 10 minutes and 1:10 dilution ratio, the baseline method outperformed conventional DNA extraction on cell seeded sand samples. Further investigation with PCR inhibitors (i.e., humic acids, clay, and magnesium [Mg] showed that sand samples containing less than 10 μg/g humic acids and 70% clay may not require purifications. Interestingly, the inhibition pattern of Mg ion was different from other inhibitors due to the complexation interaction of Mg ion with DNA fragments. It was concluded that DNA extraction method without purification is suitable for soil samples that have less than 10 μg/g of humic acids, less than 70% clay content and less than 0.01% Mg ion content.

  1. A scintillator purification system for the Borexino solar neutrino detector

    Science.gov (United States)

    Benziger, J.; Cadonati, L.; Calaprice, F.; Chen, M.; Corsi, A.; Dalnoki-Veress, F.; Fernholz, R.; Ford, R.; Galbiati, C.; Goretti, A.; Harding, E.; Ianni, Aldo; Ianni, Andrea; Kidner, S.; Leung, M.; Loeser, F.; McCarty, K.; McKinsey, D.; Nelson, A.; Pocar, A.; Salvo, C.; Schimizzi, D.; Shutt, T.; Sonnenschein, A.

    2008-03-01

    Purification of the 278 tons of liquid scintillator and 889 tons of buffer shielding for the Borexino solar neutrino detector is performed with a system that combines distillation, water extraction, gas stripping, and filtration. This paper describes the principles of operation, design, and construction of that purification system, and reviews the requirements and methods to achieve system cleanliness and leak-tightness.

  2. One step purification of biological active human interleukin-2 protein ...

    African Journals Online (AJOL)

    Pharmacological importance of recombinant human interleukin-2 protein has increased the demand to establish effective, reliable and cost effective chromatography method for its production and purification on large scale. One step mimetic ligand affinity chromatography method for purification of mutated human ...

  3. A scintillator purification system for the Borexino solar neutrino detector

    Energy Technology Data Exchange (ETDEWEB)

    Benziger, J. [Chemical Engineering Department, Princeton University, Princeton, NJ 08544 (United States)], E-mail: benziger@princeton.edu; Cadonati, L.; Calaprice, F.; Chen, M.; Corsi, A.; Dalnoki-Veress, F.; Fernholz, R.; Ford, R.; Galbiati, C. [Physics Department, Princeton University, Princeton, NJ 08544 (United States); Goretti, A. [INFN, Laboratori Nazionale di Gran Sasso (Italy); Harding, E. [Physics Department, Princeton University, Princeton, NJ 08544 (United States); Ianni, Aldo [INFN, Laboratori Nazionale di Gran Sasso (Italy); Ianni, Andrea; Kidner, S.; Leung, M.; Loeser, F.; McCarty, K.; McKinsey, D.; Nelson, A.; Pocar, A. [Physics Department, Princeton University, Princeton, NJ 08544 (United States)] (and others)

    2008-03-21

    Purification of the 278 tons of liquid scintillator and 889 tons of buffer shielding for the Borexino solar neutrino detector is performed with a system that combines distillation, water extraction, gas stripping, and filtration. This paper describes the principles of operation, design, and construction of that purification system, and reviews the requirements and methods to achieve system cleanliness and leak-tightness.

  4. One-step purification of E. coli elongation factor Tu

    DEFF Research Database (Denmark)

    Knudsen, Charlotte Rohde; Clark, Brian F. C.; Degn, B

    1993-01-01

    The tuf A gene, encoding the E. coli elongation factor Tu, was cloned in the pGEX gene fusion system. Upon expression EF-Tu is fused to glutathione-S-transferase serving as a purification handle with affinity for glutathione immobilised on agarose. This allows purification of EF-Tu in a one...

  5. Expression and purification of recombinant Shiga toxin 2B from ...

    African Journals Online (AJOL)

    The two step purification trains were used to purify native Stx2B. First step purification was Ni-immobilized metal ion affinity chromatography (IMAC) column, followed by second step using HaloLink resin. The native Stx2B was obtained after column cleavage of halo-tag using HaloTEV protease. Maximum protein expression ...

  6. Purification and characterization of a novel extracellular chitinase ...

    African Journals Online (AJOL)

    The purification and characterization of a new thermophilic chitinase from thermophilic Bacillus sp. HU1, originally isolated from a soil sample collected from hot spring of XinJiang Province, China, is presented in this paper. The purification steps included ammonium sulfate precipitation, with columns of DEAE-Sepharose ...

  7. Purification and characterization of a protease from Thermophilic ...

    African Journals Online (AJOL)

    The purification and characterization of a thermophilic neutral protease from Thermophilic bacillus strain HS08, originally isolated from a soil sample collected from the Tulufan Crater of China, is presented in this paper. The purification steps included ammonium sulfate precipitation, with columns of DEAE-Sepharose anion ...

  8. Comparative study of peroxidase purification from apple and orange ...

    African Journals Online (AJOL)

    This paper reports the isolation and purification of peroxidase from low cost material; moreover, no significant work has been done on the isolation and purification of peroxidase from such cost effective sources (apple and orange seeds). Peroxidases had attracted considerable interest in recent years because of their ...

  9. The technology of large-scale pharmaceutical plasmid purification ...

    African Journals Online (AJOL)

    STORAGESEVER

    2010-01-04

    Jan 4, 2010 ... As an alternative, a new plasmid purification technology with cetyltrimethylammonium bromide (CTAB) is ... into account the application of animal. DNA vaccine, the cost of purification must be decreased. .... mixture was immediately transfered to 20, 26, 32 and 42°C water bath for another 10 min, afterwards ...

  10. Optimized conditions for high-level solubilization and purification of ...

    African Journals Online (AJOL)

    NH Computers

    2011-10-10

    Oct 10, 2011 ... In this report, we describe the cloning, over-expression, efficient solubilization, purification and evaluation of bioactivity of camel ... Key words: Recombinant growth hormone, somatotropin, cloning, expression, inclusion bodies, solubilization, purification ... 2003; Guakhar and Bernard, 2004). Thus, camel ...

  11. Surface processes during purification of InP quantum dots.

    Science.gov (United States)

    Mordvinova, Natalia; Emelin, Pavel; Vinokurov, Alexander; Dorofeev, Sergey; Abakumov, Artem; Kuznetsova, Tatiana

    2014-01-01

    Recently, a new simple and fast method for the synthesis of InP quantum dots by using phosphine as phosphorous precursor and myristic acid as surface stabilizer was reported. Purification after synthesis is necessary to obtain samples with good optical properties. Two methods of purification were compared and the surface processes which occur during purification were studied. Traditional precipitation with acetone is accompanied by a small increase in photoluminescence. It occurs that during the purification the hydrolysis of the indium precursor takes place, which leads to a better surface passivation. The electrophoretic purification technique does not increase luminescence efficiency but yields very pure quantum dots in only a few minutes. Additionally, the formation of In(OH)3 during the low temperature synthesis was explained. Purification of quantum dots is a very significant part of postsynthetical treatment that determines the properties of the material. But this subject is not sufficiently discussed in the literature. The paper is devoted to the processes that occur at the surface of quantum dots during purification. A new method of purification, electrophoresis, is investigated and described in particular.

  12. Surface processes during purification of InP quantum dots

    Directory of Open Access Journals (Sweden)

    Natalia Mordvinova

    2014-08-01

    Full Text Available Recently, a new simple and fast method for the synthesis of InP quantum dots by using phosphine as phosphorous precursor and myristic acid as surface stabilizer was reported. Purification after synthesis is necessary to obtain samples with good optical properties. Two methods of purification were compared and the surface processes which occur during purification were studied. Traditional precipitation with acetone is accompanied by a small increase in photoluminescence. It occurs that during the purification the hydrolysis of the indium precursor takes place, which leads to a better surface passivation. The electrophoretic purification technique does not increase luminescence efficiency but yields very pure quantum dots in only a few minutes. Additionally, the formation of In(OH3 during the low temperature synthesis was explained. Purification of quantum dots is a very significant part of postsynthetical treatment that determines the properties of the material. But this subject is not sufficiently discussed in the literature. The paper is devoted to the processes that occur at the surface of quantum dots during purification. A new method of purification, electrophoresis, is investigated and described in particular.

  13. Effect of Purification Procedures on DIF Analysis in IRTPRO

    Science.gov (United States)

    Fikis, David R. J.; Oshima, T. C.

    2017-01-01

    Purification of the test has been a well-accepted procedure in enhancing the performance of tests for differential item functioning (DIF). As defined by Lord, purification requires reestimation of ability parameters after removing DIF items before conducting the final DIF analysis. IRTPRO 3 is a recently updated program for analyses in item…

  14. Comparative study of peroxidase purification from apple and orange ...

    African Journals Online (AJOL)

    Jane

    2011-07-06

    Jul 6, 2011 ... This paper reports the isolation and purification of peroxidase from low cost material; moreover, no significant work has been done on the isolation and purification of peroxidase from such cost effective sources (apple and orange seeds). Peroxidases had attracted considerable interest in recent years.

  15. Biopharmaceuticals from microorganisms: from production to purification.

    Science.gov (United States)

    Jozala, Angela Faustino; Geraldes, Danilo Costa; Tundisi, Louise Lacalendola; Feitosa, Valker de Araújo; Breyer, Carlos Alexandre; Cardoso, Samuel Leite; Mazzola, Priscila Gava; Oliveira-Nascimento, Laura de; Rangel-Yagui, Carlota de Oliveira; Magalhães, Pérola de Oliveira; Oliveira, Marcos Antonio de; Pessoa, Adalberto

    2016-12-01

    The use of biopharmaceuticals dates from the 19th century and within 5-10 years, up to 50% of all drugs in development will be biopharmaceuticals. In the 1980s, the biopharmaceutical industry experienced a significant growth in the production and approval of recombinant proteins such as interferons (IFN α, β, and γ) and growth hormones. The production of biopharmaceuticals, known as bioprocess, involves a wide range of techniques. In this review, we discuss the technology involved in the bioprocess and describe the available strategies and main advances in microbial fermentation and purification process to obtain biopharmaceuticals. Copyright © 2016 Sociedade Brasileira de Microbiologia. Published by Elsevier Editora Ltda. All rights reserved.

  16. Biopharmaceuticals from microorganisms: from production to purification

    Directory of Open Access Journals (Sweden)

    Angela Faustino Jozala

    Full Text Available ABSTRACT The use of biopharmaceuticals dates from the 19th century and within 5-10 years, up to 50% of all drugs in development will be biopharmaceuticals. In the 1980s, the biopharmaceutical industry experienced a significant growth in the production and approval of recombinant proteins such as interferons (IFN α, β, and γ and growth hormones. The production of biopharmaceuticals, known as bioprocess, involves a wide range of techniques. In this review, we discuss the technology involved in the bioprocess and describe the available strategies and main advances in microbial fermentation and purification process to obtain biopharmaceuticals.

  17. Twist-off purification of hair bundles.

    Science.gov (United States)

    Shin, Jung-Bum; Pagana, James; Gillespie, Peter G

    2009-01-01

    Purification of hair bundles from inner-ear organs allows biochemical analysis of bundle constituents, including proteins and lipids. We describe here the "twist-off" method of bundle isolation, where dissected inner-ear organs are embedded in agarose, then subjected to a mechanical disruption that shears off bundles and leaves them in agarose blocks. With care in the dissection and in clean-up of the isolated bundles, contamination from cell bodies can be kept to a minimum. Isolated bundles can be analyzed by a variety of techniques, including immunocytochemistry, SDS-PAGE, immunoblotting, and mass spectrometry.

  18. Enzymatic Purification of Microplastics in Environmental Samples.

    Science.gov (United States)

    Löder, Martin G J; Imhof, Hannes K; Ladehoff, Maike; Löschel, Lena A; Lorenz, Claudia; Mintenig, Svenja; Piehl, Sarah; Primpke, Sebastian; Schrank, Isabella; Laforsch, Christian; Gerdts, Gunnar

    2017-12-19

    Micro-Fourier transform infrared (micro-FTIR) spectroscopy and Raman spectroscopy enable the reliable identification and quantification of microplastics (MPs) in the lower micron range. Since concentrations of MPs in the environment are usually low, the large sample volumes required for these techniques lead to an excess of coenriched organic or inorganic materials. While inorganic materials can be separated from MPs using density separation, the organic fraction impedes the ability to conduct reliable analyses. Hence, the purification of MPs from organic materials is crucial prior to conducting an identification via spectroscopic techniques. Strong acidic or alkaline treatments bear the danger of degrading sensitive synthetic polymers. We suggest an alternative method, which uses a series of technical grade enzymes for purifying MPs in environmental samples. A basic enzymatic purification protocol (BEPP) proved to be efficient while reducing 98.3 ± 0.1% of the sample matrix in surface water samples. After showing a high recovery rate (84.5 ± 3.3%), the BEPP was successfully applied to environmental samples from the North Sea where numbers of MPs range from 0.05 to 4.42 items m-3. Experiences with different environmental sample matrices were considered in an improved and universally applicable version of the BEPP, which is suitable for focal plane array detector (FPA)-based micro-FTIR analyses of water, wastewater, sediment, biota, and food samples.

  19. Design strategies for integrated protein purification processes: challenges, progress and outlook

    NARCIS (Netherlands)

    Nfor, B.; Ahamed, T.; Dedem, G.; Wielen, van der L.; Sandt, van de E.; Eppink, M.H.M.; Ottens, M.

    2008-01-01

    The key to successful and efficient protein purification is the selection of the most appropriate purification techniques and their combination in a logical way to obtain the desired purification in the minimum number of steps. However, the rationalization of protein purification process development

  20. CdTe substrate purification from impurities by gettering

    Science.gov (United States)

    Kosyachenko, Leonid A.; Zakharuk, Z. I.; Rarenko, A. I.; Nykonyuk, E. S.

    2001-02-01

    A possibility of CdTe substrate purification from impurities by structure-breakdown layer gettering, formed by laser irradiation, is considered. For profile calculation of diffusive distribution of point defects during heat treatment, and also substrate purification degree after heat treatment, a model, based on diffusion equation with consideration of impurity absorption by dislocations, is proposed. Impurity redistribution task in structure CdTe-CdHgTe during annealing is solved also. Investigations, carried out on specially prepared samples, confirmed CdTe purification effectiveness by gettering: impurity concentration decreased in 5 - 10 times.

  1. Nylon wool purification alters the activation of T cells.

    Science.gov (United States)

    Wohler, Jillian E; Barnum, Scott R

    2009-02-01

    Purification of lymphocytes, particularly T cells, is commonly performed using nylon wool. This enrichment method selectively retains B cells and some myeloid cells allowing a significantly more pure T cell population to flow through a nylon wool column. T cells purified in this fashion are assumed to be unaltered and functionally naïve, however some studies have suggested aberrant in vitro T cell responses after nylon wool treatment. We found that nylon wool purification significantly altered T cell proliferation, expression of activation markers and production of cytokines. Our results suggest that nylon wool treatment modifies T cell activation responses and that caution should be used when choosing this purification method.

  2. Hydrogen Purification Using Natural Zeolite Membranes

    Science.gov (United States)

    DelValle, William

    2003-01-01

    The School of Science at Universidad del Turabo (UT) have a long-lasting investigation plan to study the hydrogen cleaning and purification technologies. We proposed a research project for the synthesis, phase analysis and porosity characterization of zeolite based ceramic perm-selective membranes for hydrogen cleaning to support NASA's commitment to achieving a broad-based research capability focusing on aerospace-related issues. The present study will focus on technology transfer by utilizing inorganic membranes for production of ultra-clean hydrogen for application in combustion. We tested three different natural zeolite membranes (different particle size at different temperatures and time of exposure). Our results show that the membranes exposured at 900 C for 1Hr has the most higher permeation capacity, indicated that our zeolite membranes has the capacity to permeate hydrogen.

  3. Overview of Albumin and Its Purification Methods.

    Science.gov (United States)

    Raoufinia, Ramin; Mota, Ali; Keyhanvar, Neda; Safari, Fatemeh; Shamekhi, Sara; Abdolalizadeh, Jalal

    2016-12-01

    As the most frequent plasma protein, albumin constitutes more than 50% of the serum proteins in healthy individuals. It has a key role in oncotic pressure maintenance and it is known as a versatile protein carrier for transportation of various endogenous and exogenous ligands. Reduced amounts of albumin in the body will lead to different kinds of diseases such as hypovolemia and hypoproteinemia. It also has various indications in shocks, burns, cardiopulmonary bypass, acute liver failure and etc. Further applications in research consist of cell culture supplement, drug delivery carrier and protein/drug stabilizer. So, the demand for albumin increased annually worldwide. Due to different applications of albumin, many efforts have been accomplished to achieve albumin during a long period of time. In this review, an overview of serum albumin and different purification methods are summarized.

  4. Purification of equine Gc-globulin

    DEFF Research Database (Denmark)

    Houen, Gunnar; Pihl, Tina Holberg; Andersen, Pia Haubro

    Objectives With the aim of producing antibodies for an equine Group specific component (Gc)-globulin assay, the protein was purified from normal equine plasma. Methods Equine Gc-globulin was purified from healthy horse plasma using ion exchange chromatography (Q-Sepharose, CM......-Sepharose) and preparative PAGE. Results Equine Gc-globulin has successfully been purified from healthy horse plasma and rabbits and mice are being immunized to produce specific antibodies. Conclusions Purification of equine Gc-globulin and the production of specific antibodies will make it possible to develop an assay...... to be a sensitive marker of acute tissue injury and fatal outcome in humans. Patients with a low plasma concentration of Gc-globulin due to severe tissue injury might potentially benefit from infusions with purified Gc-globulin [1]. With an equine Gc-globulin assay, future studies will investigate the concentration...

  5. Concentration and purification of plutonium or thorium

    Science.gov (United States)

    Hayden, John A.; Plock, Carl E.

    1976-01-01

    In this invention a first solution obtained from such as a plutonium/thorium purification process or the like, containing plutonium (Pu) and/or thorium (Th) in such as a low nitric acid (HNO.sub.3) concentration may have the Pu and/or Th separated and concentrated by passing an electrical current from a first solution having disposed therein an anode to a second solution having disposed therein a cathode and separated from the first solution by a cation permeable membrane, the Pu or Th cation permeating the cation membrane and forming an anionic complex within the second solution, and electrical current passage affecting the complex formed to permeate an anion membrane separating the second solution from an adjoining third solution containing disposed therein an anode, thereby effecting separation and concentration of the Pu and/or Th in the third solution.

  6. [Periphyton and its application in water purification].

    Science.gov (United States)

    Chen, Chong-Jun; Han, Zhi-Ying; Zhu, Yin-Mei; Wu, Wei-Xiang

    2009-11-01

    Periphyton widely exists in natural water bodies, with the characteristics of huge biomass generation, strong ecological function, and sensitive response to water quality. It removes the pollutants in water bodies mainly through the processes of absorption, metabolism, adsorption, and complexation, etc. Owing to its high tolerance against pollution and high removal efficiency for nitrogen and phosphorus, as well as the feasibility of recycling its cells at low cost, periphyton is a promising candidate for developing the treatment techniques of water purification. The newly-developed artificial periphyton systems, e.g., algal turf scrubbers, periphyton biofilm systems, periphyton aquaculture systems, have been successfully applied in treating livestock manure, aquaculture wastewater, and municipal sewage. However, further researches are needed to understand the growth patterns of periphyton, its physiological responses to pollutants concentration, and its molecular biological mechanisms in removing pollutants.

  7. Nanocellulose-Based Materials for Water Purification.

    Science.gov (United States)

    Voisin, Hugo; Bergström, Lennart; Liu, Peng; Mathew, Aji P

    2017-03-05

    Nanocellulose is a renewable material that combines a high surface area with high strength, chemical inertness, and versatile surface chemistry. In this review, we will briefly describe how nanocellulose is produced, and present-in particular, how nanocellulose and its surface modified versions affects the adsorption behavior of important water pollutants, e.g., heavy metal species, dyes, microbes, and organic molecules. The processing of nanocellulose-based membranes and filters for water purification will be described in detail, and the uptake capacity, selectivity, and removal efficiency will also be discussed. The processing and performance of nanocellulose-based membranes, which combine a high removal efficiency with anti-fouling properties, will be highlighted.

  8. Nanocellulose-Based Materials for Water Purification

    Directory of Open Access Journals (Sweden)

    Hugo Voisin

    2017-03-01

    Full Text Available Nanocellulose is a renewable material that combines a high surface area with high strength, chemical inertness, and versatile surface chemistry. In this review, we will briefly describe how nanocellulose is produced, and present—in particular, how nanocellulose and its surface modified versions affects the adsorption behavior of important water pollutants, e.g., heavy metal species, dyes, microbes, and organic molecules. The processing of nanocellulose-based membranes and filters for water purification will be described in detail, and the uptake capacity, selectivity, and removal efficiency will also be discussed. The processing and performance of nanocellulose-based membranes, which combine a high removal efficiency with anti-fouling properties, will be highlighted.

  9. PURIFICATION OF STAPHYLOCOCCAL ALPHA-HEMOLYSIN

    Science.gov (United States)

    Madoff, Morton A.; Weinstein, Louis

    1962-01-01

    Madoff, Morton A. (New England Center Hospital, Boston, Mass.) and Louis Weinstein. Purification of staphylococcal alpha-hemolysin. J. Bacteriol. 83:914–918. 1962. — Staphylococcal alpha-toxin has been purified by metal-ion precipitation, column chromatography, and continuous-flow paper electrophoresis. The resultant preparation is highly unstable. A single line is obtained on agar-gel diffusion analysis against staphylococcal antiserum. Two bands, both associated with hemolytic activity and migrating cathodally at pH 8.4, are obtained on starch-gel electrophoresis. Levels of lytic, dermonecrotic, and lethal toxin activity maintain a constant relationship in the crude and purified materials, suggesting the unity of these effects. PMID:14468121

  10. Submersible purification system for radioactive water

    Science.gov (United States)

    Abbott, Michael L.; Lewis, Donald R.

    1989-01-01

    A portable, submersible water purification system for use in a pool of water containing radioactive contamination includes a prefilter for filtering particulates from the water. A resin bed is then provided for removal of remaining dissolved, particulate, organic, and colloidal impurities from the prefiltered water. A sterilizer then sterilizes the water. The prefilter and resin bed are suitably contained and are submerged in the pool. The sterilizer is water tight and located at the surface of the pool. The water is circulated from the pool through the prefilter, resin bed, and sterilizer by suitable pump or the like. In the preferred embodiment, the resin bed is contained within a tank which stands on the bottom of the pool and to which a base mounting the prefilter and pump is attached. An inlet for the pump is provided adjacent the bottom of the pool, while the sterilizer and outlet for the system is located adjacent the top of the pool.

  11. Purification of biomaterials by phase partitioning

    Science.gov (United States)

    Harris, J. M.

    1984-01-01

    A technique which is particularly suited to microgravity environments and which is potentially more powerful than electrophoresis is phase partitioning. Phase partitioning is purification by partitioning between the two immiscible aqueous layers formed by solution of the polymers poly(ethylene glycol) and dextran in water. This technique proved to be very useful for separations in one-g but is limited for cells because the cells are more dense than the phase solutions thus tend to sediment to the bottom of the container before reaching equilibrium with the preferred phase. There are three phases to work in this area: synthesis of new polymers for affinity phase partitioning; development of automated apparatus for ground-based separations; and design of apparatus for performing simple phase partitioning space experiments, including examination of mechanisms for separating phases in the absence of gravity.

  12. Purification and biochemical characterization of a novel glutathione ...

    African Journals Online (AJOL)

    SERVER

    2008-02-05

    dinitrobenzene as a substrate. ..... purification of GST enzyme from other insect species, which were from 3 to 26% in Hyphantria .... Glutathione S-transferases from the larval gut of the silkworm. Bombyx mori: cDNA cloning, gene ...

  13. Renaissance of protein crystallization and precipitation in biopharmaceuticals purification.

    Science.gov (United States)

    Dos Santos, Raquel; Carvalho, Ana Luísa; Roque, A Cecília A

    The current chromatographic approaches used in protein purification are not keeping pace with the increasing biopharmaceutical market demand. With the upstream improvements, the bottleneck shifted towards the downstream process. New approaches rely in Anything But Chromatography methodologies and revisiting former techniques with a bioprocess perspective. Protein crystallization and precipitation methods are already implemented in the downstream process of diverse therapeutic biological macromolecules, overcoming the current chromatographic bottlenecks. Promising work is being developed in order to implement crystallization and precipitation in the purification pipeline of high value therapeutic molecules. This review focuses in the role of these two methodologies in current industrial purification processes, and highlights their potential implementation in the purification pipeline of high value therapeutic molecules, overcoming chromatographic holdups. Copyright © 2016 Elsevier Inc. All rights reserved.

  14. Purification of heat labile toxin from Bordetella pertussis vaccine ...

    African Journals Online (AJOL)

    Purification of heat labile toxin from Bordetella pertussis vaccine strain 134 employed indigenous technology. KC Shivanandappa, S Jagannathan, S Pandiyarajan, P Tamilvanan, T Umadevi, Jeeva Kalaiselvan, B Seker ...

  15. Functionalized Solvents for Olefin Isomer Purification by Reactive Extractive Distillation

    NARCIS (Netherlands)

    Kuipers, N.J.M.; Wentink, A.E.; de Haan, A.B.; Scholtz, J.; Mulder, H.

    2007-01-01

    Olefin isomer separations are difficult, energy intensive and thus expensive. An overview is presented to investigate the feasibility of metal–ligand complexes as functionalized solvents applied in a novel separation technology, reactive extractive distillation, for the separation and purification

  16. Kinetic modelling and thermodynamic studies on purification of ...

    African Journals Online (AJOL)

    user

    adsorption mechanism might be a chemisorption process; thermodynamic studies showed that this process is spontaneous and endothermic. These results are very important in optimization of this purification process. Keywords: Polyvinylpyrrolidone (PVP), adsorption, activated carbon, resin, modelling. 1. Introduction.

  17. Economic Methods of Ginger Protease'sextraction and Purification

    Science.gov (United States)

    Qiao, Yuanyuan; Tong, Junfeng; Wei, Siqing; Du, Xinyong; Tang, Xiaozhen

    This article reports the ginger protease extraction and purification methods from fresh ginger rhizome. As to ginger protease extraction, we adapt the steps of organic solvent dissolving, ammonium sulfate depositing and freeze-drying, and this method can attain crude enzyme powder 0.6% weight of fresh ginger rhizome. The purification part in this study includes two steps: cellulose ion exchange (DEAE-52) and SP-Sephadex 50 chromatography, which can purify crude ginger protease through ion and molecular weight differences respectively.

  18. Recommended methods for purification of solvents and tests for impurities

    CERN Document Server

    Coetzee, J F

    1982-01-01

    Recommended Methods for Purification of Solvents and Tests for Impurities is a compilation of recommended procedures for purification of solvents and tests for solvent impurities. Ten solvents are covered: acetonitrile, sulfolane, propylene carbonate, dimethyl sulfoxide, dimethylformamide, hexamethylphosphoramide, pyridine, ethylenediamine, N-methylacetamide, and N-methylpropionamide. This book is comprised of 12 chapters and opens with an introduction to general aspects of impurity effects. The rationale for the selection of solvent is explained, and the relative reactivities of solutes in di

  19. A simple method for purification of herpesvirus DNA

    DEFF Research Database (Denmark)

    Christensen, Laurids Siig; Normann, Preben

    1992-01-01

    A rapid and reliable method for purification of herpesvirus DNA from cell cultures is described. The method is based on the isolation of virus particles and/or nucleocapsids by differential centrifugation and exploits the solubilizing and denaturing capabilities of cesium trifluoroacetate during...... isopycnic centrifugation, so that phenol/chloroform extractions can be omitted. The method was used for the purification of DNA from several members of the Alfaherpesvirinae subfamily....

  20. Purification and Characterization of Lipase from Aspergillus flavus ...

    African Journals Online (AJOL)

    Lipase from Aspergillus flavus was purified in a single step purification using MnFeO4 magnetic nano particles to achieve a 20.53- fold purification with specific activity of 11.29 U/mg and a 59% recovery yield. SDS-PAGE of lipase showed a single pure band with corresponding molecular weight of 35 kDa. The optimal ...

  1. Production, purification, and capsid stability of rhinovirus C types.

    Science.gov (United States)

    Griggs, Theodor F; Bochkov, Yury A; Nakagome, Kazuyuki; Palmenberg, Ann C; Gern, James E

    2015-06-01

    The rhinovirus C (RV-C) were discovered in 2006 and these agents are an important cause of respiratory morbidity. Little is known about their biology. RV-C15 (C15) can be produced by transfection of recombinant viral RNA into cells and subsequent purification over a 30% sucrose cushion, even though yields and infectivity of other RV-C genotypes with this protocol are low. The goal of this study was to determine whether poor RV-C yields were due to capsid instability, and moreover, to develop a robust protocol suitable for the purification of many RV-C types. Capsid stability assays indicated that virions of RV-C41 (refractory to purification) have similar tolerance for osmotic and temperature stress as RV-A16 (purified readily), although C41 is more sensitive to low pH. Modification to the purification protocol by removing detergent increased the yield of RV-C. Addition of nonfat dry milk to the sucrose cushion increased the virus yield but sacrificed purity of the viral suspension. Analysis of virus distribution following centrifugation indicated that the majority of detectable viral RNA (vRNA) was found in pellets refractory to resuspension. Reduction of the centrifugal force with commiserate increase in spin-time improved the recovery of RV-C for both C41 and C2. Transfection of primary lung fibroblasts (WisL cells) followed by the modified purification protocol further improved yields of infectious C41 and C2. Described herein is a higher yield purification protocol suitable for RV-C types refractory to the standard purification procedure. The findings suggest that aggregation-adhesion problems rather than capsid instability influence RV-C yield during purification. Copyright © 2015 Elsevier B.V. All rights reserved.

  2. Purification of human platelet-derived growth factor

    Energy Technology Data Exchange (ETDEWEB)

    Raines, E.W.; Ross, R.

    1985-01-01

    The paper describes a method for purification of human platelet-derived growth factor (PDGF) from outdated platelet-rich plasma (PRP) using commonly available laboratory reagents and yielding a mitogen purified 800,000-fold over the starting material. (/sup 3/H)thymidine incorporation into DNA of cultured cells responsive to PDGF represents the most readily available method to follow its purification and define the biological activity of a purified preparation. Other assays to quantitate PDGF include radioreceptor assay and radioimmunoassay.

  3. Catalysis in air purification. Katalyse in der Abluftreinigung

    Energy Technology Data Exchange (ETDEWEB)

    Fink, K.; Joisten, M.; Neufert, R.; Sprehe, J.; Zuerbig, J. (Siemens AG, Redwitz (Germany). Bereich Energieerzeugung, Keramik- und Porzellanwerk Redwitz); Gajewski, W. (Siemens AG, Erlangen (Germany). Bereich Energieerzeugung, Zentrale Forschung und Entwicklung)

    1992-05-01

    In most cases katalytic purification of waste gas allows the reliable reduction of industrial emission to meet existing regulations. This method can remove numerous organic and inorganic compounds from waste air. This review deals with design, applications and function of such air purification units with special regard to the catalysts used. Furthermore, aspects of chemical engineering relevant for planning and construction are also discussed. (orig.).

  4. Purification of N-Acetylgalactosaminidase by Isoelectric Focusing.

    Science.gov (United States)

    1986-08-01

    obtained from Joan Quay at NBL. Dr . Bong Park, a Visiting Scientist, undertook the culture of the organism and studied the early steps in purification of...matrix% The affinity agent used was originally developed by Harpaz, Flowers and Sharon (1974) for the purification of coffee bean aipha-galactosidase...collaboration with Dr % Myron Leon of the Department of Immunology and Microbiology. These radioimmunoassays had a larger than desirable background when type

  5. Development of fuzzy logic algorithm for water purification plant

    OpenAIRE

    SUDESH SINGH RANA; SUDESH SINGH RANA

    2015-01-01

    This paper propose the design of FLC algorithm for industrial application such application is water purification plant. In the water purification plant raw water or ground water is promptly purified by injecting chemical at rates related to water quality. The feed of chemical rates judged and determined by the skilled operator. Yagishita et al.[1] structured a system based on fuzzy logic so that the feed rate of the coagulant can be judged automatically without any skilled operator. We perfor...

  6. Purification of cerium, neodymium and gadolinium for low background experiments

    Directory of Open Access Journals (Sweden)

    Boiko R.S.

    2014-01-01

    Full Text Available Cerium, neodymium and gadolinium contain double beta active isotopes. The most interesting are 150Nd and 160Gd (promising for 0ν2β search, 136Ce (2β+ candidate with one of the highest Q2β. The main problem of compounds containing lanthanide elements is their high radioactive contamination by uranium, radium, actinium and thorium. The new generation 2β experiments require development of methods for a deep purification of lanthanides from the radioactive elements. A combination of physical and chemical methods was applied to purify cerium, neodymium and gadolinium. Liquid-liquid extraction technique was used to remove traces of Th and U from neodymium, gadolinium and for purification of cerium from Th, U, Ra and K. Co-precipitation and recrystallization methods were utilized for further reduction of the impurities. The radioactive contamination of the samples before and after the purification was tested by using ultra-low-background HPGe gamma spectrometry. As a result of the purification procedure the radioactive contamination of gadolinium oxide (a similar purification efficiency was reached also with cerium and neodymium oxides was decreased from 0.12 Bq/kg to 0.007 Bq/kg in 228Th, from 0.04 Bq/kg to <0.006 Bq/kg in 226Ra, and from 0.9 Bq/kg to 0.04 Bq/kg in 40K. The purification methods are much less efficient for chemically very similar radioactive elements like actinium, lanthanum and lutetium.

  7. TMI-2 purification demineralizer resin study

    Energy Technology Data Exchange (ETDEWEB)

    Thompson, J D; Osterhoudt, T R

    1984-05-01

    Study of the Makeup and Purification System demineralizers at TMI-2 has established that fuel quantities in the vessels are low, precluding criticality, that the high radioactive cesium concentration on the demineralizer resins can be chemically removed, and that the demineralizer resins can probably be removed from the vessels by sluicing through existing plant piping. Radiation measurements from outside the demineralizers establishing that there is between 1.5 and 5.1 (probably 3.3) lb of fuel in the A vessel and less than that amount in the B vessel. Dose rates up to 2780 R per hour were measured on contact with the A demineralizer. Remote visual observation of the A demineralizer showed a crystalline crust overlaying amber-colored resins. The cesium activity in solid resin samples ranged from 220 to 16,900 ..mu..Ci/g. Based on this information, researchers concluded that the resins cannot be removed through the normal pathway in their present condition. Studies do show that the resins will withstand chemical processing designed to rinse and elute cesium from the resins. The process developed should work on the TMI-2 resins.

  8. Purification and electron cryomicroscopy of coronavirus particles.

    Science.gov (United States)

    Neuman, Benjamin W; Adair, Brian D; Yeager, Mark; Buchmeier, Michael J

    2008-01-01

    Intact, enveloped coronavirus particles vary widely in size and contour, and are thus refractory to study by traditional structural means such as X-ray crystallography. Electron microscopy (EM) overcomes some problems associated with particle variability and has been an important tool for investigating coronavirus ultrastructure. However, EM sample preparation requires that the specimen be dried onto a carbon support film before imaging, collapsing internal particle structure in the case of coronaviruses. Moreover, conventional EM achieves image contrast by immersing the specimen briefly in heavy-metal-containing stain, which reveals some features while obscuring others. Electron cryomicroscopy (cryo-EM) instead employs a porous support film, to which the specimen is adsorbed and flash-frozen. Specimens preserved in vitreous ice over holes in the support film can then be imaged without additional staining. Cryo-EM, coupled with single-particle image analysis techniques, makes it possible to examine the size, structure and arrangement of coronavirus structural components in fully hydrated, native virions. Two virus purification procedures are described.

  9. Production, purification and properties of microbial phytases.

    Science.gov (United States)

    Pandey, A; Szakacs, G; Soccol, C R; Rodriguez-Leon, J A; Soccol, V T

    2001-05-01

    Phytases (myo-inositol hexakisphosphate phosphohydrolase, EC 3.1.3.8) catalyse the release of phosphate from phytate (mycoinositol hexakiphosphate). Several cereal grains, legumes and oilseeds, etc., store phosphorus as phytate. Environmental pollution due to the high-phosphate manure, resulting in the accumulation of P at various locations has raised serious concerns. Phytases appear of significant value in effectively controlling P pollution. They can be produced from a host of sources including plants, animals and micro-organisms. Microbial sources, however, are promising for their commercial exploitations. Strains of Aspergillus sp., chiefly A. ficuum and A. niger have most commonly been employed for industrial purposes. Phytases are considered as a monomeric protein, generally possessing a molecular weight between 40 and 100 kDa. They show broad substrate specificity and have generally pH and temperature optima around 4.5-6.0 and 45-60 degrees C. The crystal structure of phytase has been determined at 2.5 A resolution. Immobilization of phytase has been found to enhance its thermostability. This article reviews recent trends on the production, purification and properties of microbial phytases.

  10. Variant Purification of an Allogeneic Bone Block

    Science.gov (United States)

    Lorenz, Jonas; Schlee, Markus; Al-Maawi, Sarah; Chia, Poju; Sader, Robert A.

    2017-01-01

    Objective This short communication reports on a histological analysis of the composition of the commercially available Maxgraft® allogeneic bone block. Materials and Methods Based on previously published, easily applicable histological methods, blanc samples of the Maxgraft® allogeneic bone block have been decalcified, dehydrated and embedded in paraffin before histological and histochemical staining. Afterwards, the slides were evaluated for their material characteristics, such as the bone matrix structure and other components, including collagen or cells/cell remnants. Results The results show that this bone block exhibits a trabecular structure with lamellar sub-organization. Additionally, cellular remnants within the osteocyte lacunae and at the outer trabecular surfaces reside together with remnants of the former inter-trabecular fatty and connective tissue, i.e., collagenous structures and connective tissue cells or cell remnants. Conclusion Consistent with a previous study on this topic, the data presented here demonstrate that some of the certified purification techniques might not allow for the production of allogeneic materials free of organic cell and tissue components. PMID:28827851

  11. Extraction, purification and characterisation of fulvic acid

    Energy Technology Data Exchange (ETDEWEB)

    Higgo, J.J.W.; Davis, J.R.; Smith, B.; Milne, C.

    1998-04-01

    The aim of this project was to extract and purify gram quantities of fulvic acid for distribution to the group members. Ideally the material should have come from groundwater in the Sellafield repository area but the total organic carbon content of water samples taken from boreholes in the area was found to be between 0.03 and 0.05 ppm. In order to extract sufficient fulvic for distribution to group members it would, therefore, be necessary to process more than 100 000 litres of groundwater. This was considered impractical and instead fulvic acid was extracted from four thousand litres of water taken from the Derwent Reservoir (Derbyshire, UK) using DEAE-cellulose. Several stages of purification included anion exchange on Amberlite XAD-8 resin and cation exchange on Amberlite AG50W-X8. The final purified fulvic acid concentrate, volume 1300 ml, contained 4.1 g/l TOC (total fulvic acid [approx] 11 g). It was characterised in terms of elemental analysis, molecular weight, proton-binding properties, UV absorbance. Uranium binding measurements were also made for comparison with previous work. (author)

  12. Extraction, purification and characterization of fulvic acid

    Energy Technology Data Exchange (ETDEWEB)

    Higgo, J.J.W.; Davies, J.R.; Smith, B.; Milne, C. [British Geological Survey, Nottingham (United Kingdom)

    1998-08-01

    The aim of this project was to extract and purify gram quantities of fulvic acid for distribution to the group members. Ideally the material should have come from groundwater in the Sellafield repository area but the total organic carbon content of water samples taken from boreholes in the area was found to be between 0.03 and 0.05 ppm. In order to extract sufficient fulvic for distribution to group members it would, therefore, be necessary to process more than 100 000 litres of groundwater. This was considered impractical and instead fulvic acid was extracted from four thousand litres of water taken from the Derwent Reservoir (Derbyshire, UK) using DEAE-cellulose. Several stages of purification included anion exchange on Amberlite XAD-8 resin and cation exchange on Amberlite AG50W-X8. The final purified fulvic acid concentrate, volume 1300 ml, contained 4.1 g/l TOC (total fulvic acid {approx}11 g). It was characterised in terms of elemental analysis, molecular weight, proton-binding properties, UV absorbance. Uranium binding measurements were also made for comparison with previous work. (orig.)

  13. Escherichia coli enterotoxin. Purification and partial characterization.

    Science.gov (United States)

    Dorner, F

    1975-11-25

    Enterotoxin, a diarrheagenic protein elaborated by pathogenic Escherichia coli strains has been isolated from the supernatant of fermenter cultures of E. coli strain P263, a porcine enteropathogen. Purification steps involving Bio-Gel agarose A-5m, Sephadex G-75 chromatography, and preparative isotachophoresis were used in the isolation. The resulting product appears to be pure according to immunoelectrophoretic, disc electrophoretic, ultracentrifugal, and immunologic criteria. The entertoxin has an apparent molecular weight of 102,000 as judged by gel filtration and sodium dodecyl sulfate polyacrylamide gel electrophoresis, and its isoelectric point is 6.90. The isolated product is highly active in inducing experimental diarrhea in adult rabbits and piglets. It also elicits, in small dosage, a marked increase in adenylate cyclase activity in broken cell preparations of cat heart tissue. The enterotoxin activity is acid-labile and is destroyed by heating at 65 degrees for 30 min. It is suggested that the heat-stable enterotoxin material is derived from heat-labile enterotoxin by forming a complex with endotoxin or capsular material present in the culture supernatant.

  14. Purification and characterization of canine intestinal motilin.

    Science.gov (United States)

    Poitras, P; Reeve, J R; Hunkapiller, M W; Hood, L E; Walsh, J H

    1983-02-01

    A 22 amino acid peptide with motilin-like immunoreactivity was purified from acetic acid extracts of small intestinal mucosa from mongrel dogs. Sequential chromatography on carboxymethyl-cellulose, Sephadex G-50, CM cellulose, Biogel P6, and two steps of high-performance liquid chromatography were used for purification. Microsequence analysis of the purified product permitted unambiguous identification of residues 2-22 as -VPIFTHSELQKIREKERNKGQ. The sequence of porcine intestinal motilin is FVPIFTYGELQRMQEKERNKGQ. The amino terminal residue of the canine peptide could not be assigned with certainty since Phe, Lys, and Ser all were identified by analysis of PTH derivatives on the first sequencing cycle. Definite amino acid differences between canine and porcine motilin thus were identified in positions 7, 8, 12, 13 and 14. These differences did not alter immunoreactivity of canine motilin with antibodies specific for the carboxyl-terminal portion of porcine motilin, but probably explain markedly diminished immunoreactivity with antibodies to the amino or mid-portion of porcine motilin. Synthetic Phe1 canine motilin was prepared by a solid phase method. The synthetic peptide had the same pattern of immunoreactivity as natural canine motilin and was biologically active with a potency similar to synthetic porcine motilin for induction of premature activity fronts of the interdigestive motor complex in the small intestine of fasting dogs.

  15. Myxoma virus: propagation, purification, quantification, and storage.

    Science.gov (United States)

    Smallwood, Sherin E; Rahman, Masmudur M; Smith, Dorothy W; McFadden, Grant

    2010-05-01

    Myxoma virus (MYXV) is a member of the Poxviridae family and prototype for the genus Leporipoxvirus. It is pathogenic only for European rabbits, in which it causes the lethal disease myxomatosis, and two North American species, in which it causes a less severe disease. MYXV replicates exclusively in the cytoplasm of the host cell. Although not infectious in humans, its genome encodes proteins that can interfere with or modulate host defense mechanisms; it is able to productively infect a number of human cancer cell lines, but not normal human cells, and has also been shown to increase survival time in mouse models of human glioma. These characteristics suggest that MYXV could be a viable therapeutic agent, e.g., in anti-inflammatory or anti-immune therapy, or as an oncolytic agent. MYXV is also an excellent model for poxvirus biology, pathogenesis, and host tropism studies. It is easily propagated in a number of cell lines, including adherent cells and suspension cultures, and minimal purification is required to provide a stock for in vivo and in vitro studies.

  16. Purification of transthyretin as nutritional biomarker of selenium status.

    Science.gov (United States)

    Mahn, Andrea; Lienqueo, María Elena; Quilodrán, Claudia; Olivera-Nappa, Alvaro

    2012-11-01

    Transthyretin has been proposed as nutritional biomarker of selenium intake. Previous transthyretin purification methods used different procedures to isolate transthyretin either from plasma or from pathological urine of humans. In general, the procedure for purification of transthyretin is laborious and expensive, and extensive sample recycling is necessary for purification in appreciable amounts. This work proposes a new, promissory, and cheap two-step process to purify transthyretin from blood plasma, composed by a first aqueous two-phase system fractionation followed by affinity chromatography, using thyroxine-immobilized on epoxy-activated Sepharose CL-6B. The aqueous two-phase system fractionation was demonstrated to perform better than commercial immunoaffinity-based kits for albumin depletion in blood plasma samples and is an effective first step for transthyretin purification. Thyroxine affinity chromatography was designed to bind transthyretin with high affinity, and was demonstrated to be useful to purify transthyretin, but was unable to completely resolve transthyretin from thyroxine-binding globulin and serum albumin, although the relative amount of albumin was lowered in the eluates. This purification process could be used in nutritional diagnosis tools or as a first step in structural and functional studies. © 2012 WILEY-VCH Verlag GmbH & Co. KGaA, Weinheim.

  17. Oestrogen, testosterone, cytotoxin and cholinesterase inhibitor ...

    African Journals Online (AJOL)

    Namibia is the driest sub-Saharan country in Africa. Namibia's capital, Windhoek, reclaims sewage water for domestic use at the Goreangab Water Reclamation Plant (GWRP). Risks associated with sewage effluent and reclaimed sewage should be closely monitored; therefore water at the Gammams Sewage Treatment ...

  18. Oestrogen, testosterone, cytotoxin and cholinesterase inhibitor ...

    African Journals Online (AJOL)

    2013-07-08

    Jul 8, 2013 ... removal during reclamation of sewage to drinking water. AK Faul1,2, E Julies1, and ... closely monitored; therefore water at the Gammams Sewage Treatment Plant (GSTP) inlet and outlet, as well as reclaimed water from the ... led to the upgrade of the conventional water treatment plant in the vicinity of the ...

  19. Carbon Nanotube Membranes for Water Purification

    Science.gov (United States)

    Bakajin, Olgica

    2009-03-01

    Carbon nanotubes are an excellent platform for the fundamental studies of transport through channels commensurate with molecular size. Water transport through carbon nanotubes is also believed to be similar to transport in biological channels such as aquaporins. I will discuss the transport of gas, water and ions through microfabricated membranes with sub-2 nanometer aligned carbon nanotubes as ideal atomically-smooth pores. The measured gas flow through carbon nanotubes exceeded predictions of the Knudsen diffusion model by more than an order of magnitude. The measured water flow exceeded values calculated from continuum hydrodynamics models by more than three orders of magnitude and is comparable to flow rates extrapolated from molecular dynamics simulations and measured for aquaporins. More recent reverse osmosis experiments reveal ion rejection by our membranes. Based on our experimental findings, the current understanding of the fundamentals of water and gas transport and of ion rejection will be discussed. The potential application space that exploits these unique nanofluidic phenomena will be explored. The extremely high permeabilities of these membranes, combined with their small pore size will enable energy efficient filtration and eventually decrease the cost of water purification.[4pt] In collaboration with Francesco Fornasiero, Biosciences and Biotechnology Division, PLS, LLNL, Livermore, CA 94550; Sangil Kim, NSF Center for Biophotonics Science & Technology, University of California at Davis, Sacramento CA 95817; Jung Bin In, Mechanical Engineering Department, UC Berkeley, Berkeley CA 94720; Hyung Gyu Park, Jason K Holt, and Michael Stadermann, Biosciences and Biotechnology Division, PLS, LLNL; Costas P. Grigoropoulos, Mechanical Engineering Department, UC Berkeley; Aleksandr Noy, Biosciences and Biotechnology Division, PLS, LLNL and School of Natural Sciences, University of California at Merced.

  20. Affinity purification of Car9-tagged proteins on silica matrices: Optimization of a rapid and inexpensive protein purification technology.

    Science.gov (United States)

    Soto-Rodríguez, Jessica; Coyle, Brandon L; Samuelson, Ariana; Aravagiri, Kannan; Baneyx, François

    2017-07-01

    Car9, a dodecapeptide identified by cell surface display for its ability to bind to the edge of carbonaceous materials, also binds to silica with high affinity. The interaction can be disrupted with l-lysine or l-arginine, enabling a broad range of technological applications. Previously, we reported that C-terminal Car9 extensions support efficient protein purification on underivatized silica. Here, we show that the Car9 tag is functional and TEV protease-excisable when fused to the N-termini of target proteins, and that it supports affinity purification under denaturing conditions, albeit with reduced yields. We further demonstrate that capture of Car9-tagged proteins is enhanced on small particle size silica gels with large pores, that the concomitant problem of nonspecific protein adsorption can be solved by lysing cells in the presence of 0.3% Tween 20, and that efficient elution is achieved at reduced l-lysine concentrations under alkaline conditions. An optimized small-scale purification kit incorporating the above features allows Car9-tagged proteins to be inexpensively recovered in minutes with better than 90% purity. The Car9 affinity purification technology should prove valuable for laboratory-scale applications requiring rapid access to milligram-quantities of proteins, and for preparative scale purification schemes where cost and productivity are important factors. Copyright © 2017 Elsevier Inc. All rights reserved.

  1. Purification and fluorescent labeling of the human serotonin transporter

    DEFF Research Database (Denmark)

    Rasmussen, Søren G F; Gether, Ulrik

    2005-01-01

    To establish a purification procedure for the human serotonin transporter (hSERT) we expressed in Sf9 insect cells an epitope-tagged version of the transporter containing a FLAG epitope at the N-terminus and a polyhistidine tail at the C-terminus (FLAG-hSERT-12H). For purification, the transporter...... was solubilized in digitonin followed by nickel affinity and subsequent concanavalin A chromatography. Using this procedure we were able to obtain an overall purification of 700-fold and a yield of approximately 0.1 mg/L of cell culture. The purified transporter displayed pharmacological properties similar...... quenching experiments revealed that the aqueous quencher iodide was able to cause marked quenching of the fluorescence of the IANBD labeled transporter with a K(SV) of 3.4 +/- 0.10 M(-)(1). In a mutant transporter with five cysteines mutated (5CysKO) we observed a significant reduction in this quenching (K...

  2. Bromelain: an overview of industrial application and purification strategies.

    Science.gov (United States)

    Arshad, Zatul Iffah Mohd; Amid, Azura; Yusof, Faridah; Jaswir, Irwandi; Ahmad, Kausar; Loke, Show Pau

    2014-09-01

    This review highlights the use of bromelain in various applications with up-to-date literature on the purification of bromelain from pineapple fruit and waste such as peel, core, crown, and leaves. Bromelain, a cysteine protease, has been exploited commercially in many applications in the food, beverage, tenderization, cosmetic, pharmaceutical, and textile industries. Researchers worldwide have been directing their interest to purification strategies by applying conventional and modern approaches, such as manipulating the pH, affinity, hydrophobicity, and temperature conditions in accord with the unique properties of bromelain. The amount of downstream processing will depend on its intended application in industries. The breakthrough of recombinant DNA technology has facilitated the large-scale production and purification of recombinant bromelain for novel applications in the future.

  3. Submerged type water purification system using Hollow fiber Microfiltration membrane

    Energy Technology Data Exchange (ETDEWEB)

    Chung, Kyu-Young [Genix Engineering, Seoul (Korea); Kim, Hyung-Soo [Sung Kyun Kwan University, Suwon (Korea); Im, Jong-Sung [Kumho Industrial Company, Seoul (Korea)

    1999-06-30

    Membrane separation process is considered as an alternative of conventional water purification system using coagulation - sedimentation - sand filtration. In this study, it was examined that the application possibility of Hollowfiber Microfiltration membrane for water purification process. A 20 m{sup 3}/day scale pilot plant was used for studying the possibility of long-term operation and the stability of water quality under the optimum condition, 0.03 m/h permeate flux, filtration for 10 minutes, pause for 2 minutes (including air-scrubbing for 30 seconds), obtained by lab-scale experiment. As a result, it was proved stability of pilot plant over one year and filtrate quality(Turbidity, SS etc.). Therefore, it was proved that membrane separation process using Hollowfiber Microfiltration membrane can be applied for water purification system. (author). 13 refs., 3 tabs., 16 figs.

  4. 24 CFR 203.52 - Acceptance of individual residential water purification equipment.

    Science.gov (United States)

    2010-04-01

    ... residential water purification equipment. 203.52 Section 203.52 Housing and Urban Development Regulations... water purification equipment. If a property otherwise eligible for insurance under this part does not have access to a continuing supply of safe and potable water without the use of a water purification...

  5. 21 CFR 884.6170 - Assisted reproduction water and water purification systems.

    Science.gov (United States)

    2010-04-01

    ... 21 Food and Drugs 8 2010-04-01 2010-04-01 false Assisted reproduction water and water purification... Devices § 884.6170 Assisted reproduction water and water purification systems. (a) Identification. Assisted reproduction water purification systems are devices specifically intended to generate high quality...

  6. Squeezed-state purification with linear optics and feedforward.

    Science.gov (United States)

    Glöckl, O; Andersen, U L; Filip, R; Bowen, W P; Leuchs, G

    2006-08-04

    A scheme for optimal and deterministic linear optical purification of mixed squeezed Gaussian states is proposed and experimentally demonstrated. The scheme requires only linear optical elements and homodyne detectors, and allows the balance between purification efficacy and squeezing degradation to be controlled. One particular choice of parameters gave a tenfold reduction of the thermal noise with a corresponding squeezing degradation of only 11%. We prove optimality of the protocol, and show that it can be used to enhance the performance of quantum informational protocols such as dense coding and entanglement generation.

  7. Solid support resins and affinity purification mass spectrometry.

    Science.gov (United States)

    Havis, Spencer; Moree, Wilna J; Mali, Sujina; Bark, Steven J

    2017-02-28

    Co-affinity purification-mass spectrometry (CoAP-MS) is a primary technology for elucidating the protein-protein interactions that form the basis of all biological processes. A critical component of CoAP-MS is the affinity purification (AP) of the bait protein, usually by immobilization of an antibody to a solid-phase resin. This Minireview discusses common resins, reagents, tagging methods, and their consideration for successful AP of tagged proteins. We discuss our experiences with different solid supports, their impact in AP experiments, and propose areas where chemistry can advance this important technology.

  8. Expression and purification of recombinant nattokinase in Spodoptera frugiperda cells.

    Science.gov (United States)

    Li, Xiaoxiang; Wang, Xiaoli; Xiong, Shaoling; Zhang, Jing; Cai, Litao; Yang, Yanyan

    2007-10-01

    A recombinant baculovirus, rv-egfp-NK, containing a reporter gene encoding the enhanced green fluorescent protein (EGFP), was used to express nattokinase (NK), a fibrinolytic enzyme, in Spodoptera frugiperda (SF-9) cells. The recombinant protein also included a histidine tag for purification using Ni(2+) resins. The recombinant NK, approximately 30 kDa, retained fibrinolytic activity (60 U/ml). The integration of the EGFP expression cassette in the Bac-to-Bac system is thus an effective method for the expression and purification of recombinant NK protein in Spodoptera frugiperda insect cells.

  9. High quality protein microarray using in situ protein purification

    Directory of Open Access Journals (Sweden)

    Fleischmann Robert D

    2009-08-01

    Full Text Available Abstract Background In the postgenomic era, high throughput protein expression and protein microarray technologies have progressed markedly permitting screening of therapeutic reagents and discovery of novel protein functions. Hexa-histidine is one of the most commonly used fusion tags for protein expression due to its small size and convenient purification via immobilized metal ion affinity chromatography (IMAC. This purification process has been adapted to the protein microarray format, but the quality of in situ His-tagged protein purification on slides has not been systematically evaluated. We established methods to determine the level of purification of such proteins on metal chelate-modified slide surfaces. Optimized in situ purification of His-tagged recombinant proteins has the potential to become the new gold standard for cost-effective generation of high-quality and high-density protein microarrays. Results Two slide surfaces were examined, chelated Cu2+ slides suspended on a polyethylene glycol (PEG coating and chelated Ni2+ slides immobilized on a support without PEG coating. Using PEG-coated chelated Cu2+ slides, consistently higher purities of recombinant proteins were measured. An optimized wash buffer (PBST composed of 10 mM phosphate buffer, 2.7 mM KCl, 140 mM NaCl and 0.05% Tween 20, pH 7.4, further improved protein purity levels. Using Escherichia coli cell lysates expressing 90 recombinant Streptococcus pneumoniae proteins, 73 proteins were successfully immobilized, and 66 proteins were in situ purified with greater than 90% purity. We identified several antigens among the in situ-purified proteins via assays with anti-S. pneumoniae rabbit antibodies and a human patient antiserum, as a demonstration project of large scale microarray-based immunoproteomics profiling. The methodology is compatible with higher throughput formats of in vivo protein expression, eliminates the need for resin-based purification and circumvents

  10. Purification of highly chlorinated dioxins degrading enzyme

    Energy Technology Data Exchange (ETDEWEB)

    Ishii, K.; Furuichi, T.; Koike, K.; Kuboshima, M. [Hokkaido Univ. (Japan). Division of Environment Resource Engineering, Graduate School of Engineering

    2004-09-15

    Soil contamination caused by dioxins in and around sites of incinerators for municipal solid waste (MSW) is a concern in Japan. For example, scattering wastewater from a wet gas scrubber at an MSW incinerator facility in Nose, Osaka caused soil and surface water contamination. The concentration of dioxins in the soil was about 8,000 pg-TEQ/g. Other contamination sites include soils on which fly ash has been placed directly or improperly stored and landfill sites that have received bottom and fly ash over a long period. Some countermeasures are required immediately at these dioxins-contaminated sites. We have previously developed bioreactor systems for dioxin-contaminated water and soil. We have shown that a fungus, Pseudallescheria boydii (P. boydii), isolated from activated sludge treating wastewater that contained dioxins, has the ability to degrade highly chlorinated dioxins. A reaction product of octachlorinated dibenzo-p-dioxin (OCDD) was identified as heptachlorinated dibenzo-p-dioxin. Therefore, one of the pathways for degradation of OCDD by this fungus was predicted to be as follows: OCDD is transformed by dechlorination and then one of the remaining aromatic rings is oxidized. To apply P. boydii to on-site technologies (e.g., bioreactor systems), as well as in situ technologies, enzyme treatment using a dioxin-degrading enzyme from P. boydii needs to be developed because P. boydii is a weak pathogenic fungus, known to cause opportunistic infection. As a result, we have studied enzyme purification of nonchlorinated dioxin, namely, dibenzo-pdioxin (DD). However, we did not try to identify enzymes capable of degrading highly chlorinated dioxins. This study has elucidated a method of enzyme assay for measuring OCDD-degrading activity, and has attempted to purify OCDD-degrading enzymes from P. boydii using enzyme assay. In addition, as first step toward purifying 2,3,7,8-tetrachlorodibenzo-p-dioxin (2,3,7,8-TCDD), 2,3,7,8-TCDD degradation tests were carried out

  11. Effect of the purification of antidermatophytic proteins from Nigella ...

    African Journals Online (AJOL)

    17 amino acids were found in the four tested plant proteins with N. sativa protein having the highest content of amino acid (347.21 μg/ml). ... Complete purification of the most active fraction (Pr2) on diethylaminoethyl (DEAE)-Sephadex eluted one single peak with increasing antidermatophytic activity (7.6 cm) against the ...

  12. New purification process of fungal immunomodulatory protein, FIP ...

    African Journals Online (AJOL)

    cy l

    protein, FIP-fve from Flammulina velutipes via filtration and column .... electrophoresis (SDS-PAGE). The practice is shown in the following. Figure 1. Cao et al. 513. Downstream protein purification by ultrafiltration concentration. The clear liquor ..... Liao CH, Hsiao YM, Hsu CP, Lin MY, Wang JC, Huang YL, Ko JL. (2006).

  13. Purification of cadmium up to 5N+ by vacuum distillation

    Indian Academy of Sciences (India)

    Cadmium was refined by vacuum distillation, a technique suitable for low boiling and melting point materials, to remove the heavy and low vapour pressure impurities at ppm level. The detailed analysis of the purified Cd as well as raw Cd was done by ICP–OES techniques for 27 impurity elements. Purification was carried ...

  14. The purification and characterisation of allergenic hazelnut seed proteins

    NARCIS (Netherlands)

    Rigby, Neil M.; Marsh, Justin; Sancho, Ana I.; Wellner, Klaus; Akkerdaas, Jaap; van Ree, Ronald; Knulst, Andre; Fernández-Rivas, Montserrat; Brettlova, Vlasta; Schilte, Piet P.; Summer, Colin; Pumphrey, Richard; Shewry, Peter R.; Mills, E. N. Clare

    2008-01-01

    A lipid transfer protein (LTP, Cor a 8) together with the 11S (Cor a 9) and 7S seed storage globulins (Cor a 11) are major food allergens present in hazelnut. Methods are described for their purification and characterisation using in-gel tryptic digestion mass spectrometry to confirm their

  15. Production and purification of polyclonal anti-hamster ...

    African Journals Online (AJOL)

    Polyclonal antibodies are mixtures of monoclonal antibodies that were produced against different epitops. The goal of this project is to know the production, purification and horseradish peroxidase (HRP) conjugation of polyclonal antibodies against hamster immunoglobulins in rabbits. 300 ìg/300 ìl of ten hamster ...

  16. Fungal laccase: copper induction, semi-purification, immobilization ...

    African Journals Online (AJOL)

    Semi-purification of the crude laccase was carried out through precipitation and the column separation with NaCl gradient. In order to apply in an effluent treatment, laccase was immobilized on different vitroceramics supports, pyrolytic graphite and also on a carbon fiber electrode as biosensor. The maximum laccase activity ...

  17. Extraction, Purification, and Spectroscopic Characterization of a Mixture of Capsaicinoids

    Science.gov (United States)

    Wagner, Carl E.; Cahill, Thomas M.; Marshall, Pamela A.

    2011-01-01

    This laboratory experiment provides a safe and effective way to instruct undergraduate organic chemistry students about natural-product extraction, purification, and NMR spectroscopic characterization. On the first day, students extract dried habanero peppers with toluene, perform a pipet silica gel column to separate carotenoids from…

  18. Purification and deodorization of structured lipids by short path distillation

    DEFF Research Database (Denmark)

    Xu, Xuebing; Jacobsen, Charlotte; Nielsen, Nina Skall

    2002-01-01

    Purification of structured lipids (SL), produced from lipase- catalyzed acidolysis of rapeseed oil and capric acid, and deodorization of randomized SL, produced from chemical randomization of fish oil and tricaprin, were studied in a bench-scale short path distillation (SPD). SL obtained from...

  19. Extraction and Purification of Flavonoids from Radix Puerariae | Li ...

    African Journals Online (AJOL)

    Purpose: To develop an efficient method for the purification of flavonoids from Radix puerariae. Methods: Optimal extraction technology was obtained using orthogonal test. Through adsorption and desorption tests, 8 resins with different polarity, diameter, and surface area were studied. Finally, a novel macroporous resin, ...

  20. Turnkey Helium Purification and Liquefaction Plant for DARWIN, Australia

    Science.gov (United States)

    Lindemann, U.; Boeck, S.; Blum, L.; Kurtcuoglu, K.

    2010-04-01

    The Linde Group, through its Australian subsidiary BOC Limited, has signed an agreement with Darwin LNG Pty Ltd for the supply of feed-gas to Linde's new helium refining and liquefaction facility in Darwin, Australia. Linde Kryotechnik AG, located in Switzerland, has carried out the engineering and fabrication of the equipment for the turn key helium plant. The raw feed gas flow of 20'730 Nm3/h contains up to of 3 mol% helium. The purification process of the feed gas consists of partial condensation of nitrogen in two stages, cryogenic adsorption and finally catalytic oxidation of hydrogen followed by a dryer system. Downstream of the purification the refined helium is liquefied using a modified Bryton process and stored in a 30'000 gal LHe tank. For further distribution and export of the liquid helium there are two stations available for filling of truck trailers and containers. The liquid nitrogen, required for refrigeration capacity to the nitrogen removal stages in the purification process as well as for the pre-cooling of the pure helium in the liquefaction process, is generated on site during the feed gas purification process. The optimized process provides low power consumption, maximum helium recovery and a minimum helium loss.

  1. Development of Purification Protocol Specific for Bacteriocin 105B

    Science.gov (United States)

    2017-02-09

    S.M Lemon, M. Najafi, et al., editors; “The Resistance Phenomenon in Microbes and Infectious Disease Vectors: Implications for Human Health and...Bacillus anthracis. As the current application of broad-spectrum antimicrobials promotes the development of multi- drug resistant microorganisms...SPECTRUM TARGETED ANTIMICROBIALS ASSAYS PURIFICATION BACILLUS ANTHRACIS DRUG- RESISTANT MICROORGANISMS

  2. Purification and Some Properties of Rubber Seed Lipoxygenase ...

    African Journals Online (AJOL)

    Lipoxygenase was extracted from rubber seed and purified by ammonium sulphate precipitation, gel filtration on Sephadex G – 25 and ion – exchange chromatographies on DEAE – cellulose columns. The enzyme was purified 78 fold and 38.56% of the enzyme activity was recovered in the purification. The molecular ...

  3. Production and purification of polyclonal antibody against bovine ...

    African Journals Online (AJOL)

    Antibodies are important tools in medical researches which have led to many advances in this field. Anti-bovine immunoglobulins and its conjugate with horse radish peroxidase (HRP) is used to diagnose cows' disease by ELISA or western blotting tests. In this study, the production, purification and horse radish peroxidase ...

  4. Expression, purification and preliminary diffraction studies of PhnP

    DEFF Research Database (Denmark)

    Podzelinska, Kateryna; He, Shumei; Soares, Alexei

    2008-01-01

    PhnP belongs to a 14-gene operon that supports the growth of Escherichia coli on alkylphosphonates as a sole source of phosphorus; however, the exact biochemistry of phosphonate degradation by this pathway is poorly understood. The protein was recombinantly expressed in Escherichia coli and purif...

  5. The Partial Purification and Characterization of Lactate Dehydrogenase.

    Science.gov (United States)

    Wolf, Edward C.

    1988-01-01

    Offers several advantages over other possibilities as the enzyme of choice for a student's first exposure to a purification scheme. Uses equipment and materials normally found in biochemistry laboratories. Incorporates several important biochemical techniques including spectrophotometry, chromatography, centrifugation, and electrophoresis. (MVL)

  6. Ligand-modified metal clusters for gas separation and purification

    Science.gov (United States)

    Okrut, Alexander; Ouyang, Xiaoying; Runnebaum, Ron; Gates, Bruce C.; Katz, Alexander

    2017-02-21

    Provided is an organic ligand-bound metal surface that selects one gaseous species over another. The species can be closely sized molecular species having less than 1 Angstrom difference in kinetic diameter. In one embodiment, the species comprise carbon monoxide and ethylene. Such organic ligand-bound metal surfaces can be successfully used in gas phase separations or purifications, sensing, and in catalysis.

  7. Production, purification and partial characterization of lipase from ...

    African Journals Online (AJOL)

    Production, purification and partial characterization of lipase from Trichoderma Viride. MA Kashmiri, A Adnan, BW Butt. Abstract. A new strain of lipolytic Trichoderma viride was isolated from the soil on a selective mediumthat contained olive oil as the only source of carbon and energy. The isolated strain was cultivated for ...

  8. A study on the extraction and purification process of lily ...

    African Journals Online (AJOL)

    The objective of this paper was to extract and purify lily polysaccharide and to study its anti-H22 hepatoma effect in mice. Orthogonal experimental method was used to analyze the factors influencing the extraction and purification of lily polysaccharide, and the anti-tumor effect of lily polysaccharide was studied by acting it on ...

  9. Purification and characterization of a peroxidase present in ...

    African Journals Online (AJOL)

    In this work, we report the purification of a peroxidase (POX) from umbu xilopodium exsudate by direct extraction from polyacrylamide electrophoresis gels. ... When assayed with metal ions, umbu POX activity was shown to be inhibited by Mn2+ and stimulated by Ca2+ and Mg2+; it was also inhibited by sodium azide ...

  10. Novel heat-pump-assisted extractive distillation for bioethanol purification

    NARCIS (Netherlands)

    Luo, Hao; Bildea, Costin Sorin; Kiss, Anton A.

    2015-01-01

    The purification of bioethanol fuel involves an energy-intensive separation process to concentrate the diluted streams obtained in the fermentation stage and to overcome the azeotropic behavior of the ethanol-water mixture. The conventional separation sequence employs three distillation columns that

  11. Energy Efficient Bioethanol Purification by Heat Pump Assisted Extractive Distillation

    NARCIS (Netherlands)

    Kiss, Anton A.; Luo, Hao; Bildea, Costin Sorin

    2015-01-01

    The purification of bioethanol fuel requires an energy demanding separation process to concentrate the diluted streams obtained in the fermentation stage and to overcome the azeotropic behaviour of ethanol-water mixture. The classic separation sequence consists of three distillation columns that

  12. Purification of kappa (k)-carrageenase from locally isolated ...

    African Journals Online (AJOL)

    Ehab Beltagy

    2012-06-28

    Jun 28, 2012 ... Key words: Cellulosimicrobium cellulans, k-carrageenase, purification, sephadex G-100, diethylaminoethyl. (DEAE) sephadex A-52. ... been found in several marine bacteria (Bellion et al.,. 1982). Many bacterial ... ingredients (g/l): filtered sea water 800 ml; distilled water 200 ml; yeast extract, 1; peptone, 5; ...

  13. Characterization and partial purification of phospholipase D from human placenta

    DEFF Research Database (Denmark)

    Vinggaard, Anne Marie; Hansen, Harald S.

    1995-01-01

    . The PLD activity was enriched 15-fold by solubilization and purification on a DEAE-Sepharose column. N-Ethylmaleimide (10 mM) markedly inhibited the purified enzyme, indicating the presence of free thiol groups on PLD. Sphingosine (20) µM) and (±) propranolol (53 µM) had no direct effect on PLD activity...

  14. Purification and characterization of angiotensin-1 converting enzyme

    African Journals Online (AJOL)

    Purification yield of the active peptide was estimated to be 0.2 ± 0.1%, starting from the lyophilized jellyfish. The infrared (IR), proton nuclear magnetic resonance spectroscopy (1H NMR), carbon nuclear magnetic resonance (13C NMR) and mass spectrometry (MS) spectrometer analyses elucidated that the structure of the ...

  15. Purification of kappa (k)-carrageenase from locally isolated ...

    African Journals Online (AJOL)

    Partial purification of the crude kappa (k)-carrageenase present in the culture filtrates of Cellulosimicrobium cellulans was carried out by fractional precipitation, using ammonium sulphate, acetone and ethanol individually. The highest recovered protein (37.08%) combined with enzyme activity was obtained with ammonium ...

  16. Purification of bacteriocins using size-exclusion chromatography

    Directory of Open Access Journals (Sweden)

    Vivek K. Bajpai

    2016-06-01

    Full Text Available The bacteriocin purification involves following main steps. a. Extraction of cell-free-supernatant of bacteria. b. Ammonium sulfate precipitation. c. Dialysis. d. Diafiltration using PVP and e. Size-exclusion chromatography. However, depending on the nature of work, the compound could be further analyzed by reverse-phase HPLC, NMR, mass spectrometry and sequencing.

  17. Solubilization and purification of Escherichia coli expressed GST ...

    African Journals Online (AJOL)

    STORAGESEVER

    2009-05-18

    May 18, 2009 ... prognosis, monitoring of therapy and diagnosis. VEGF has been identified as the target for the treatment of cancer. Though prokaryotic expression of VEGF has been done, the solubilization and purification is time consuming and empirical. In this study, VEGF165 and VEGF121 were cloned into. pGEX-4T-1 ...

  18. Optimization of membrane protein overexpression and purification using GFP fusions

    NARCIS (Netherlands)

    Drew, David; Lerch, Mirjam; Kunji, Edmund; Slotboom, Dirk-Jan; de Gier, Jan-Willem

    Optimizing conditions for the overexpression and purification of membrane proteins for functional and structural studies is usually a Laborious and time-consuming process. This process can be accelerated using membrane protein-GFP fusions(1-3), which allows direct monitoring and visualization of

  19. Purification and biochemical characterization of a novel glutathione ...

    African Journals Online (AJOL)

    SERVER

    2008-02-05

    Feb 5, 2008 ... Purification and biochemical characterization of a novel glutathione S-transferase of the silkworm, ... silkworm as a model for lepidopteran insects is also useful to obtain data on its detoxification .... The effect of temperature on CDNB conjugation reaction catalyzed by BmGST was examined under the ...

  20. Purification the surface of detail from biological contaminations

    Science.gov (United States)

    Gabdrakhmanov, Az T.; Israphilov, I. H.; Galiakbarov, A. T.; Gabdrakhmanov, Al T.

    2017-01-01

    More than 70% of biodegradation occur due to the corrosion processes. A biological corrosion causes the greatest damage to the oil and gas-production industry, the Navy and pipelines, constructions of water supply, means of communication. This paper proposes an effective method of purification various surfaces from biological contaminations by using of cold plasma.

  1. Rapid purification of high activity Taq DNA polymerase expressed in ...

    African Journals Online (AJOL)

    A simplified method is described here for the preparation of a thermostable Taq DNA polymerase enzyme from Escherichia coli (E. coli) strain DH5a carrying the pTTQ18 expression vector transformed with the Taq polymerase gene. Standard purifications were done with 1 litre batch cultures of E. coli cells and produced ...

  2. Purification and characterization of laccase from Trametes hirsuta ...

    African Journals Online (AJOL)

    Purification and characterization of laccase from Trametes hirsuta Bm-2 and its contribution to dye and effluent decolorization. P Zapata-Castillo, MdeL Villalonga-Santana, J Tamayo-Cortés, G Rivera-Muñoz, S Solís-Pereira ...

  3. Optimized conditions for high-level solubilization and purification of ...

    African Journals Online (AJOL)

    In this report, we describe the cloning, over-expression, efficient solubilization, purification and evaluation of bioactivity of camel growth hormone (cGH). The total cellular RNA was extracted from pituitary glands of freshly slaughtered animals and cDNA of cGH was synthesized by a pair of sequence specific primers with a ...

  4. Isolation and purification of psoralen and isopsoralen and their ...

    African Journals Online (AJOL)

    Isolation and purification of psoralen and isopsoralen and their efficacy and safety in the treatment of osteosarcoma in nude rats. Honghui Lu¥ , Lihai Zhang¥, Daohong Liu, Peifu Tang2, Feixiang Song. Orthopaedic Department , People's Liberation Army General Hospital, Beijing 100000, China. ¥Authors Contributed ...

  5. Purification and characterization of the chitosanase from Aeromonas ...

    African Journals Online (AJOL)

    Purification and characterization of the chitosanase from Aeromonas sp. HG08. Y Sun, J Zhang, S Wang. Abstract. In this work, the characterization of a chitosanase-producing bacterium isolated from soil was reported and this strain was grouped under the genus Aeromonas by virtue of its morphological, physiological ...

  6. Kinetic modelling and thermodynamic studies on purification of ...

    African Journals Online (AJOL)

    ... kinetic model suggesting that the adsorption mechanism might be a chemisorption process; thermodynamic studies showed that this process is spontaneous and endothermic. These results are very important in optimization of this purification process. Keywords: Polyvinylpyrrolidone (PVP), adsorption, activated carbon, ...

  7. Purification and properties of Rhizobial DehL expressed in ...

    African Journals Online (AJOL)

    STORAGESEVER

    2008-06-17

    Jun 17, 2008 ... Full Length Research Paper. Purification and properties of Rhizobial DehL expressed in Escherichia coli. Fahrul Huyop1*, Nooraini Abdul Rashid1, Roswanira A. B. Wahab2 and Ronald A. Cooper3. 1Industrial Biotechnology Department, University Technology Malaysia, 81310 Skudai, Johor, Malaysia.

  8. Purification and Characterization of Lipase from Aspergillus flavus ...

    African Journals Online (AJOL)

    USER

    application. Key words: Lipase, purification, Aspergillus flavus PW2961, magnetic nanoparticles.______. Correspondence: sharafkareem@yahoo.co.uk ..... in solid state fermentation. Process Biochem. 38(5): 715-721. Okoli, C., Boutonnet, M., Mariey, L., Jaras, S. And Rajarao, G. (2011). Application of magnetic iron oxide ...

  9. Purification performances of common reed beds based on the ...

    African Journals Online (AJOL)

    SARAH

    2013-11-30

    Nov 30, 2013 ... technology. Organic treatments are cheap, efficient, and require little electric energy. Objective: This study aims to determine the purification performances of ... water crops. According to the INSAE (2002), Benin had a population of 6 million people in 2000; in 2012, this population would reach 9 million.

  10. Effective Purification of Biogas by Condensing-Liquid Membrane

    Czech Academy of Sciences Publication Activity Database

    Poloncarzová, Magda; Vejražka, Jiří; Veselý, Václav; Izák, Pavel

    2010-01-01

    Roč. 50, č. 3 (2010), s. 669-671 ISSN 1433-7851 R&D Projects: GA MPO FR-TI1/245 Institutional research plan: CEZ:AV0Z40720504 Keywords : biogas purification * condensing liquid * gas permeation Subject RIV: CI - Industrial Chemistry, Chemical Engineering Impact factor: 12.730, year: 2010

  11. Comparative study of methods for extraction and purification of ...

    African Journals Online (AJOL)

    USER

    2010-08-02

    Aug 2, 2010 ... et al., 1999; Yu and Mohn, 1999) and water samples. (England et al., 2001; Rivera et al., 2003). These methods are generally based on three steps: physical, chemical and or enzymatic lysis for direct or indirect extraction of. DNA followed by purification. Unlike pure culture samples, factors such as the ...

  12. Cloning, high-level expression, purification and characterization of a ...

    African Journals Online (AJOL)

    The staphylokinase (Sak) is emerging as an important thrombolytic agent for the treatment of patients suffering from cardiovascular disease. Hence in this study, we reported the cloning, high-level expression, purification and characterization of the Sak variant SakøC from Staphylococcus aureus QT08 in Escherichia coli ...

  13. Extraction and purification of formonometin from Trifolium pratense L ...

    African Journals Online (AJOL)

    Extraction and purification of formonometin from Trifolium pratense L: Physicochemical properties of its complex with ... been limited by the poor solubility in water. Solubility is essential in the oral bio-availability of functional food ... A copper testing stub was used as the carrier of the samples. A layer of gold was sputtered on.

  14. Purification and Some Properties of a Thermostable α-Amylase ...

    African Journals Online (AJOL)

    This work reports the isolation, purification and some properties of a thermostable α-amylase producing Bacillus subtilis isolated from the soil. Soil samples were collected and screened for thermophilic bacterial strains with amylase activity and to examine the amylase heat tolerance potentiality. The isolate was Gram ...

  15. Purification and characterization of a protease from Thermophilic ...

    African Journals Online (AJOL)

    AJB SERVER

    2006-10-19

    Oct 19, 2006 ... presented in this paper. The purification steps included ammonium sulfate precipitation, with columns of DEAE-Sepharose anion exchange chromatography and Sephacryl S-100HR on AKTA purifier 100 protein liquid chromatography. The method gave a 4.25 fold increase of the specific activity and had a.

  16. Purification of recombinant C-terminus polyhistidine tagged human ...

    African Journals Online (AJOL)

    Dell

    2012-05-03

    May 3, 2012 ... Bringing the excitement and motivation of research to students; Using inquiry and research-based learning in a year-long biochemistry laboratory: Part. I-guided inquiry-purification and characterization of a fusion protein: Histidine tag, malate dehydrogenase, and green fluorescent protein. Biochem. Mol.

  17. Synthesis Gas Purification Purification des gaz de synthèse

    Directory of Open Access Journals (Sweden)

    Chiche D.

    2013-10-01

    Full Text Available Fischer-Tropsch (FT based B-XTL processes are attractive alternatives for future energy production. These processes aim at converting lignocellulosic biomass possibly in co-processing with petcoke, coal, or vacuum residues into synthetic biofuels. A gasification step converts the feed into a synthesis gas (CO and H2 mixture , which undergoes the Fischer-Tropsch reaction after H2/CO ratio adjustment and CO2 removal. However synthesis gas also contains various impurities that must be removed in order to prevent Fischer-Tropsch catalyst poisoning. Due to the large feedstocks variety that can be processed, significant variations of the composition of the synthesis gas are expected. Especially, this affects the nature of the impurities that are present (element, speciation, as well as their relative contents. Moreover, due to high FT catalyst sensitivity, severe syngas specifications regarding its purity are required. For these reasons, synthesis gas purification constitutes a major challenge for the development of B-XTL processes. In this article, we focus on these major hurdles that have to be overcome. The different kinds of syngas impurities are presented. The influence of the nature of feedstocks, gasification technology and operating conditions on the type and content of impurities is discussed. Highlight is given on the fate of sulfur compounds, nitrogen compounds, halides, transition and heavy metals. Main synthesis gas purification technologies (based on adsorption, absorption, catalytic reactions, etc. are finally described, as well as the related challenges. Les procédés de synthèse de biocarburants par voie Fischer-Tropsch (FT, voies B-XTL, représentent des alternatives prometteuses pour la production d’énergie. Ces procédés permettent la conversion en carburants de synthèse de biomasse lignocellulosique, éventuellement mise en oeuvre en mélange avec des charges fossiles telles que petcoke, charbons ou résidus sous vide. Pour

  18. INVESTIGATIONS ON BIOCHEMICAL PURIFICATION OF GROUND WATER FROM HYDROGEN SULFIDE

    Directory of Open Access Journals (Sweden)

    Yu. P. Sedlukho

    2015-01-01

    Full Text Available The paper considers problems and features of biochemical removal of hydrogen sulfide from ground water. The analysis of existing methods for purification of ground water from hydrogen sulfide has been given in the paper. The paper has established shortcomings of physical and chemical purification of ground water. While using aeration methods for removal of hydrogen sulfide formation of colloidal sulfur that gives muddiness and opalescence to water occurs due to partial chemical air oxidation. In addition to this violation of sulfide-carbonate equilibrium taking place in the process of aeration due to desorption of H2S and CO2, often leads to clogging of degasifier nozzles with formed CaCO3 that causes serious operational problems. Chemical methods require relatively large flow of complex reagent facilities, storage facilities and transportation costs.In terms of hydrogen sulfide ground water purification the greatest interest is given to the biochemical method. Factors deterring widespread application of the biochemical method is its insufficient previous investigation and necessity to execute special research in order to determine optimal process parameters while purifying groundwater of a particular water supply source. Biochemical methods for oxidation of sulfur compounds are based on natural biological processes that ensure natural sulfur cycle. S. Vinogradsky has established a two-stage mechanism for oxidation of hydrogen sulfide with sulfur bacteria (Beggiatoa. The first stage presupposes oxidation of hydrogen sulphide to elemental sulfur which is accumulating in the cytoplasm in the form of globules. During the second stage sulfur bacteria begin to oxidize intracellular sulfur to sulfuric acid due to shortage of hydrogen sulfide.The paper provides the results of technological tests of large-scale pilot plants for biochemical purification of groundwater from hydrogen sulfide in semi-industrial conditions. Dependences of water quality

  19. Purification and characterization of osteopontin from human milk.

    Science.gov (United States)

    Sørensen, Steen; Justesen, Steen Just; Johnsen, Anders H

    2003-08-01

    Osteopontin (OPN) is expressed in many organs and tissues and has different biological properties related to different molecular forms in respect to size and posttranslational modifications. However, a purification procedure for authentic intact OPN as well as fragments of OPN from an accessible biological source is missing. A four-step procedure was used to purify OPN from human milk, based on its crystal growth inhibitory activity, including anion exchange chromatography, the elimination of casein, hydroxyapatite chromatography, and negative affinity chromatography. Purified OPN was further separated into its different molecular forms by means of a two-step procedure, involving size exclusion chromatography and reverse phase chromatography. A rabbit polyclonal antibody was raised to purified intact OPN and high M(r) OPN components; the immunoreactivity of both forms was almost equal when investigated by enzyme immunoassay (EIA). The procedures facilitate the purification of intact OPN and OPN fragments for purposes of standardization, preparation of monospecific antibodies, and functional studies.

  20. Uranium hexafluoride purification; Purificacao de hexafluoreto de uranio

    Energy Technology Data Exchange (ETDEWEB)

    Araujo, Eneas F. de

    1986-07-01

    Uranium hexafluoride might contain a large amount of impurities after manufacturing or handling. Three usual methods of purification of uranium hexafluoride were presented: selective sorption, sublimation, and distillation. Since uranium hexafluoride usually is contaminated with hydrogen fluoride, a theoretical study of the phase equilibrium properties was performed for the binary system UF{sub 6}-HF. A large deviation from the ideal solution behaviour was observed. A purification unity based on a constant reflux batch distillation process was developed. A procedure was established in order to design the re boiler, condenser and packed columns for the UF{sub 6}-HF mixture separation. A bench scale facility for fractional distillation of uranium hexafluoride was described. Basic operations for that facility and results extracted from several batches were discussed. (author)

  1. Chemical purification of lanthanides for low-background experiments

    Science.gov (United States)

    Boiko, R. S.

    2017-10-01

    There are many potentially active isotopes among the lanthanide elements which are possible to use for low-background experiments to search for double β decay, dark matter, to investigate rare α and β decays. These kind of experiments require very low level of radioactive contamination, but commercially available compounds of lanthanides are always contamined by uranium, thorium, radium, potassium, etc. A simple chemical method based on liquid-liquid extraction has been applied for the purification of CeO2, Nd2O3 and Gd˙2O˙3 from radioactive traces. Detailed schemes of purification procedure are described. Measurements by using HPGe spectrometry demonstrate high efficiency in K, Ra, Th, U contaminations reduction on at least one order of magnitude.

  2. Preparative Purification of Recombinant Proteins: Current Status and Future Trends

    Directory of Open Access Journals (Sweden)

    Mayank Saraswat

    2013-01-01

    Full Text Available Advances in fermentation technologies have resulted in the production of increased yields of proteins of economic, biopharmaceutical, and medicinal importance. Consequently, there is an absolute requirement for the development of rapid, cost-effective methodologies which facilitate the purification of such products in the absence of contaminants, such as superfluous proteins and endotoxins. Here, we provide a comprehensive overview of a selection of key purification methodologies currently being applied in both academic and industrial settings and discuss how innovative and effective protocols such as aqueous two-phase partitioning, membrane chromatography, and high-performance tangential flow filtration may be applied independently of or in conjunction with more traditional protocols for downstream processing applications.

  3. A scintillator purification plant and fluid handling system for SNO+

    Science.gov (United States)

    Ford, Richard J.

    2015-08-01

    A large capacity purification plant and fluid handling system has been constructed for the SNO+ neutrino and double-beta decay experiment, located 6800 feet underground at SNOLAB, Canada. SNO+ is a refurbishment of the SNO detector to fill the acrylic vessel with liquid scintillator based on Linear Alkylbenzene (LAB) and 2 g/L PPO, and also has a phase to load natural tellurium into the scintillator for a double-beta decay experiment with 130Te. The plant includes processes multi-stage dual-stream distillation, column water extraction, steam stripping, and functionalized silica gel adsorption columns. The plant also includes systems for preparing the scintillator with PPO and metal-loading the scintillator for double-beta decay exposure. We review the basis of design, the purification principles, specifications for the plant, and the construction and installations. The construction and commissioning status is updated.

  4. Experimental nested purification for a linear optical quantum repeater

    Science.gov (United States)

    Chen, Luo-Kan; Yong, Hai-Lin; Xu, Ping; Yao, Xing-Can; Xiang, Tong; Li, Zheng-Da; Liu, Chang; Lu, He; Liu, Nai-Le; Li, Li; Yang, Tao; Peng, Cheng-Zhi; Zhao, Bo; Chen, Yu-Ao; Pan, Jian-Wei

    2017-11-01

    Quantum repeaters1-4 are essential elements for demonstrating global-scale quantum communication. Over the past few decades, tremendous efforts have been dedicated to implementing a practical quantum repeater5-10. However, nested purification1, the backbone of a quantum repeater, remains a challenge because the capacity for successive entanglement manipulation is still absent. Here, we propose and demonstrate an architecture of nested purification using spontaneous parametric downconversion sources11. A heralded entangled photon pair with higher fidelity is successfully purified from two copies of low-fidelity pairs that experience entanglement swapping and noisy channels. By delicately designing the optical circuits, double-pair emission noise is eliminated automatically and the purified state can be used for scalable entanglement connections to extend the communication distance. Combined with a quantum memory, our approach can be applied immediately in the implemention of a practical quantum repeater.

  5. Ion-exchange chromatography purification of extracellular vesicles.

    Science.gov (United States)

    Kosanović, Maja; Milutinović, Bojana; Goč, Sanja; Mitić, Ninoslav; Janković, Miroslava

    2017-08-01

    Despite numerous studies, isolating pure preparations of extracellular vesicles (EVs) has proven challenging. Here, we compared ion-exchange chromatography (IEC) to the widely used sucrose density gradient (SDG) centrifugation method for the purification of EVs. EVs in bulk were isolated from pooled normal human amniotic fluid (AF) by differential centrifugation followed by IEC or sucrose density gradient separation. The purity of the isolated EVs was evaluated by electrophoresis and lectin blotting/immuno blotting to monitor the distribution of total proteins, different EVs markers, and selected N-glycans. Our data showed efficient separation of negatively charged EVs from other differently charged molecules, while comparative profiling of EVs using SDG centrifugation confirmed anion-exchange chromatography is advantageous for EV purification. Finally, although this IEC-based method was validated using AF, the approach should be readily applicable to isolation of EVs from other sources as well.

  6. [Purification and breaking techniques for cysts of Giardia spp].

    Science.gov (United States)

    Polverino, D; Molina, N B; Minvielle, M C; Lozano, M E; Basualdo, J A

    2004-01-01

    The purpose of this study was to optimize and evaluate the purification techniques, isolation and breaking of cysts of Giardia spp from fecal samples to isolate DNA. Filtrated fecal samples were tested in 3 purification techniques: Telleman solution, sucrose and Telleman plus sucrose. The sucrose solution let us to isolate the cysts with less detritus. The cleaned cysts were splited in 3 techniques to test the breaking: osmotic shock and heat, chemistry degradation and thermic shock, enzymatic action and mechanic effect. Only the last method was successful and showed bands in agarose gel. The result of this study shows a routine and common method which could be used in the previous steps to the PCR technique for the genotypification of these parasites.

  7. Technical project for a new water purification solution

    Directory of Open Access Journals (Sweden)

    Toma Adina

    2018-01-01

    Full Text Available This research is part of the RO-BG Cross-Border Cooperation Program, project “CLEANDANUBE”, MIS-ETC 653, which has finalised by providing a common strategy to prevent the Danube’s pollution technological risks with oil and oil products. This paper presents a new sustainable water purification solution. A short introduction will be offered and an overview regarding the research and new methods to greening the waste is provided. The theoretical aspects of the centrifugal separation phenomenon are studied and the preliminary project bases were established. The paper conveys the possible constructive variations and the technological implications of those. Ultimately, the technical project for a new water purification solution and conclusions with critical points encountered during the designing phase are presented.

  8. A scintillator purification plant and fluid handling system for SNO+

    Energy Technology Data Exchange (ETDEWEB)

    Ford, Richard J., E-mail: ford@snolab.ca [SNOLAB, Creighton Mine #9, 1039 R.R.24, Lively, Ontario, Canada. (Canada)

    2015-08-17

    A large capacity purification plant and fluid handling system has been constructed for the SNO+ neutrino and double-beta decay experiment, located 6800 feet underground at SNOLAB, Canada. SNO+ is a refurbishment of the SNO detector to fill the acrylic vessel with liquid scintillator based on Linear Alkylbenzene (LAB) and 2 g/L PPO, and also has a phase to load natural tellurium into the scintillator for a double-beta decay experiment with {sup 130}Te. The plant includes processes multi-stage dual-stream distillation, column water extraction, steam stripping, and functionalized silica gel adsorption columns. The plant also includes systems for preparing the scintillator with PPO and metal-loading the scintillator for double-beta decay exposure. We review the basis of design, the purification principles, specifications for the plant, and the construction and installations. The construction and commissioning status is updated.

  9. Purification process of recombinant monoclonal antibodies with mixed mode chromatography.

    Science.gov (United States)

    Maria, Sophie; Joucla, Gilles; Garbay, Bertrand; Dieryck, Wilfrid; Lomenech, Anne-Marie; Santarelli, Xavier; Cabanne, Charlotte

    2015-05-08

    An innovative process to purify mAb from CHO cell culture supernatant was developed. This three-step process involved two mixed mode resins and an anion exchange membrane. We used a human IgG mixture to determine the optimal conditions for each purification step. Thereafter, the whole process was evaluated and improved for the purification of a recombinant mAb produced in the supernatant of CHO cells. Once optimized, yield and purity of 88% and 99.9%, respectively were comparable to those obtained in a conventional process based on a capture step using protein A. In addition, aggregates, HCPs and DNA levels in the purified fraction were below regulatory specifications. Then we used mass spectrometry to identify contaminating proteins in the antibody fraction in order to highlight the behavior of HCPs. Copyright © 2015 Elsevier B.V. All rights reserved.

  10. Antibody Fragments and Their Purification by Protein L Affinity Chromatography

    Directory of Open Access Journals (Sweden)

    Gustav Rodrigo

    2015-09-01

    Full Text Available Antibodies and related proteins comprise one of the largest and fastest-growing classes of protein pharmaceuticals. A majority of such molecules are monoclonal antibodies; however, many new entities are antibody fragments. Due to their structural, physiological, and pharmacological properties, antibody fragments offer new biopharmaceutical opportunities. In the case of recombinant full-length antibodies with suitable Fc regions, two or three column purification processes centered around Protein A affinity chromatography have proven to be fast, efficient, robust, cost-effective, and scalable. Most antibody fragments lack Fc and suitable affinity for Protein A. Adapting proven antibody purification processes to antibody fragments demands different affinity chromatography. Such technology must offer the unit operation advantages noted above, and be suitable for most of the many different types of antibody fragments. Protein L affinity chromatography appears to fulfill these criteria—suggesting its consideration as a key unit operation in antibody fragment processing.

  11. Superhydrophobic coated apparatus for liquid purification by evaporative condensation

    Science.gov (United States)

    Simpson, John T; McNeany, Steve R; Dinsmore, Thomas V; Hunter, Scott R; Ivanov, Ilia N

    2014-03-11

    Disclosed are examples of apparatuses for evaporative purification of a contaminated liquid. In each example, there is a first vessel for storing the contaminated fluid. The first vessel includes a surface coated with a layer of superhydrophobic material and the surface is at least partially in contact with the contaminated liquid. The contaminants do not adhere to the surface as the purified liquid evaporates, thus simplifying maintenance of the apparatus.

  12. Production and Purification of Peroxidase from Aspergillus niger.

    OpenAIRE

    Mohammed A. Jebor

    2017-01-01

    This study was conducted in the laboratories of Biology Department, College of Science, which deals with isolation and purification of peroxidase and optimization of process parameters to achieve maximum yield of peroxidase by Aspergillus niger. Solid-state fermentation of Aspergillus niger was carried out for enhanced production of peroxidase using hydrogen peroxide as the substrate of enzyme maximum activity of the enzyme was achieved under optimum growth conditions. The optimum conditi...

  13. Modular flue gas purification systems; Rauchgasreinigung in modularer Form

    Energy Technology Data Exchange (ETDEWEB)

    Mergler, R. [Lurgi Umwelt GmbH, Frankfurt am Main (Germany). Geschaeftsbereich Vertrieb

    1997-02-01

    Using the example of a flue gas purification system for a waste incinerator, the author shows how modularisation and standardisation of processes will optimize the cost and make efficient use of the available know-how on environmental engineering. (orig.) [Deutsch] Am Beispiel der Rauchgasreinigung fuer eine thermische Restabfallbehandlung wird dargestellt, wie die Modularisierung/Standardisierung von Verfahren zur Kostenoptimierung und zum effizienten Einsatz von Verfahrens-know-how in der Umwelttechnik genutzt werden kann. (orig.)

  14. Expression and purification of recombinant Shiga toxin 2B from ...

    African Journals Online (AJOL)

    sunny t

    expression of Stx2B economically was obtained using 1 mM IPTG for 4 h at 37°C. Protein identity was confirmed by a band at ~11.4 kDa using ... of recombinant Shiga toxin B subunit (rStxB) protein in. BALB/c mice was evaluated. Animal ..... tag-based purification of PKCƔ Kinase free tag protein. (Ohana et al., 2011) which ...

  15. Purification and characterization of a milk-clotting protease from ...

    African Journals Online (AJOL)

    Crude enzymatic extract obtained from five fermentations (300 g of wheat bran) was characterized by a clotting activity of 0.34 ± 0.08 UP/ml with a strength ratio of 1/1: 200. The comparative study of the summaries from 2 purification protocols showed that it is possible to recover 6% of the initial proteins with a 44.54% activity ...

  16. Aqueous Chloride Operations Overview: Plutonium and Americium Purification/Recovery

    Energy Technology Data Exchange (ETDEWEB)

    Gardner, Kyle Shelton [Los Alamos National Lab. (LANL), Los Alamos, NM (United States); Kimball, David Bryan [Los Alamos National Lab. (LANL), Los Alamos, NM (United States); Skidmore, Bradley Evan [Los Alamos National Lab. (LANL), Los Alamos, NM (United States)

    2016-09-28

    These are a set of slides intended for an information session as part of recruiting activities at Brigham Young University. It gives an overview of aqueous chloride operations, specifically on plutonium and americium purification/recovery. This presentation details the steps taken perform these processes, from plutonium size reduction, dissolution, solvent extraction, oxalate precipitation, to calcination. For americium recovery, it details the CLEAR (chloride extraction and actinide recovery) Line, oxalate precipitation and calcination.

  17. Peptide affinity reagents for AAV capsid recognition and purification.

    Science.gov (United States)

    Pulicherla, N; Asokan, A

    2011-10-01

    We report the discovery of AAV capsid-binding peptides identified through phage panning. The heptapeptide motif GYVSRHP selectively recognized AAV serotype 8 capsids and blocked transduction in vitro. Recombinant AAV8 vectors were purified directly from crude cell lysate and supernatant through sequential application of peptide affinity and anion exchange chromatography. Peptide affinity reagents may serve as useful alternatives to monoclonal antibodies in AAV capsid recognition, and offer readily scalable solutions for purification of clinical grade AAV vectors.

  18. Purification of Dorsal Root Ganglion Neurons from Rat by Immunopanning

    OpenAIRE

    Zuchero, J. Bradley

    2014-01-01

    Dorsal root ganglion neurons (DRGs) are sensory neurons that facilitate somatosensation and have been used to study neurite outgrowth, regeneration, and degeneration and PNS and CNS myelination. Studies of DRGs have relied on cell isolation strategies that generally involve extended culture in the presence of antimitotic agents or other cytotoxic treatments that target dividing cells. The surviving cells typically are dependent on serum for growth. Other methods, involving purification of DRG...

  19. Two-Step Vapor/Liquid/Solid Purification

    Science.gov (United States)

    Holland, L. R.

    1986-01-01

    Vertical distillation system combines in single operation advantages of multiple zone refining with those of distillation. Developed specifically to load Bridgman-Stockbarger (vertical-solidification) growth ampoules with ultrapure tellurium and cadmium, system, with suitable modifications, serves as material refiner. In first phase of purification process, ampoule heated to drive off absorbed volatiles. Second phase, evaporator heated to drive off volatiles in charge. Third phase, slowly descending heater causes distillation from evaporator to growing crystal in ampoule.

  20. Secretory immunoglobulin purification from whey by chromatographic techniques.

    Science.gov (United States)

    Matlschweiger, Alexander; Engelmaier, Hannah; Himmler, Gottfried; Hahn, Rainer

    2017-08-15

    Secretory immunoglobulins (SIg) are a major fraction of the mucosal immune system and represent potential drug candidates. So far, platform technologies for their purification do not exist. SIg from animal whey was used as a model to develop a simple, efficient and potentially generic chromatographic purification process. Several chromatographic stationary phases were tested. A combination of two anion-exchange steps resulted in the highest purity. The key step was the use of a small-porous anion exchanger operated in flow-through mode. Diffusion of SIg into the resin particles was significantly hindered, while the main impurities, IgG and serum albumin, were bound. In this step, initial purity was increased from 66% to 89% with a step yield of 88%. In a second anion-exchange step using giga-porous material, SIg was captured and purified by step or linear gradient elution to obtain fractions with purities >95%. For the step gradient elution step yield of highly pure SIg was 54%. Elution of SIgA and SIgM with a linear gradient resulted in a step yield of 56% and 35%, respectively. Overall yields for both anion exchange steps were 43% for the combination of flow-through and step elution mode. Combination of flow-through and linear gradient elution mode resulted in a yield of 44% for SIgA and 39% for SIgM. The proposed process allows the purification of biologically active SIg from animal whey in preparative scale. For future applications, the process can easily be adopted for purification of recombinant secretory immunoglobulin species. Copyright © 2017 Elsevier B.V. All rights reserved.

  1. Reverse osmosis membrane of high urea rejection properties. [water purification

    Science.gov (United States)

    Johnson, C. C.; Wydeven, T. J. (Inventor)

    1980-01-01

    Polymeric membranes suitable for use in reverse osmosis water purification because of their high urea and salt rejection properties are prepared by generating a plasma of an unsaturated hydrocarbon monomer and nitrogen gas from an electrical source. A polymeric membrane is formed by depositing a polymer of the unsaturated monomer from the plasma onto a substrate, so that nitrogen from the nitrogen gas is incorporated within the polymer in a chemically combined form.

  2. Purification and characterization of the chitosanase from Aeromonas ...

    African Journals Online (AJOL)

    STORAGESEVER

    2009-06-17

    Jun 17, 2009 ... Q-Sepharose Fast Flow. 10. 2.0. 5. 55.6. 12.5. Sephadex G-75. 8.5. 1.0. 8.5. 47.2. 21.25. aThe chitosanase was purified by the following chromatographic procedure. All purification steps were performed at 4°C. Step. 1: The bacterial cells were pelleted out from the culture medium by centrifugation (10 ...

  3. The Blood Compatibilities of Blood Purification Membranes and Other Materials Developed in Japan

    Directory of Open Access Journals (Sweden)

    Takaya Abe

    2011-01-01

    Full Text Available The biocompatibilities in blood purification therapy are defined as “a concept to stipulate safety of blood purification therapy by an index based on interaction in the body arising from blood purification therapy itself.” The biocompatibilities are associated with not only materials to be used but also many factors such as sterilization method and eluted substance. It is often evaluated based on impacts on cellular pathways and on humoral pathways. Since the biocompatibilities of blood purification therapy in particular hemodialysis are not just a prognostic factor for dialysis patients but a contributory factor for long-term complications, it should be considered with adequate attention. It is important that blood purification therapy should be performed by consistently evaluating not only risks associated with these biocompatibilities but also the other advantages obtained from treatments. In this paper, the biocompatibilities of membrane and adsorption material based on Japanese original which are used for blood purification therapy are described.

  4. Identification and Purification of Montmorillonite Mineral of Ghaen Mine Bentonite

    Directory of Open Access Journals (Sweden)

    P. Mirhoseini Moosavi

    2015-06-01

    Full Text Available Montmorillonite is the major mineral of Bentonite with many applications in industrial fields but some impurities decreases the quality of the bentonite. The main objective of this study was to investigate the suitable method for purification of Ghaen mine bentonite. A combination of methods was considered including wet sieving and sedimentation, centrifuge and ultrasound. The efficiency of purification methods was determined based on X-ray, particle size, cation exchange capacity (CEC and ratio peak of the Quartz/Montmorillonite analysis before and after experiments. The results showed that such methods were efficient for preparing of the materials having high quantity of montmorillonite with less than 2 microns particle sizes. Cristobalite was the only mineral remained in samples, however many of particles were exempted from the samples. Cristobalite was the main impurity remained with montmorillonite. Chemical treatment is the only way for its complete removal. The results of this study revealed that by using easy, cheap and fast methods, it is possible for acceptable purification of bentonite.

  5. Countercurrent tangential chromatography for large-scale protein purification.

    Science.gov (United States)

    Shinkazh, Oleg; Kanani, Dharmesh; Barth, Morgan; Long, Matthew; Hussain, Daniar; Zydney, Andrew L

    2011-03-01

    Recent advances in cell culture technology have created significant pressure on the downstream purification process, leading to a "downstream bottleneck" in the production of recombinant therapeutic proteins for the treatment of cancer, genetic disorders, and cardiovascular disease. Countercurrent tangential chromatography overcomes many of the limitations of conventional column chromatography by having the resin (in the form of a slurry) flow through a series of static mixers and hollow fiber membrane modules. The buffers used in the binding, washing, and elution steps flow countercurrent to the resin, enabling high-resolution separations while reducing the amount of buffer needed for protein purification. The results obtained in this study provide the first experimental demonstration of the feasibility of using countercurrent tangential chromatography for the separation of a model protein mixture containing bovine serum albumin and myoglobin using a commercially available anion exchange resin. Batch uptake/desorption experiments were used in combination with critical flux data for the hollow fiber filters to design the countercurrent tangential chromatography system. A two-stage batch separation yielded the purified target protein at >99% purity with 94% recovery. The results clearly demonstrate the potential of using countercurrent tangential chromatography for the large-scale purification of therapeutic proteins. Copyright © 2010 Wiley Periodicals, Inc.

  6. Different purification methods and quality of sunflower biodiesel

    Energy Technology Data Exchange (ETDEWEB)

    Pighinelli, A.L.M.T.; Park, K.J. [Campinas State Univ., Sao Paulo (Brazil). School of Agricultural Engineering; Ferrari, R.A.; Miguel, A.M.R.O. [Food Technology Inst., Sao Paulo (Brazil)

    2010-07-01

    Biodiesel is derived from triacylglycerides and is produced primarily through transesterification, a chemical reaction of vegetable oils with alcohol, methanol or ethanol. The cost of raw material should be considered since 85 per cent of production cost is related to vegetable oil. The purpose of this study was to evaluate oil expression of sunflower seed. It also examined the sunflower crude oil as a raw material for biodiesel by transesterification in both laboratory and pilot scale studies. Three different biodiesel purification methods were examined. The best result for oil expelling (68.4 per cent) at the experimental stage was obtained for seeds with a moisture content of 6.9 per cent at 25 degrees C and at a screw speed of 114 rpm. For biodiesel production at the laboratory scale, the best result for oil expelling was 87.5 per cent. It was obtained with an ethanol:oil molar ratio of 4.7:1 and with a 4.42 per cent catalyst concentration related to the quantity of oil that had to be transesterified. The experimental condition was applied at a bigger scale with a batch stirred tank reactor. For purification with washing, the biodiesel yield was 84.2 per cent. Purification with silica resulted in a yield of 84.6 per cent. A better quality biofuel was obtained through distillation of biodiesel.

  7. A comprehensive review on biodiesel purification and upgrading

    Directory of Open Access Journals (Sweden)

    Hamed Bateni

    2017-09-01

    Full Text Available Serious environmental concerns regarding the use of fossil-based fuels have raised awareness regarding the necessity of alternative clean fuels and energy carriers. Biodiesel is considered a clean, biodegradable, and non-toxic diesel substitute produced via the transesterification of triglycerides with an alcohol in the presence of a proper catalyst. After initial separation of the by-product (glycerol, the crude biodiesel needs to be purified to meet the standard specifications prior to marketing. The presence of impurities in the biodiesel not only significantly affects its engine performance but also complicates its handling and storage. Therefore, biodiesel purification is an essential step prior to marketing. Biodiesel purification methods can be classified based on the nature of the process into equilibrium-based, affinity-based, membrane-based, reaction-based, and solid-liquid separation processes. The main adverse properties of biodiesel – namely moisture absorption, corrosiveness, and high viscosity – primarily arise from the presence of oxygen. To address these issues, several upgrading techniques have been proposed, among which catalytic (hydrodeoxygenation using conventional hydrotreating catalysts, supported metallic materials, and most recently transition metals in various forms appear promising. Nevertheless, catalyst deactivation (via coking and/or inadequacy of product yields necessitate further research. This paper provides a comprehensive overview on the techniques and methods used for biodiesel purification and upgrading.

  8. The modified swirl sedimentation tanks for water purification.

    Science.gov (United States)

    Ochowiak, Marek; Matuszak, Magdalena; Włodarczak, Sylwia; Ancukiewicz, Małgorzata; Krupińska, Andżelika

    2017-03-15

    This paper discusses design, evaluation, and application for the use of swirl/vortex technologies as liquid purification system. A study was performed using modified swirl sedimentation tanks. The vortex separators (OW, OWK, OWR and OWKR) have been studied under laboratory conditions at liquid flow rate from 2.8⋅10(-5) to 5.1⋅10(-4) [m(3)/s]. The pressure drop and the efficiency of purification of liquid stream were analyzed. The suspended particles of different diameters were successfully removed from liquid with the application of swirl chambers of proposed constructions. It was found that damming of liquid in the tank increases alongside liquid stream at the inlet and depends on the tank construction. The efficiency of the sedimentation tanks increases alongside the diameters of solid particles and decrease in the liquid flow rate. The best construction proved to be the OWR sedimentation tank due to smallest liquid damming, even at high flow rates, and the highest efficiency of the purification liquid stream for solid particles of the smallest diameter. The proposed solution is an alternative to the classical constructions of sedimentation tanks. Copyright © 2016 Elsevier Ltd. All rights reserved.

  9. Simple method for purification of enterotoxigenic Escherichia coli fimbriae.

    Science.gov (United States)

    Curtis, Brittany; Grassel, Christen; Laufer, Rachel S; Sears, Khandra T; Pasetti, Marcela F; Barry, Eileen M; Simon, Raphael

    2016-03-01

    Enterotoxigenic Escherichia coli (ETEC) are endemic pathogens in the developing world. They frequently cause illness in travelers, and are among the most prevalent causes of diarrheal disease in children. Pathogenic ETEC strains employ fimbriae as adhesion factors to bind the luminal surface of the intestinal epithelium and establish infection. Accordingly, there is marked interest in immunoprophylactic strategies targeting fimbriae to protect against ETEC infections. Multiple strategies have been reported for purification of ETEC fimbriae, however none is ideal. Purification has typically involved the use of highly virulent wild-type strains. We report here a simple and improved method to purify ETEC fimbriae, which was applied to obtain two different Class 5 fimbriae types of clinical relevance (CFA/I and CS4) expressed recombinantly in E. coli production strains. Following removal from cells by shearing, fimbriae proteins were purified by orthogonal purification steps employing ultracentrifugation, precipitation, and ion-exchange membrane chromatography. Purified fimbriae demonstrated the anticipated size and morphology by electron microscopy analysis, contained negligible levels of residual host cell proteins, nucleic acid, and endotoxin, and were recognized by convalescent human anti-sera. Copyright © 2015 Elsevier Inc. All rights reserved.

  10. Purification of Sardine Fish Oil Through Degumming and Neutralization

    Directory of Open Access Journals (Sweden)

    Stephanie Bija

    2017-06-01

    Full Text Available The quality of sardine fish oil can be improved by purification method through the step of degumming and neutralization. The aimed of this this study was analysis characteristic of crude sardin fish oil and determined the best method of purification. Degumming was carried out using 30% water and salt at concentration 5%, 8%, 10% b/v. Neutralization process using  NaOH with 16°Be and bleaching using 5% Magnesol XL. All step of refining was done at 50°C, 60°C, 70°C, and 80°C. The result of analysis showed that sardine crude fish oil had 24.86% of palmitic acid as the highest fatty acid, heavy metal was not detected,dencity was 0.92 g/cm3 and viscocity was 51 cPs. The best treatment of purification method was degumming using 5% NaCl at 50°C with rendement 65.37±0.72%; free fatty acid (FFA 0.38±0.03%; peroxide (PV 1.07±0.12 mEq/kg; anisidine (p-AnV 15.18±0.16 mEq/kg; total oxidation value (TOTOX 17.31±0.39 mEq/kg; and clarity was 75.09± 1.20%.

  11. Preparative Purification of Liriodendrin from Sargentodoxa cuneata by Macroporous Resin

    Science.gov (United States)

    Li, Di-Hua; Wang, Yan; Lv, Yuan-Shan; Liu, Jun-Hong; Yang, Lei; Zhang, Shu-Kun; Zhuo, Yu-Zhen

    2015-01-01

    The preparative purification of liriodendrin from Sargentodoxa cuneata using macroporous resin combined with crystallization process was evaluated. The properties of adsorption/desorption of liriodendrin on eight macroporous resins were investigated systematically. X-5 resin was selected as the most suitable medium for liriodendrin purification. The adsorption of liriodendrin on X-5 resin fitted well with the pseudo-second-order kinetic model and Langmuir isotherm model. Dynamic adsorption/desorption tests were performed using a glass column packed with X-5 resin to optimize the separation process of liriodendrin. After one treatment with X-5 resin, the content of liriodendrin in the product was increased 48.73-fold, from 0.85% to 41.42%, with a recovery yield of 88.9%. 97.48% liriodendrin was obtained by further crystallization and determined by HPLC. The purified product possessed strong antioxidant activity. In conclusion, purification of liriodendrin might expend its further pharmacological researches and further applications in pharmacy. PMID:26236742

  12. Expression, purification, and characterization of formaldehyde dehydrogenase from Pseudomonas aeruginosa.

    Science.gov (United States)

    Zhang, Wangluo; Chen, Shuai; Liao, Yuanping; Wang, Dingli; Ding, Jianfeng; Wang, Yingming; Ran, Xiaoyuan; Lu, Daru; Zhu, Huaxing

    2013-12-01

    As a member of zinc-containing medium-chain alcohol dehydrogenase family, formaldehyde dehydrogenase (FDH) can oxidize toxic formaldehyde to less active formate with NAD(+) as a cofactor and exists in both prokaryotes and eukaryotes. Most FDHs are well known to be glutathione-dependent in the catalysis of formaldehyde oxidation, but the enzyme from Pseudomonas putida is an exception, which is independent of glutathione. To identify novel glutathione-independent FDHs from other bacterial strains and facilitate the corresponding structural and enzymatic studies, high-level soluble expression and efficient purification of these enzymes need to be achieved. Here, we present molecular cloning, expression, and purification of the FDH from Pseudomonas aeruginosa, which is a Gram-negative pathogenic bacterium causing opportunistic human infection. The FDH of P. aeruginosa shows high sequence identity (87.97%) with that of P. putida. Our results indicated that coexpression with molecular chaperones GroES, GroEL, and Tig has significantly attenuated inclusion body formation and improved the solubility of the recombinant FDH in Escherichiacoli cells. A purification protocol including three chromatographic steps was also established to isolate the recombinant FDH to homogeneity with a yield of ∼3.2 mg from 1L of cell culture. The recombinant P. aeruginosa FDH was properly folded and biologically functional, as demonstrated by the mass spectrometric, crystallographic, and enzymatic characterizations of the purified proteins. Copyright © 2013 Elsevier Inc. All rights reserved.

  13. Affinity purification of soluble lysosomal proteins for mass spectrometric identification.

    Science.gov (United States)

    Jaquinod, Sylvie Kieffer-; Chapel, Agnès; Garin, Jérôme; Journet, Agnøs

    2008-01-01

    This chapter describes the process of production, purification, separation, and mass spectrometry identification of soluble lysosomal proteins. The rationale for purification of these proteins resides in their characteristic sugar, the mannose-6-phosphate (M6P), which allows an easy purification by affinity chromatography on immobilized M6P receptor (MPR). The secretion of M6P proteins (essentially soluble lysosomal proteins) from cells in culture is induced by adding a weak base in the culture medium. Secreted proteins are ammonium sulfate precipitated, dialyzed, and loaded onto the immobilized MPR column. After specific elution and collection of the M6P proteins, these are resolved by either bidimensional or monodimensional gel electrophoresis (designated as 2-DE or 1-DE, respectively). Mass spectrometry analysis is performed on spots excised from the 2-DE gel, or on discrete bands covering altogether the whole length of the 1-DE gel lane: these spots or bands are in-gel digested with trypsin and protein identification is obtained, thanks to peptide mass fingerprints [provided by analysis of the digests by matrix-assisted laser desorption ionization-mass spectrometry (MALDI-MS)] or peptide amino acid sequences (provided by analysis of the digests by the coupling between liquid chromatography and tandem mass spectrometry, LC-MS/MS).

  14. Extraction, Separation, and Purification of Blueberry Anthocyanin Using Ethyl Alcohol

    Directory of Open Access Journals (Sweden)

    Zhe Gao

    2017-11-01

    Full Text Available Blueberry contains many substances that are important to the human body and can prevent cardiovascular diseases, protect the retina, and soften blood vessels. Anthocyanin, which is extracted from blueberry, can activate the retina, strengthen vision, reduce serum cholesterol, triglyceride and high-density lipoprotein, and protect cell nucleus tissues from radical oxidation; hence, blueberry is of importance to scientists from different countries. In this study, anthocyanin was extracted and separated from blueberry using ethyl alcohol to investigate the effects of factors, such as ethyl alcohol volume ratio on anthocyanin extraction and separation technologies. The extracting solution was then purified using the macroreticular resin purification method to investigate the effects of ethyl alcohol concentration and eluent dosage on anthocyanin extraction during purification. The research results demonstrated that 60 % ethyl alcohol volume fraction, 1 : 10 mass ratio of solid to liquid, and 60 °C ultrasonic temperature were the best conditions for anthocyanin extraction. The best purification conditions were 95 % ethyl alcohol, which had been acidized by 0.3 % hydrochloric acid and 70 ml of eluent. This work provides a reference for the application of ethyl alcohol in anthocyanin extraction.

  15. Purification of a Multidrug Resistance Transporter for Crystallization Studies

    Directory of Open Access Journals (Sweden)

    Kamela O. Alegre

    2015-03-01

    Full Text Available Crystallization of integral membrane proteins is a challenging field and much effort has been invested in optimizing the overexpression and purification steps needed to obtain milligram amounts of pure, stable, monodisperse protein sample for crystallography studies. Our current work involves the structural and functional characterization of the Escherichia coli multidrug resistance transporter MdtM, a member of the major facilitator superfamily (MFS. Here we present a protocol for isolation of MdtM to increase yields of recombinant protein to the milligram quantities necessary for pursuit of structural studies using X-ray crystallography. Purification of MdtM was enhanced by introduction of an elongated His-tag, followed by identification and subsequent removal of chaperonin contamination. For crystallization trials of MdtM, detergent screening using size exclusion chromatography determined that decylmaltoside (DM was the shortest-chain detergent that maintained the protein in a stable, monodispersed state. Crystallization trials of MdtM performed using the hanging-drop diffusion method with commercially available crystallization screens yielded 3D protein crystals under several different conditions. We contend that the purification protocol described here may be employed for production of high-quality protein of other multidrug efflux members of the MFS, a ubiquitous, physiologically and clinically important class of membrane transporters.

  16. Chitinase III in Euphorbia characias latex: Purification and characterization.

    Science.gov (United States)

    Spanò, Delia; Pospiskova, Kristyna; Safarik, Ivo; Pisano, Maria Barbara; Pintus, Francesca; Floris, Giovanni; Medda, Rosaria

    2015-12-01

    This paper deals with the purification of a class III endochitinase from Euphorbia characias latex. Described purification method includes an effective novel separation step using magnetic chitin particles. Application of magnetic affinity adsorbent noticeably simplifies and shortens the purification procedure. This step and the subsequently DEAE-cellulose chromatography enable to obtain the chitinase in homogeneous form. One protein band is present on PAGE in non-denaturing conditions and SDS-PAGE profile reveals a unique protein band of 36.5 ± 2 kDa. The optimal chitinase activity is observed at 50 °C, pH 5.0. E. characias latex chitinase is able to hydrolyze colloidal chitin giving, as reaction products, N-acetyl-D-glucosamine, chitobiose and chitotriose. Moreover, we observed that calcium and magnesium ions enhance chitinase activity. Finally, we cloned the cDNA encoding the E. characias latex chitinase. The partial cDNA nucleotide sequence contains 762 bp, and the deduced amino acid sequence (254 amino acids) is homologous to the sequence of several plant class III endochitinases. Copyright © 2015 Elsevier Inc. All rights reserved.

  17. Large-scale functional purification of recombinant HIV-1 capsid.

    Directory of Open Access Journals (Sweden)

    Magdeleine Hung

    Full Text Available During human immunodeficiency virus type-1 (HIV-1 virion maturation, capsid proteins undergo a major rearrangement to form a conical core that protects the viral nucleoprotein complexes. Mutations in the capsid sequence that alter the stability of the capsid core are deleterious to viral infectivity and replication. Recently, capsid assembly has become an attractive target for the development of a new generation of anti-retroviral agents. Drug screening efforts and subsequent structural and mechanistic studies require gram quantities of active, homogeneous and pure protein. Conventional means of laboratory purification of Escherichia coli expressed recombinant capsid protein rely on column chromatography steps that are not amenable to large-scale production. Here we present a function-based purification of wild-type and quadruple mutant capsid proteins, which relies on the inherent propensity of capsid protein to polymerize and depolymerize. This method does not require the packing of sizable chromatography columns and can generate double-digit gram quantities of functionally and biochemically well-behaved proteins with greater than 98% purity. We have used the purified capsid protein to characterize two known assembly inhibitors in our in-house developed polymerization assay and to measure their binding affinities. Our capsid purification procedure provides a robust method for purifying large quantities of a key protein in the HIV-1 life cycle, facilitating identification of the next generation anti-HIV agents.

  18. Review of Membranes for Helium Separation and Purification

    Science.gov (United States)

    Scholes, Colin A.; Ghosh, Ujjal K.

    2017-01-01

    Membrane gas separation has potential for the recovery and purification of helium, because the majority of membranes have selectivity for helium. This review reports on the current state of the research and patent literature for membranes undertaking helium separation. This includes direct recovery from natural gas, as an ancillary stage in natural gas processing, as well as niche applications where helium recycling has potential. A review of the available polymeric and inorganic membranes for helium separation is provided. Commercial gas separation membranes in comparable gas industries are discussed in terms of their potential in helium separation. Also presented are the various membrane process designs patented for the recovery and purification of helium from various sources, as these demonstrate that it is viable to separate helium through currently available polymeric membranes. This review places a particular focus on those processes where membranes are combined in series with another separation technology, commonly pressure swing adsorption. These combined processes have the most potential for membranes to produce a high purity helium product. The review demonstrates that membrane gas separation is technically feasible for helium recovery and purification, though membranes are currently only applied in niche applications focused on reusing helium rather than separation from natural sources. PMID:28218644

  19. The effect of water purification systems on fluoride content of drinking water

    OpenAIRE

    Prabhakar A; Raju O; Kurthukoti A; Vishwas T

    2008-01-01

    Objective: The purpose of the present study was to determine the effect of different water purification systems on the fluoride content of drinking water and to compare the efficacy of these water purification systems in reducing the fluoride content. Materials and Methods: Five different water purification systems were tested in this study. They were reverse osmosis, distillation, activated carbon, Reviva ® , and candle filter. The water samples in the study were of two types, viz, bo...

  20. Post-treatment sludge analyses and purification of paint effluent by coag-flocculation method

    National Research Council Canada - National Science Library

    Menkiti, M C; Ezemagu, I G; Nwoye, C I; Ejimofor, M I

    ...) for the purification of paint effluent (PE). Tympanotonos fuscatus shells (TFS) are of crustacean origin consisting mainly of chitin, calcium carbonate, entrained protein and other organic matrixes...

  1. The Borexino Solar Neutrino Experiment: Scintillator purification and surface contamination

    Science.gov (United States)

    Leung, Michael

    The Borexino Solar Neutrino Experiment will observe the monoenergetic (862 keV) 7Be neutrinos, produced in the solar reaction 7Be+e- →7 Li+nue. These neutrinos are the second most abundant species of solar neutrinos, with an expected flux at earth of 5 x 109/cm2/s. Using nu - e scattering in an aromatic liquid scintillator, Borexino will make the first real time measurement of the solar neutrino flux at energies less than 1 MeV. In addition to checking Standard Solar Model and neutrino oscillation predictions at low energies, Borexino will test the MSW vacuum-matter transition, luminosity constraint, and non-standard theories such as mass varying neutrinos. The Borexino detector will also be sensitive to supernova neutrinos, geoneutrinos, reactor neutrinos, and pep solar neutrinos. The pep measurement will tightly constrain the primary pp solar neutrino flux whose energy is below the Borexino threshold. With an expected rate of 35 events per day from solar 7Be neutrinos, the maximum tolerable background rate is one count per day. Removal of radioactive isotopes from the liquid scintillator is essential for the experiment's success and will be achieved with purification techniques including filtration, distillation, water extraction, nitrogen stripping, and silica gel adsorption. Results from small-scale purification efficiency tests are presented. Water extraction showed moderate but inadequate removal of 210Po which is a dominant background. Distillation reduced 210Po by a factor of more than 500. Online purification involves cycling over 300 m3 of scintillator from the detector though the purification plants. Flow patterns within the detector that influence the purification efficiency were determined with numerical simulations. Poor flow in the prototype Counting Test Facility showed effectively stagnant volumes within the detector. These are not present in the larger Borexino detector. Surface contamination in Borexino arises primarily from contact with

  2. Efficient biotinylation and single-step purification of tagged transcription factors in mammalian cells and transgenic mice.

    NARCIS (Netherlands)

    E. de Boer (Ernie); P. Rodriguez (Patrick); E. Bonte (Edgar); J. Krijgsveld (Jeroen); E. Katsantoni (Eleni); A.J.R. Heck (Albert); F.G. Grosveld (Frank); J. Strouboulis (John)

    2003-01-01

    textabstractProteomic approaches require simple and efficient protein purification methodologies that are amenable to high throughput. Biotinylation is an attractive approach for protein complex purification due to the very high affinity of avidin/streptavidin for biotinylated

  3. Tall fescue seed extraction and partial purification of ergot alkaloids

    Science.gov (United States)

    Bush, Lowell

    2014-12-01

    Many substances in the tall fescue/endophyte association (Schedonorus arundinaceus/Epichloë coenophiala) have biological activity. Of these compounds only the ergot alkaloids are known to have significant mammalian toxicity and the predominant ergot alkaloids are ergovaline and ergovalinine. Because synthetically produced ergovaline is difficult to obtain, we developed a seed extraction and partial purification protocol for ergovaline/ergovalinine that provided a biologically active product. Tall fescue seed was ground and packed into several different sized columns for liquid extraction. Smaller particle size and increased extraction time increased efficiency of extraction. Our largest column was a 114 × 52 × 61 cm (W×L×D) stainless steel tub. Approximately 150 kg of seed could be extracted in this tub. The extraction was done with 80% ethanol. When the solvent front migrated to bottom of the column, flow was stopped and seed was allowed to steep for at least 48 h. Light was excluded from the solvent from the beginning of this step to the end of the purification process. Following elution, ethanol was removed from the eluate by evaporation at room temperature. Resulting syrup was freeze-dried. About 80% recovery of alkaloids was achieved with 18-fold increase in concentration of ergovaline. Initial purification of the dried product was accomplished by extracting with hexane/water (6:1, v/v) and the hexane fraction was discarded. The aqueous fraction was extracted with chloroform, the aqueous layer discarded, after which the chloroform was removed with a resulting 20-fold increase of ergovaline. About 65% of the ergovaline was recovered from the chloroform residue for an overall recovery of 50%. The resultant partially purified ergovaline had biological activities in in vivo and in vitro bovine bioassays that approximate that of synthetic ergovaline.

  4. Fractionation for Biodiesel Purification Using Supercritical Carbon Dioxide

    Directory of Open Access Journals (Sweden)

    Chao-Yi Wei

    2014-02-01

    Full Text Available In recent years, biodegradable and alternative biodiesel has attracted increased attention worldwide. Producing biodiesel from biomass involves critical separation and purification technology. Conventional technologies such as gravitational settling, decantation, filtration, water washing, acid washing, organic solvent washing and absorbent applications are inefficient, less cost effective and environmentally less friendly. In this study supercritical carbon dioxide (SC-CO2 with few steps and a low environmental impact, was used for biodiesel fractionation from impure fatty acid methyl ester (FAME solution mixes. The method is suitable for application in a variety of biodiesel production processes requiring subsequent stages of purification. The fractionation and purification was carried out using continuous SC-CO2 fractionation equipment, consisting of three columns filled with stainless steel fragments. A 41.85% FAME content solution mix was used as the raw material in this study. Variables were a temperature range of 40–70 °C, pressure range of 10–30 MPa, SC-CO2 flow rate range of 7–21 mL/min and a retention time range of 30–90 min. The Taguchi method was used to identify optimal operating conditions. The results show that a separated FAME content of 99.94% was verified by GC-FID under optimal fractionation conditions, which are a temperature of 40 °C of, a pressure level of 30MPa and a flow rate of 7 mL/min of SC-CO2 for a retention time of 90 min.

  5. A Simple Slow-Sand Filter for Drinking Water Purification

    Directory of Open Access Journals (Sweden)

    K. O. Yusuf

    2017-04-01

    Full Text Available Water-borne diseases are commonly encountered when pathogen-contaminated water is consumed. In rural areas, water is usually obtained from ponds, open shallow wells, streams and rain water during rainy season. Rain water is often contaminated by pathogens due to unhygienic of physical and chemical conditions of the roofs thereby making it unsafe for consumption. A simple slow sand filter mechanism was designed and fabricated for purification of water in rural areas where electricity is not available to power water purification devices. Rain water samples were collected from aluminum roof, galvanized roof and thatched roof. The waters samples were allowed to flow through the slow sand filter. The values of turbidity, total dissolved solids, calcium, nitrite, faecal coliform and total coliform from unfiltered water through thatched roof were 0.92 NTU, 27.23 mg/l, 6 mg/l, 0.16 mg/l, 5cfu/100ml and 6.0 cfu/100ml, respectively while the corresponding values for slow sand filter from thatched roof were 0.01 NTU, 0.23 mg/l, 2.5 mg/l, 0.1 mg/l, 0 cfu/100ml and 0 cfu/100ml, respectively. The values of turbidity, total dissolved solid, nitrite, calcium, faecal coliform and total coliform from unfiltered water for aluminum roof were 0.82 NTU, 23.68 mg/l, 2.70 mg/l, 1.0 mg/l, 4 cfu/100ml and 4cfu/100ml, respectively while the corresponding values for slow sand filter were 0.01 NTU, 0.16 mg/l, 0.57 mg/l, 0.2 mg/l, 0 cfu/100ml and 0 cfu/100ml, respectively. The values obtained for galvanized roof were also satisfactory. The slow sand filter is recommended for used in rural areas for water purification to prevent risk of water-borne diseases.

  6. High-throughput protein purification under denaturating conditions by the use of cation exchange chromatography.

    Science.gov (United States)

    Alm, Tove; Steen, Johanna; Ottosson, Jenny; Hober, Sophia

    2007-06-01

    A high-throughput protein purification strategy using the polycationic Z(basic) tag has been developed. In order for the strategy to be useful both for soluble and less soluble proteins, a denaturating agent, urea, was used in all purification steps. First, four target proteins were genetically fused to the purification tag, Z(basic). These protein constructs were purified by cation exchange chromatography and eluted using a salt gradient. From the data achieved, a purification strategy was planned including stepwise elution to enable parallel protein purification using a laboratory robot. A protocol that includes all steps, equilibration of the chromatography resin, load of sample, wash, and elution, all without any manual handling steps, was handled by the laboratory robot. The program allows automated purification giving milligram amounts of pure recombinant protein of up to 60 cell lysates. In this study 22 different protein constructs, with different characteristics regarding pI and solubility, were successfully purified by the laboratory robot. The data show that Z(basic) can be used as a general purification tag also under denaturating conditions. Moreover, the strategy enables purification of proteins with different pI and solubility using ion exchange chromatography (IEXC). The procedure is highly reproducible and allows for high protein yield and purity and is therefore a good complement to the commonly used His(6)-tag.

  7. Purification of heat labile toxin from Bordetella pertussis vaccine strain 134 employed indigenous technology

    Directory of Open Access Journals (Sweden)

    K.C. Shivanandappa

    2016-06-01

    Conclusion: The B. pertussis HLT could be purified through two phase with G50 and DEAE, cost effective techniques, the G50 purification has reduced the bioburden problems during DEAE purification and at the same time the quality of the product was high.

  8. Using an FPLC to Promote Active Learning of the Principles of Protein Structure and Purification

    Science.gov (United States)

    Robinson, Rebekah L.; Neely, Amy E.; Mojadedi, Wais; Threatt, Katie N.; Davis, Nicole Y.; Weiland, Mitch H.

    2017-01-01

    The concepts of protein purification are often taught in undergraduate biology and biochemistry lectures and reinforced during laboratory exercises; however, very few reported activities allow students to directly gain experience using modern protein purification instruments, such as Fast Protein Liquid Chromatography (FPLC). This laboratory…

  9. Purification of a Recombinant Glutathione Transferase from the Causative Agent of Hydatidosis, "Echinococcus granulosus"

    Science.gov (United States)

    Fleitas, Andrea L.; Randall, Lía M.; Möller, Matías N.; Denicola, Ana

    2016-01-01

    This practical class activity was designed to introduce students to recombinant protein expression and purification. The principal goal is to shed light on basic aspects concerning recombinant protein production, in particular protein expression, chromatography methods for protein purification, and enzyme activity as a tool to evaluate purity and…

  10. Item Purification Does Not Always Improve DIF Detection: A Counterexample with Angoff's Delta Plot

    Science.gov (United States)

    Magis, David; Facon, Bruno

    2013-01-01

    Item purification is an iterative process that is often advocated as improving the identification of items affected by differential item functioning (DIF). With test-score-based DIF detection methods, item purification iteratively removes the items currently flagged as DIF from the test scores to get purified sets of items, unaffected by DIF. The…

  11. Exploiting interfacial water properties for desalination and purification applications.

    Energy Technology Data Exchange (ETDEWEB)

    Xu, Hongwu (Los Alamos National Laboratory, Los Alamos, NM); Varma, Sameer; Nyman, May Devan; Alam, Todd Michael; Thuermer, Konrad; Holland, Gregory P.; Leung, Kevin; Liu, Nanguo (University of New Mexico Albuquerque, NM); Xomeritakis, George K. (University of New Mexico Albuquerque, NM); Frankamp, Benjamin L.; Siepmann, J. Ilja (University of Minnesota, Minneapolis, MN); Cygan, Randall Timothy; Hartl, Monika A. (Los Alamos National Laboratory, Los Alamos, NM); Travesset, Alex (Iowa State University, Ames, IA); Anderson, Joshua A. (Iowa State University, Ames, IA); Huber, Dale L.; Kissel, David J. (University of New Mexico Albuquerque, NM); Bunker, Bruce Conrad; Lorenz, Christian Douglas; Major, Ryan C. (University of Minnesota, Minneapolis, MN); McGrath, Matthew J. (University of Minnesota, Minneapolis, MN); Farrow, Darcie; Cecchi, Joseph L. (University of New Mexico Albuquerque, NM); van Swol, Frank B.; Singh, Seema; Rempe, Susan B.; Brinker, C. Jeffrey; Clawson, Jacalyn S.; Feibelman, Peter Julian; Houston, Jack E.; Crozier, Paul Stewart; Criscenti, Louise Jacqueline; Chen, Zhu (University of New Mexico Albuquerque, NM); Zhu, Xiaoyang (University of Minnesota, Minneapolis, MN); Dunphy, Darren Robert (University of New Mexico Albuquerque, NM); Orendorff, Christopher J.; Pless, Jason D.; Daemen, Luke L. (Los Alamos National Laboratory, Los Alamos, NM); Gerung, Henry (University of New Mexico Albuquerque, NM); Ockwig, Nathan W.; Nenoff, Tina Maria; Jiang, Ying-Bing; Stevens, Mark Jackson

    2008-09-01

    A molecular-scale interpretation of interfacial processes is often downplayed in the analysis of traditional water treatment methods. However, such an approach is critical for the development of enhanced performance in traditional desalination and water treatments. Water confined between surfaces, within channels, or in pores is ubiquitous in technology and nature. Its physical and chemical properties in such environments are unpredictably different from bulk water. As a result, advances in water desalination and purification methods may be accomplished through an improved analysis of water behavior in these challenging environments using state-of-the-art microscopy, spectroscopy, experimental, and computational methods.

  12. Process for the biological purification of waste water

    DEFF Research Database (Denmark)

    1992-01-01

    Process for the biological purification of waste water by the activated sludge method, the waste water being mixed with recirculated sludge and being subjected to an anaerobic treatment, before the waste water thus treated is alternately subjected to anoxic and aerobic treatments and the waste...... water thus treated is led into a clarification zone for settling sludge, which sludge is recirculated in order to be mixed with the crude waste water. As a result, a simultaneous reduction of the content both of nitrogen and phosphorus of the waste water is achieved....

  13. Purification and characterization of osteopontin from human milk

    DEFF Research Database (Denmark)

    Sørensen, Steen; Justesen, Steen Just; Johnsen, Anders H

    2003-01-01

    Osteopontin (OPN) is expressed in many organs and tissues and has different biological properties related to different molecular forms in respect to size and posttranslational modifications. However, a purification procedure for authentic intact OPN as well as fragments of OPN from an accessible...... biological source is missing. A four-step procedure was used to purify OPN from human milk, based on its crystal growth inhibitory activity, including anion exchange chromatography, the elimination of casein, hydroxyapatite chromatography, and negative affinity chromatography. Purified OPN was further...

  14. Semiconductor Grade, Solar Silicon Purification Project. [photovoltaic solar energy conversion

    Science.gov (United States)

    Ingle, W. M.; Rosler, R. S.; Thompson, S. W.; Chaney, R. E.

    1979-01-01

    A low cost by-product, SiF4, is reacted with mg silicon to form SiF2 gas which is polymerized. The (SiF2)x polymer is heated forming volatile SixFy homologues which disproportionate on a silicon particle bed forming silicon and SiF4. The silicon analysis procedure relied heavily on mass spectroscopic and emission spectroscopic analysis. These analyses demonstrated that major purification had occured and some samples were indistinguishable from semiconductor grade silicon (except possibly for phosphorus). However, electrical analysis via crystal growth reveal that the product contains compensated phosphorus and boron.

  15. Systems and methods for separation and purification of products

    Energy Technology Data Exchange (ETDEWEB)

    Weiss, Michael Joseph; Gilliam, Ryan J.; Self, Kyle; Mariansky, Gal; Leclerc, Margarete K.; Shipchandler, Riyaz Mohammed; Nagar, Jacob

    2017-11-28

    There are provided methods and systems for an electrochemical cell including an anode and a cathode where the anode is contacted with a metal ion that converts the metal ion from a lower oxidation state to a higher oxidation state. The metal ion in the higher oxidation state is reacted with an unsaturated hydrocarbon and/or a saturated hydrocarbon to form products. Separation and/or purification of the products as well as of the metal ions in the lower oxidation state and the higher oxidation state, is provided herein.

  16. Engineered graphite oxide materials for application in water purification.

    Science.gov (United States)

    Gao, Wei; Majumder, Mainak; Alemany, Lawrence B; Narayanan, Tharangattu N; Ibarra, Miguel A; Pradhan, Bhabendra K; Ajayan, Pulickel M

    2011-06-01

    Retaining the inherent hydrophilic character of GO (graphite-oxide) nanosheets, sp(2) domains on GO are covalently modified with thiol groups by diazonium chemistry. The surface modified GO adsorbs 6-fold higher concentration of aqueous mercuric ions than the unmodified GO. "Core-shell" adsorbent granules, readily useable in filtration columns, are synthesized by assembling aqueous GO over sand granules. The nanostructured GO-coated sand retains at least 5-fold higher concentration of heavy metal and organic dye than pure sand. The research results could open avenues for developing low-cost water purification materials for the developing economies. © 2011 American Chemical Society

  17. Novel Hydrogen Purification Device Integrated with PEM Fuel Cells

    Energy Technology Data Exchange (ETDEWEB)

    Joseph Schwartz; Hankwon Lim; Raymond Drnevich

    2010-12-31

    A prototype device containing twelve membrane tubes was designed, built, and demonstrated. The device produced almost 300 scfh of purified hydrogen at 200 psig feed pressure. The extent of purification met the program target of selectively removing enough impurities to enable industrial-grade hydrogen to meet purity specifications for PEM fuel cells. An extrusion process was developed to produce substrate tubes. Membranes met several test objectives, including completing 20 thermal cycles, exceeding 250 hours of operating life, and demonstrating a flux of 965 scfh/ft2 at 200 psid and 400 C.

  18. Molecular Techniques for Dicistrovirus Detection without RNA Extraction or Purification

    Directory of Open Access Journals (Sweden)

    Jailson F. B. Querido

    2013-01-01

    Full Text Available Dicistroviridae is a new family of small, nonenveloped, and +ssRNA viruses pathogenic to both beneficial arthropods and insect pests as well. Triatoma virus (TrV, a dicistrovirus, is a pathogen of Triatoma infestans (Hemiptera: Reduviidae, one of the main vectors of Chagas disease. In this work, we report a single-step method to identify TrV, a dicistrovirus, isolated from fecal samples of triatomines. The identification method proved to be quite sensitive, even without the extraction and purification of RNA virus.

  19. Preparation and Characterization of Zeolite Membrane for Bioethanol Purification

    Directory of Open Access Journals (Sweden)

    Aprilina Purbasari

    2013-06-01

    Full Text Available The use of bioethanol as an alternative fuel with a purity of more than 99.5% wt has prompted research on bioethanol purification. One of the promising methods used for bioethanol purification is pervaporation membrane. This research is aimed to prepare and characterize zeolite membranes for pervaporation membrane. The membrane preparation consisted of two stages, namely support preparation and zeolite deposition on the support. In support preparation, α- alumina and kaolin with specific composition (50:30; 40:40; 50:30 was mixed with additives and water. After pugging and aging process, the mixture became paste and extruded into tubular shape. The tube was then calcined at temperature of 1250 °C for 3 hours. After that, zeolite 4A was deposited on the tubes using clear solution made of 10 %wt zeolite and 90 %wt water and heated at temperature of 80 °C for 3 hours. Furthermore, the resulting zeolite membranes was washed with deionized water for 5 minutes and dried in oven at temperature of 100 °C for 24 hours. Characterization of zeolite membranes included mechanical strength test, XRD, and SEM. In the mechanical strength test, the membrane sample with α- alumina:kaolin = 50:30 (membrane A has the highest mechanical strength of 46.65 N/mm2. Result of XRD analysis for the membrane A indicated that mullite and corundum phases were formed, which mullite phase was more dominant. Meanwhile the result of SEM analysis shows that zeolite crystals have been formed and covered the pores support, but the deposition of zeolite has not been optimal yet. The performance examination for bioethanol purification showed that the membrane could increase the purity of bioethanol from 95% to 98.5% wt. © 2013 BCREC UNDIP. All rights reservedReceived: 23rd October 2012; Revised: 15th February 2013; Accepted: 16th February 2013[How to Cite: Purbasari, A., Istirokhatun, T., Devi, A.M., Mahsunnah, L. , Susanto, H. (2013. Preparation and Characterization of Zeolite

  20. Purification and characterization of a soluble calnexin from human placenta

    DEFF Research Database (Denmark)

    Olsen, Dorthe T; Peng, Li; Træholt, Sofie D

    2013-01-01

    (sCnx) was consistently identified in a separate ion exchange chromatography peak. The sCnx was further purified and characterised. This showed that the protein had been cleaved after residue 472 (between Gln and Met), thus liberating it from the transmembrane and cytoplasmic parts of Cnx......Calreticulin (Crt) and calnexin (Cnx) are homologous endoplasmic reticulum (ER) chaperones involved in protein folding and quality control. Crt is a soluble ER luminal Mr 46 kDa protein and Cnx is a Mr 67kDa ER membrane protein. During purification of Crt from human placenta a soluble form of Cnx...

  1. Preparation and purification of 7-Iodoclonazepam for use in radioimmunoassay

    Energy Technology Data Exchange (ETDEWEB)

    Goddard, C.P.; Law, B.; Mason, P.A.; Stead, A.H.

    1986-04-01

    A method is described for the preparation and purification of 7-(/sup 125/I)-Iodoclonazepam (5-(o-Chlorophenyl)-2,3-Dihydro-7-(/sup 125/I)-Iodo-1H-1,4-Benzodiazepin-2-one). The structure was confirmed by mass spectrometry using 7-(/sup 127/I)iodoclonazepam prepared by the same method. 7-(/sup 125/I)-Iodoclonazepam binds well to a benzodiazepine antiserum. Although readily displaced by all the benzodiazepines commercially available in the UK, it is not displaced by structurally related nonbenzodiazepines except at very high concentrations. 7-(/sup 125/I) Iodoclonazepam should therefore be useful for the development of a screening radioimmunoassay (RIA) for benzodiazepines.

  2. Magnetic purification of curcumin from Curcuma longa rhizome by novel naked maghemite nanoparticles.

    Science.gov (United States)

    Magro, Massimiliano; Campos, Rene; Baratella, Davide; Ferreira, Maria Izabela; Bonaiuto, Emanuela; Corraducci, Vittorino; Uliana, Maíra Rodrigues; Lima, Giuseppina Pace Pereira; Santagata, Silvia; Sambo, Paolo; Vianello, Fabio

    2015-01-28

    Naked maghemite nanoparticles, namely, surface active maghemite nanoparticles (SAMNs), characterized by a diameter of about 10 nm, possessing peculiar colloidal stability, surface chemistry, and superparamagnetism, present fundamental requisites for the development of effective magnetic purification processes for biomolecules in complex matrices. Polyphenolic molecules presenting functionalities with different proclivities toward iron chelation were studied as probes for testing SAMN suitability for magnetic purification. Thus, the binding efficiency and reversibility on SAMNs of phenolic compounds of interest in the pharmaceutical and food industries, namely, catechin, tyrosine, hydroxytyrosine, ferulic acid, coumaric acid, rosmarinic acid, naringenin, curcumin, and cyanidin-3-glucoside, were evaluated. Curcumin emerged as an elective compound, suitable for magnetic purification by SAMNs from complex matrices. A combination of curcumin, demethoxycurcumin, and bis-demethoxycurcumin was recovered by a single magnetic purification step from extracts of Curcuma longa rhizomes, with a purity >98% and a purification yield of 45%, curcumin being >80% of the total purified curcuminoids.

  3. Isolation and Purification of Jacalin from Artocarpus Heterophyllus Lam

    Directory of Open Access Journals (Sweden)

    M.R. Othman

    2010-09-01

    Full Text Available This paper presents investigation results of saturation conditions needed for purification of jacalin lectin from the extract seeds of Artocarpus heterophyllus by ammonium precipitation and affinity chromatography on Galactose-Affi gel Hz. Three different aspects of parameters encompassing the percentage of saturation of ammonium sulfate precipitation, the presence of ammonium sulfate on Lowry method and the suitable galactose concentration for optimum elution of the protein from Galactose-Affi gel Hz were investigated. With three different sets of fractional saturation of jacalin purification using ammonium sulfate precipitation, the maximum yield of 0.463 g/g was achieved at 0-90% saturation range in the absence of dialysis. Maximum yield of 0.425 g/g was obtained at 30-60% and 0-90% saturation range in the presence of dialysis. The result from this work also indicates that excessive quantity of NH4SO4 interferes with Lowry method for protein determination substantially. The 0-90% saturation range was found to be more potentially appropriate for large scale application than 30-60% saturation, since the former involves only 1 step NH4SO4 addition. From the affinity chromatography, elution of 0.2 M galactose (in 0.15 M NaCl from Galactose-Affi gel Hz produced the maximum peak profile and jacalin concentration. A reduction or increase in galactose concentration of more than 0.2 M did not increase concentration of purified jacalin purified using this method.

  4. Expression and affinity purification of recombinant proteins from plants

    Science.gov (United States)

    Desai, Urvee A.; Sur, Gargi; Daunert, Sylvia; Babbitt, Ruth; Li, Qingshun

    2002-01-01

    With recent advances in plant biotechnology, transgenic plants have been targeted as an inexpensive means for the mass production of proteins for biopharmaceutical and industrial uses. However, the current plant purification techniques lack a generally applicable, economic, large-scale strategy. In this study, we demonstrate the purification of a model protein, beta-glucuronidase (GUS), by employing the protein calmodulin (CaM) as an affinity tag. In the proposed system, CaM is fused to GUS. In the presence of calcium, the calmodulin fusion protein binds specifically to a phenothiazine-modified surface of an affinity column. When calcium is removed with a complexing agent, e.g., EDTA, calmodulin undergoes a conformational change allowing the dissociation of the calmodulin-phenothiazine complex and, therefore, permitting the elution of the GUS-CaM fusion protein. The advantages of this approach are the fast, efficient, and economical isolation of the target protein under mild elution conditions, thus preserving the activity of the target protein. Two types of transformation methods were used in this study, namely, the Agrobacterium-mediated system and the viral-vector-mediated transformation system. Copyright 2002 Elsevier Science (USA).

  5. Production and Purification of Peroxidase from Aspergillus niger.

    Directory of Open Access Journals (Sweden)

    Mohammed A. Jebor

    2017-02-01

    Full Text Available This study was conducted in the laboratories of Biology Department, College of Science, which deals with isolation and purification of peroxidase and optimization of process parameters to achieve maximum yield of peroxidase by Aspergillus niger. Solid-state fermentation of Aspergillus niger was carried out for enhanced production of peroxidase using hydrogen peroxide as the substrate of enzyme maximum activity of the enzyme was achieved under optimum growth conditions. The optimum conditions were the isolated of Aspergillus niger from soil and growth in synthetic medium, it gave high titer of peroxidase activity, the fructose as carbon source, peptone as nitrogen source, after 12 days of incubation, incubation temperature 25 °C and pH = 6.5. Peroxidase purified in four purification steps; precipitation with 70% saturation of ammonium sulfate, step of dialysis, the third by ion exchange chromatography using DEAE-Cellulose and fourth by gel filtration throughout Sephadex G-100. The specific activity of the purified enzyme was 150U/mg with 7.75 folds. The peroxidase was shown to have molecular weight of 40kDa in SDS-PAGA and about 40kDa in gel filtration.The optimum pH and temperature for peroxidase activity 7 and 35 C0 respectively.

  6. Bacteriocin production and different strategies for their recovery and purification.

    Science.gov (United States)

    Garsa, Anita Kumari; Kumariya, Rashmi; Sood, S K; Kumar, Anil; Kapila, Suman

    2014-03-01

    Bacteriocins from lactic acid bacteria (LAB) are a diverse group of antimicrobial proteins/peptides, offering potential as biopreservatives, and exhibit a broad spectrum of antimicrobial activity at low concentrations along with thermal as well as pH stability in foods. High bacteriocin production usually occurs in complex media. However, such media are expensive for an economical production process. For effective use of bacteriocins as food biopreservatives, there is a need to have heat-stable wide spectrum bacteriocins produced with high-specific activity in food-grade medium. The main hurdles concerning the application of bacteriocins as food biopreservatives is their low yield in food-grade medium and time-consuming, expensive purification processes, which are suitable at laboratory scale but not at industrial scale. So, the present review focuses on the bacteriocins production using complex and food-grade media, which mainly emphasizes on the bacteriocin producer strains, media used, different production systems used and effect of different fermentation conditions on the bacteriocin production. In addition, this review emphasizes the purification processes designed for efficient recovery of bacteriocins at small and large scale.

  7. Production and purification of polyclonal antibody against melatonin hormone

    Directory of Open Access Journals (Sweden)

    Fooladsaz K

    1999-09-01

    Full Text Available Nowadays immunochemical techniques have played a very important and valuable role in quantitative and qualitative assays of liquid compounds of the body. Producing antibody against immunogenes is the first step to make immunochemical kits. In this study production and purification of polyclonal antibody against melatonin has been considered. This hormone which has several important functions in physiological conditions such as migraine, cirrhosis, mammary gland cancer and other diseases, is the most important pineal gland secretion. This gland is a circumventricular organ of brain and according to histological and anatomical studies, it is a high secretory organ, that secretes active biological substances like melatonin, oxytocin, serotonin and ect. In this study, melatonin has been considered as hapten and has become an immunogen by being linked to the bovine serum Albumin. Then, by the immunization of three white New Zeland rabbits that had the booster injections in regular intervals, the antibody titer was detected to be 1/2000, by using checkboard curves, and with the use of melatonin linked to penicillinase as a labeled antigen, the titer was detected 1/200. Finally an antibody with high purification rate has been obtained, which can be used in immunochemical assays like RIA, ELISA, and EIA.

  8. Purification and Characterization of Recombinant Deinococcus radiodurans RNA Polymerase.

    Science.gov (United States)

    Esyunina, D M; Kulbachinskiy, A V

    2015-10-01

    The radioresistant bacterium Deinococcus radiodurans is one of the most interesting models for studies of cell stress resistance. Analysis of the mechanisms of gene expression in D. radiodurans revealed some specific features of the transcription apparatus that might play a role in cell resistance to DNA-damaging conditions. In particular, RNA polymerase from D. radiodurans forms unstable promoter complexes and during transcription elongation has a much higher rate of RNA cleavage than RNA polymerase from Escherichia coli. Analysis of the structure and functions of D. radiodurans RNA polymerase is complicated due to the absence of convenient genetic systems for making mutations in the RNA polymerase genes and difficulties with enzyme purification. In this work, we developed a system for expression of D. radiodurans RNA polymerase in E. coli cells. We obtained an expression vector encoding all core RNA polymerase subunits and defined optimal conditions for the expression and purification of the RNA polymerase. It was found that D. radiodurans RNA polymerase has much higher rates of RNA cleavage than E. coli RNA polymerase under a wide range of conditions, including variations in the concentration of catalytic magnesium ions and pH values of the reaction buffer. The expression system can be used for further studies of the RNA cleavage reaction and the mechanisms of transcription regulation in D. radiodurans, including analysis of mutant RNA polymerase variants.

  9. Purification of biodiesel by choline chloride based deep eutectic solvent

    Science.gov (United States)

    Niawanti, Helda; Zullaikah, Siti; Rachimoellah, M.

    2017-05-01

    Purification is a crucial step in biodiesel production to meet the biodiesel standard. This study purified biodiesel using choline chloride based deep eutectic solvent (DES). DES was used to reduce unreacted oil and unsaponifiable matter in rice bran oil based biodiesel. The objective of this work was to study the effect of extraction time using DES on the content and yield of fatty acid methyl ester (FAME). Rice bran used in this work contains 16.49 % of oil with initial free fatty acids (FFA) of 44.75 %. Acid catalyzed methanolysis was employed to convert rice bran oil (RBO) into biodiesel under following operation conditions: T = 60 °C, t = 8 h, molar ratio of oil to methanol = 1/10, H2SO4 = 1% w/w of oil. Rice bran oil based biodiesel obtained contain 89.05 % of FAME with very low FFA content (0.05 %). DES was made from a mixture of choline chloride and ethylene glycol with molar ratio of 1/2. Molar ratio of crude biodiesel to DES were 1/2 and 1/4. Extraction time was varied from 15 minutes to 240 minutes at 30 °C. The highest FAME content was obtained after purification for 240 min. at molar ratio crude biodiesel to DES 1/4 was 96.60 %. This work shows that DES has potential to purify biodiesel from non-edible raw material, such as RBO.

  10. Partial Purification of a Tonoplast ATPase from Corn Coleoptiles 1

    Science.gov (United States)

    Mandala, Suzanne; Taiz, Lincoln

    1985-01-01

    The tonoplast ATPase from corn coleoptile membranes was solubilized using a two-step procedure consisting of a pretreatment with 0.15% (w/v) deoxycholate to remove 60% of the protein, and 40 millimolar octyl-glucoside to solubilize the ATPase. During ultracentrifugation, the solublized ATPase entered a linear sucrose gradient faster than the majority of the protein, resulting in an 11-fold purification over the initial specific activity. The partially purified ATPase was almost completely inhibited by KNO3 with an estimated Ki of 10 millimolar. The specific activity of the KNO3-sensitive ATPase was increased 29-fold during purification. N,N′-Dicyclohexylcarbodiimide also completely inhibited the ATPase with half-maximal effects at a concentration of 4 micromolar. Neither vanadate nor azide inhibited enzyme activity. The purified ATPase was stimulated by Cl− and preferred Mg-ATP as substrate. Analysis of frations from the sucrose gradient by sodium dodecyl sulfate-polyacrylamide gel electrophoresis led to the identification of two major polypeptides at 72,000 and 62,000 daltons which were best correlated with ATPase activity. Several minor bands also appeared to copurify with enzyme activity, but were less consistent. Radiation inactivation experiments with intact membranes indicated that the functional molecular size of the tonoplast ATPase was nearly 400,000 daltons. This suggests that the ATPase is composed of several polypeptides, possibly including the 72,000- and 62,000-dalton proteins. Images Fig. 4 PMID:16664239

  11. The purification mechanism of wastewater by underwater discharge

    Science.gov (United States)

    Kim, Kangil; Ma, Suk Hwal; Huh, Jin Young; Hong, Yong Cheol; National Fusion Research Institute Team; Chonbuk National University Team; Kwangwoon University Team; NPAC Team

    2016-09-01

    There is a continuing need for development of effective, cheap and environmentally friendly processes for purification of wastewater. In this regard, the plasmas can be a promising candidate for next-generation method to purify the wastewater. It is well known that the plasmas generate many reactive species and thus they are predominant for degradation of organic pollutants from water. In order to generate plasma in wastewater, the capillary electrodes are used with ac power supply. After plasma treatment, the coagulants are added to purify the wastewater. The efficiency of coagulation is significantly improved by plasma treatment of wastewater. These results may come from the reactions among radicals of plasma-treated water, electron reduction and oxidation of ions in waste water, and coagulant. In order to verify the hypothesis, we measured characteristics changes of water by underwater discharge. In this study, we propose the purification mechanism of wastewater by underwater discharge. We expect that the underwater discharge can be applied to purify wastewater in near future.

  12. PARTIAL PURIFICATION AND CHARACTERIZATION OF ALKALOPHILIC PROTEASE FROM PSEUDOMONAS AERUGINOSA

    Directory of Open Access Journals (Sweden)

    R. Satheeskumar

    2013-10-01

    Full Text Available Partial purification and characterization of alkalophilic protease production from Pseudomonas aeruginosa was isolated from the gut of marine and coastal waters shrimp Penaeus monodon. The protease production was assayed in submerged fermentation to produce maximum protease activity (423 ± 0.09 U/ml. The enzyme was precipitated with ammonium sulphate and partially purified by ion exchange chromatography through DEAE Sephadex A-50 column. In 10th fraction showed maximum protease activity (734 ± 0.18 U/ml with increase in purification fold. The molecular weight of protease from Pseudomonas aeruginosa was recorded as 60 kDa. The stability of protease was tested at various pH and temperature; it showed maximum protease activity at pH-9 and temperature 50ºC. Among the various surfactants tested for enzyme stability, maximum activity was retained in poly ethylene glycol. The compatibility of protease enzyme with various commercial detergents; the enzyme retained maximum protease activity in tide. The results are indicated that all these properties make the bacterial proteases are most suitable for wide industrial applications.

  13. Purification of genomic DNA with minimal contamination of proteins.

    Science.gov (United States)

    Hebron, Haroun R; Yang, Yu; Hang, Jun

    2009-12-01

    The purification is based on a set of solutions and a simple centrifugation procedure. Protocols are designed for an easy extraction and purification of genomic DNA from a wide range of samples, including whole blood, buffy coat, bone marrow, body fluids, buccal cells, tissues, mouse tails, etc. RBCs are lysed by dilution into a hypotonic solution. Tissues are broken down and digested by proteinase K in the presence of an anion detergent to release genomic DNA. After precipitation of the detergent and proteins, unique beads that bind proteins, lipids, and RNAs are added to achieve the supreme purity. Genomic DNA is then separated by alcohol precipitation. A proprietary nucleic acid precipitation reagent is used to enhance DNA recovery from low concentration samples. No DNA-binding beads or columns are used in the method, eliminating the problem of low yield and the risk of shearing of genomic DNA. The purified samples are free of proteins, lipids, salts, and RNA contamination. Purified samples are also stable for storage and suitable for all downstream applications.

  14. The Expanding Toolkit of Translating Ribosome Affinity Purification.

    Science.gov (United States)

    Dougherty, Joseph D

    2017-12-13

    Translating ribosome affinity purification is a method initially developed for profiling mRNA from genetically defined cell types in complex tissues. It has been applied both to identify target molecules in cell types that are important for controlling a variety of behaviors in the brain, and to understand the molecular consequences on those cells due to experimental manipulations, ranging from drugs of abuse to disease-causing mutations. Since its inception, a variety of methodological advances are opening new avenues of investigation. These advances include a variety of new methods for targeting cells for translating ribosome affinity purification by features such as their projections or activity, additional tags and mouse reagents increasing the flexibility of the system, and new modifications of the method specifically focused on studying the regulation of translation. The latter includes methods to assess cell type-specific regulation of translation in specific subcellular compartments. Here, I provide a summary of these recent advances and resources, highlighting both new experimental opportunities and areas for future technical development. Copyright © 2017 the authors 0270-6474/17/3712079-09$15.00/0.

  15. Development of tantalum–zirconium alloy for hydrogen purification

    Energy Technology Data Exchange (ETDEWEB)

    Kumar, Sanjay, E-mail: sanjay.barc@gmail.com [Fusion Reactor Materials Section, MG, BARC, Mumbai 85 (India); IAMR, Hiroshima University, Higashihiroshima 739-8530 (Japan); Singh, Anamika [GSASM Hiroshima University, Higashihiroshima 739-8530 (Japan); Jain, Uttam; Dey, Gautam Kumar [Fusion Reactor Materials Section, MG, BARC, Mumbai 85 (India)

    2016-11-01

    Highlights: • Terminal solid solubility of Ta increases with Zr addition. • Increase in lattice parameters of Ta due to Zr addition may be the possible reason. • Enhance H solubility could also be explained on the change in e-DOS of Ta–Zr alloys. • Ta–Zr alloys could be possible combination for hydrogen purification membrane. - Abstract: Terminal solid solubility of hydrogen in Ta–Zr alloys has been studied in connection with the development of tantalum based metallic membrane for hydrogen/tritium purification. The alloys were prepared by vacuum arc melting technique and subsequently cold rolled to 0.2 mm thickness. The terminal solid solubility of hydrogen in these cold rolled samples was investigated in a modified Sieverts apparatus. The terminal solid solubility of hydrogen was marginally increased with zirconium content. The change in the lattices parameter of tantalum upon zirconium addition and the higher affinity of zirconium for hydrogen as compared to tantalum could be the possible reasons.

  16. Quantifying the Effectiveness of Muon Detector Purification Systems

    Science.gov (United States)

    Mburu, Naomi; Guida, Roberto; Mandelli, Beatrice

    The experiments along the Large Hadron Collider (LHC) at the European Organization for Nuclear Research (CERN) were developed to study the fundamental particles that govern our universe. These experiments utilize gaseous detectors to track muons in the outermost portion of the experiment. For example, in the Compact Muon Solenoid (CMS) experiment three types of gaseous detectors are used as muon trackers: Resistive Plate Chambers (RPCs), Drift Tubes (DTs) and Cathode Strip Chambers (CSCs). RPCs use a gas mixture of Freon (C2H2F4) , sulfur hexafluoride (SF6) and isobutane (iC4H10) . The components of this gas mixture are both expensive and have high global warming potentials, so most of the gas mixture must be recycled and purified through a gas recirculation system. For this reason, RPCs employ purification systems that remove impurities due to the contamination and irradiation of the gas mixture that occur during normal operation of the LHC. Ion selective electrodes, gas chromatography, and mass spectrometry are set up and employed to study impurities produced in the RPCs and to quantify the ability of the purification systems to remove these impurities. National Science Foundation.

  17. Polarization entanglement purification for concatenated Greenberger-Horne-Zeilinger state

    Science.gov (United States)

    Zhou, Lan; Sheng, Yu-Bo

    2017-10-01

    Entanglement purification plays a fundamental role in long-distance quantum communication. In the paper, we put forward the first polarization entanglement purification protocol (EPP) for one type of nonlocal logic-qubit entanglement, i.e., concatenated Greenberger-Horne-Zeilinger (C-GHZ) state, resorting to the photon-atom interaction in low-quality (Q) cavity. In contrast to existing EPPs, this protocol can purify the bit-flip error and phase-flip error in both physic and logic level. Instead of measuring the photons directly, this protocol only requires to measure the atom states to judge whether the protocol is successful. In this way, the purified logic entangled states can be preserved for further application. Moreover, it makes this EPP repeatable so as to obtain a higher fidelity of logic entangled states. As the logic-qubit entanglement utilizes the quantum error correction (QEC) codes, which has an inherent stability against noise and decoherence, this EPP combined with the QEC codes may provide a double protection for the entanglement from the channel noise and may have potential applications in long-distance quantum communication.

  18. Purification of mammalian DNA repair protein XRCC1

    Energy Technology Data Exchange (ETDEWEB)

    Chen, I. [Univ. of California, Berkeley, CA (United States)

    1995-11-01

    Malfunctioning DNA repair systems lead to cancer mutations, and cell death. XRCC1 (X-ray Repair Cross Complementing) is a human DNA repair gene that has been found to fully correct the x-ray repair defect in Chinese hamster ovary (CHO) cell mutant EM9. The corresponding protein (XRCC1) encoded by this gene has been linked to a DNA repair pathway known as base excision repair, and affects the activity of DNA ligase III. Previously, an XRCC1 cDNA minigene (consisting of the uninterrupted coding sequence for XRCC1 protein followed by a decahistidine tag) was constructed and cloned into vector pET-16b for the purpose of: (1) overproduction of XRCC1 in both prokaryotic and eukaryotic cells; and (2) to facilitate rapid purification of XRCC1 from these systems. A vector is basically a DNA carrier that allows recombinant protein to be cloned and overexpressed in host cells. In this study, XRCC1 protein was overexpressed in E. coli and purified by immobilized metal affinity chromatography. Currently, the XRCC1 minigene is being inserted into a new vector [pET-26b(+)] in hopes to increase overexpression and improve purification. Once purified XRCC1 can be crystallized for structural studies, or studied in vitro for its biological function.

  19. Expression, purification and crystallization of a lyssavirus matrix (M) protein

    Energy Technology Data Exchange (ETDEWEB)

    Assenberg, René [Division of Structural Biology and Oxford Protein Production Facility, The Henry Wellcome Building for Genomic Medicine, Oxford University, Roosevelt Drive, Oxford OX3 7BN (United Kingdom); Delmas, Olivier [UPRE Lyssavirus Dynamics and Host Adaptation, WHO Collaborating Centre for Reference and Research on Rabies, Institut Pasteur, 28 Rue du Docteur Roux, 75724 Paris CEDEX 15 (France); Graham, Stephen C.; Verma, Anil; Berrow, Nick; Stuart, David I.; Owens, Raymond J. [Division of Structural Biology and Oxford Protein Production Facility, The Henry Wellcome Building for Genomic Medicine, Oxford University, Roosevelt Drive, Oxford OX3 7BN (United Kingdom); Bourhy, Hervé [UPRE Lyssavirus Dynamics and Host Adaptation, WHO Collaborating Centre for Reference and Research on Rabies, Institut Pasteur, 28 Rue du Docteur Roux, 75724 Paris CEDEX 15 (France); Grimes, Jonathan M., E-mail: jonathan@strubi.ox.ac.uk [Division of Structural Biology and Oxford Protein Production Facility, The Henry Wellcome Building for Genomic Medicine, Oxford University, Roosevelt Drive, Oxford OX3 7BN (United Kingdom)

    2008-04-01

    The expression, purification and crystallization of the full-length matrix protein from three lyssaviruses is described. The matrix (M) proteins of lyssaviruses (family Rhabdoviridae) are crucial to viral morphogenesis as well as in modulating replication and transcription of the viral genome. To date, no high-resolution structural information has been obtained for full-length rhabdovirus M. Here, the cloning, expression and purification of the matrix proteins from three lyssaviruses, Lagos bat virus (LAG), Mokola virus and Thailand dog virus, are described. Crystals have been obtained for the full-length M protein from Lagos bat virus (LAG M). Successful crystallization depended on a number of factors, in particular the addition of an N-terminal SUMO fusion tag to increase protein solubility. Diffraction data have been recorded from crystals of native and selenomethionine-labelled LAG M to 2.75 and 3.0 Å resolution, respectively. Preliminary analysis indicates that these crystals belong to space group P6{sub 1}22 or P6{sub 5}22, with unit-cell parameters a = b = 56.9–57.2, c = 187.9–188.6 Å, consistent with the presence of one molecule per asymmetric unit, and structure determination is currently in progress.

  20. Multimodal charge-induction chromatography for antibody purification.

    Science.gov (United States)

    Tong, Hong-Fei; Lin, Dong-Qiang; Chu, Wen-Ning; Zhang, Qi-Lei; Gao, Dong; Wang, Rong-Zhu; Yao, Shan-Jing

    2016-01-15

    Hydrophobic charge-induction chromatography (HCIC) has advantages of high capacity, salt-tolerance and convenient pH-controlled elution. However, the binding specificity might be improved with multimodal molecular interactions. New ligand W-ABI that combining tryptophan and 5-amino-benzimidazole was designed with the concept of mutimodal charge-induction chromatography (MCIC). The indole and benzimidazole groups of the ligand could provide orientated mutimodal binding to target IgG under neutral pH, while the imidazole groups could induce the electrostatic repulsion forces for efficient elution under acidic pH. W-ABI ligand was coupled successfully onto agarose gel, and IgG adsorption behaviors were investigated. High affinity to IgG was found with the saturated adsorption capacity of 70.4 mg/ml at pH 7, and the flow rate of mobile phase showed little impact on the dynamic binding capacity. In addition, efficient elution could be achieved at mild acidic pH with high recovery. Two separation cases (IgG separation from albumin containing feedstock and monoclonal antibody purification from cell culture supernatant) were verified with high purity and recovery. In general, MCIC with the specially-designed ligand is an expanding of HCIC with improved adsorption selectivity, which would be a potential alternative to Protein A-based capture for the cost-effective purification of antibodies. Copyright © 2015 Elsevier B.V. All rights reserved.

  1. Purification of metal electroplating waste waters using zeolites.

    Science.gov (United States)

    Alvarez-Ayuso, E; García-Sánchez, A; Querol, X

    2003-12-01

    The sorption behaviour of natural (clinoptilolite) and synthetic (NaP1) zeolites has been studied with respect to Cr(III), Ni(II), Zn(II), Cu(II) and Cd(II) in order to consider its application to purify metal finishing waste waters. The batch method has been employed using metal concentrations in solution ranged from 10 to 200 mg/l and solid/liquid ratios ranged from 2.5 to 10 g/l. The Langmuir model was found to describe well all sorption processes, allowing to establish metal sorption sequences from which the main retention mechanism involved for each metal has been inferred. Synthetic zeolite exhibited about 10 times greater sorption capacities (b(Cr)=0.838 mmol/g, b(Ni)=0.342 mmol/g, b(Zn)=0.499 mmol/g, b(Cu)=0.795 mmol/g, b(Cd)=0.452 mmol/g) than natural zeolite (b(Cr)=0.079 mmol/g, b(Ni)=0.034 mmol/g, b(Zn)=0.053 mmol/g, b(Cu)=0.093 mmol/g, b(Cd)=0.041 mmol/g), appearing, therefore, as most suitable to perform metal waste water purification processes. This mineral showed the same high sorption capacity values when used in the purification of metal electroplating waste waters.

  2. New research on bioregenerative air/water purification systems

    Science.gov (United States)

    Johnson, Anne H.; Ellender, R. D.; Watkins, Paul J.

    1991-01-01

    For the past several years, air and water purification systems have been developed and used. This technology is based on the combined activities of plants and microorganisms as they function in a natural environment. More recently, researchers have begun to address the problems associated with indoor air pollution. Various common houseplants are currently being evaluated for their abilities to reduce concentrations of volatile organic compounds (VOCS) such as formaldehyde and benzene. With development of the Space Exploration Initiative, missions will increase in duration, and problems with resupply necessitates implementation of regenerative technology. Aspects of bioregenerative technology have been included in a habitat known as the BioHome. The ultimate goal is to use this technology in conjunction with physicochemical systems for air and water purification within closed systems. This study continued the risk assessment of bioregenerative technology with emphasis on biological hazards. In an effort to evaluate the risk for human infection, analyses were directed at enumeration of fecal streptococci and enteric viruses with the BioHome waste water treatment system.

  3. Design of laboratory cyclone separator for biogas purification

    Directory of Open Access Journals (Sweden)

    Marián Fodora

    2013-01-01

    Full Text Available This article deals with calculation of a cyclone separator for biogas purification using physical and chemical methods. There is presented a methodology for determination of operating dimensions of the cyclone separator and description of principal features of the cyclone separator model. Calculations have been performed for the diameter of the cylindrical part of cyclone separator 175 mm and for the biogas volume flow rate 6.9∙10−5 m3∙s−1. The calculations can be used in practice for the design of cyclone separator depending on the flow rate of biogas, size of the biogas plants respectively. The developed cyclone separator has been used for the cleaning of biogas in operating conditions at the biogas plant in Kolinany (Slovakia. The presented method of biogas purification has been used for the removing of hydrogen sulphide, particulate matter and carbon dioxide from the raw biogas at the biogas plant. Removal of these undesirable impurities from the biogas is an important step in the production of a fully valued fuel, biomethane.

  4. [Separation and purification of the main glucosinolate from rapeseeds].

    Science.gov (United States)

    Zhou, Jinlan; Hu, Jianhu; Qiu, Aiyong

    2005-07-01

    A procedure for the preparative isolation of 1-S-[(1Z)-3-hydroxy-1-[(sulfooxy) imino]-4-pentenyl]-1-thio-beta-D-glucopyanose, potassium salt (progoitrin) from Brassica oleracea is reported. The major steps in this procedure were: (1) extraction of glucosinolates with methanol from Brassica oleracea; (2) separation and purification of glucosinolates by chromatographic column on alumina support; and (3) a follow-up reversed-phase separation by octadecyl (C18) silica support yielding progoitrin as the main content of glucosinolate. The structure of progoitrin was identified on its physicochemical properties by ultraviolet absorption spectrometry, infrared absorption, 'H nuclear magnetic resonance spectroscopy, mass spectrometry and elemental analysis. Spectroscopic data of the sample isolated were in agreement with those reported in the literature. The purity of progoitrin obtained was determined to be 99% by high performance liquid chromatography. The simplicity of the separation and purification procedure for glucosinolates and the high purity of progoitrin isolated would make the method an important reference for future research on glucosinolates and a favorable technique for future development.

  5. [Improved protein-A chromatography for monoclonal antibody purification].

    Science.gov (United States)

    Chen, Quan; Toh, Phyllicia; Hoi, Aina; Xian, Mo; Peng, Xinying; Yang, Yuansheng; Zhang, Haibo; Nian, Rui; Zhang, Wei

    2016-06-25

    Therapeutic monoclonal antibodies become the major product class within the biopharmaceutical market. Protein A as the first capture step is still dominant in current platforms for purification of monoclonal antibodies. In this study, we developed a new antibody harvest process that incorporates acidic treatment of cell harvest, demonstrating high process yield, improved clearance of host cell associated contaminants, like non-histone host cell protein, histone, DNA and heteroaggregates. Host protein contamination was reduced about 10-fold compared to protein A loaded with harvest clarified by centrifugation and microfiltration. Turbidity increase of eluted IgG upon pH neutralization was nearly eliminated. Residual levels of impurities in the protein A eluate were achieved that potentially meet requirements of drug substance and thus alleviate the burden for further impurities removal in subsequent chromatography steps. The mechanism of host cell associated contaminants removal during acidic treatment was also explored. After a polishing step by Capto adhere, host cell protein was reduced to less than 5 ppm, DNA less than 1 ppb, histone to undetectable level, heteroaggregates less than 0.01% with total IgG recovery around 87%. This efficient process can be easily integrated into current IgG purification platforms, and may overcome downstream processing challenges.

  6. A rapid Orthopoxvirus purification protocol suitable for high-containment laboratories.

    Science.gov (United States)

    Hughes, Laura; Wilkins, Kimberly; Goldsmith, Cynthia S; Smith, Scott; Hudson, Paul; Patel, Nishi; Karem, Kevin; Damon, Inger; Li, Yu; Olson, Victoria A; Satheshkumar, P S

    2017-05-01

    Virus purification in a high-containment setting provides unique challenges due to barrier precautions and operational safety approaches that are not necessary in lower biosafety level (BSL) 2 environments. The need for high risk group pathogen diagnostic assay development, anti-viral research, pathogenesis and vaccine efficacy research necessitates work in BSL-3 and BSL-4 labs with infectious agents. When this work is performed in accordance with BSL-4 practices, modifications are often required in standard protocols. Classical virus purification techniques are difficult to execute in a BSL-3 or BSL-4 laboratory because of the work practices used in these environments. Orthopoxviruses are a family of viruses that, in some cases, requires work in a high-containment laboratory and due to size do not lend themselves to simpler purification methods. Current CDC purification techniques of orthopoxviruses uses 1,1,2-trichlorotrifluoroethane, commonly known as Genetron®. Genetron® is a chlorofluorocarbon (CFC) that has been shown to be detrimental to the ozone and has been phased out and the limited amount of product makes it no longer a feasible option for poxvirus purification purposes. Here we demonstrate a new Orthopoxvirus purification method that is suitable for high-containment laboratories and produces virus that is not only comparable to previous purification methods, but improves on purity and yield. Published by Elsevier B.V.

  7. Waste water biological purification plants of dairy products industry and energy management

    Science.gov (United States)

    Stepanov, Sergey; Solkina, Olga; Stepanov, Alexander; Zhukova, Maria

    2017-10-01

    The paper presents results of engineering and economical comparison of waste water biological purification plants of dairy products industry. Three methods of purification are compared: traditional biological purification with the use of secondary clarifiers and afterpurification through granular-bed filters, biomembrane technology and physical-and-chemical treatment together with biomembrane technology for new construction conditions. The improvement of the biological purification technology using nitro-denitrification and membrane un-mixing of sludge mixture is a promising trend in this area. In these calculations, an energy management which is widely applied abroad was used. The descriptions of the three methods are illustrated with structural schemes. Costs of equipment and production areas are taken from manufacturers’ data. The research is aimed at an engineering and economical comparison of new constructions of waste water purification of dairy products industry. The experiment demonstrates advantages of biomembrane technology in waste water purification. This technology offers prospects of 122 million rubles cost saving during 25 years of operation when compared with of the technology of preparatory reagent flotation and of 13.7 million rubles cost saving compared to the option of traditional biological purification.

  8. Purification of crude glycerol from transesterification reaction of palm oil using direct method and multistep method

    Science.gov (United States)

    Nasir, N. F.; Mirus, M. F.; Ismail, M.

    2017-09-01

    Crude glycerol which produced from transesterification reaction has limited usage if it does not undergo purification process. It also contains excess methanol, catalyst and soap. Conventionally, purification method of the crude glycerol involves high cost and complex processes. This study aimed to determine the effects of using different purification methods which are direct method (comprises of ion exchange and methanol removal steps) and multistep method (comprises of neutralization, filtration, ion exchange and methanol removal steps). Two crude glycerol samples were investigated; the self-produced sample through the transesterification process of palm oil and the sample obtained from biodiesel plant. Samples were analysed using Fourier Transform Infrared Spectroscopy, Gas Chromatography and High Performance Liquid Chromatography. The results of this study for both samples after purification have showed that the pure glycerol was successfully produced and fatty acid salts were eliminated. Also, the results indicated the absence of methanol in both samples after purification process. In short, the combination of 4 purification steps has contributed to a higher quality of glycerol. Multistep purification method gave a better result compared to the direct method as neutralization and filtration steps helped in removing most excess salt, fatty acid and catalyst.

  9. Total chemical synthesis of proteins without HPLC purification.

    Science.gov (United States)

    Loibl, S F; Harpaz, Z; Zitterbart, R; Seitz, O

    2016-11-01

    The total chemical synthesis of proteins is a tedious and time-consuming endeavour. The typical steps involve solid phase synthesis of peptide thioesters and cysteinyl peptides, native chemical ligation (NCL) in solution, desulfurization or removal of ligation auxiliaries in the case of extended NCL as well as many intermediary and final HPLC purification steps. With an aim to facilitate and improve the throughput of protein synthesis we developed the first method for the rapid chemical total on-resin synthesis of proteins that proceeds without a single HPLC-purification step. The method relies on the combination of three orthogonal protein tags that allow sequential immobilization (via the N-terminal and C-terminal ends), extended native chemical ligation and release reactions. The peptide fragments to be ligated are prepared by conventional solid phase synthesis and used as crude materials in the subsequent steps. An N-terminal His6 unit permits selective immobilization of the full length peptide thioester onto Ni-NTA agarose beads. The C-terminal peptide fragment carries a C-terminal peptide hydrazide and an N-terminal 2-mercapto-2-phenyl-ethyl ligation auxiliary, which serves as a reactivity tag for the full length peptide. As a result, only full length peptides, not truncation products, react in the subsequent on-bead extended NCL. After auxiliary removal the ligation product is liberated into solution upon treatment with mild acid, and is concomitantly captured by an aldehyde-modified resin. This step allows the removal of the most frequently observed by-product in NCL chemistry, i.e. the hydrolysed peptide thioester (which does not contain a C-terminal peptide hydrazide). Finally, the target protein is released with diluted hydrazine or acid. We applied the method in the synthesis of 46 to 126 amino acid long MUC1 proteins comprising 2-6 copies of a 20mer tandem repeat sequence. Only three days were required for the parallel synthesis of 9 MUC1 proteins

  10. EM-TECHNOLOGY APPLICATION FOR MUNICIPAL WASTEWATERS PURIFICATION FROM BIOLOGICAL POLLUTANTS

    Directory of Open Access Journals (Sweden)

    Oksana Vovk

    2011-03-01

    Full Text Available Abstract. This article is devoted to the problem of municipal waste waters purification. The present daysituation with waste water treatment facilities in Ukraine, existed methods of waste waters purification andsearch for new ones are described. Much attention is paid to such kind of pollutants as microbiological andbacterial. A comparatively new method of sewage waters purification from biological contaminants andpossibilities to apply this method in Ukraine is presented in the article.Keywords: biological pollutants, disinfection, effective microorganisms, EM-technology, treatmentfacilities, wastewaters.

  11. Effect of Cryogenic Distillation and Chemical Absorption on the Argon Concentration in Krypton Purification

    Science.gov (United States)

    Wang, Zhiliang; Feng, Gaoping; Wang, Mingdong; Hong, Yanji

    2017-10-01

    Cryogenic distillation and chemical absorption are two important means of inert gas purification. Experiments show that the order of the two methods has an effect on the final purification result. In order to verify the relationship between the retention of argon and the order of the two methods, experimental verification was designed. At the same time, the numerical simulation was carried out by using the principle of low temperature distillation, and the influence results of the two methods on the argon content after gas purification was obtained.

  12. One-step hyperentanglement purification and hyperdistillation with linear optics.

    Science.gov (United States)

    Wang, Tie-Jun; Liu, Lei-Lei; Zhang, Ru; Cao, Cong; Wang, Chuan

    2015-04-06

    We investigate a new approach for achieving hyperdistillation and hyperentanglement purification operations simultaneously on two-photon systems, whose state is described by nonlocal hyperentangled Bell states on both the spatial mode and polarization degree of freedoms. Exploiting linear optics and local entanglement resource, the quantum nondemolition (QND) parity-checking measurement and the heralded two-qubit amplification could are key steps in our scheme. With the QND parity-checking measurement and heralded qubit amplification operations, both the bit-flip (phase-flip) errors caused by decoherence in noisy channels and the vacuum errors caused by the transmission losses can be corrected. We show that the proposed scheme provides a new solution to overcome the problem of photon losses and decoherence simultaneously, which could be achieved with current technologies.

  13. Recovery and purification of polysaccharides from microbial broth.

    Science.gov (United States)

    Johns, M R; Noor, E

    1991-04-01

    Current industrial practice to recover extracellular microbial polysaccharides from the broth usually requires dilution to permit cell removal followed by precipitation, typically using alcohol. This paper presents a discussion on the solvent precipitation of xanthan and the results of research performed to investigate the behaviour of xanthan solutions during membrane processing using a microporous membrane. Using crossflow microfiltration, flux rates of up to 120 L/m2h were achieved for pure xanthan solutions, with complete rejection of the polysaccharide by the membrane. The thin film model underpredicted flux for xanthan solutions. In fact, flux was independent of xanthan concentration up to 20-25 g/L, and strongly dependent on crossflow velocity. Considerable benefits in terms of purification and reduced solvent requirements can be obtained by the use of an intermediate crossflow microfiltration step during xanthan recovery.

  14. Electrospun magnetically separable calcium ferrite nanofibers for photocatalytic water purification

    Science.gov (United States)

    EL-Rafei, A. M.; El-Kalliny, Amer S.; Gad-Allah, Tarek A.

    2017-04-01

    Three-dimensional random calcium ferrite, CaFe2O4, nanofibers (NFs) were successfully prepared via the electrospinning method. The effect of calcination temperature on the characteristics of the as-spun NFs was investigated. X-ray diffraction analysis showed that CaFe2O4 phase crystallized as a main phase at 700 °C and as a sole phase at 1000 °C. Field emission scanning electron microscopy emphasized that CaFe2O4 NFs were fabricated with diameters in the range of 50-150 nm and each fiber was composed of 20-50 nm grains. Magnetic hysteresis loops revealed superparamagnetic behavior for the prepared NFs. These NFs produced active hydroxyl radicals under simulated solar light irradiation making them recommendable for photocatalysis applications in water purification. In the meantime, these NFs can be easily separated from the treated water by applying an external magnetic field.

  15. Easy and Rapid Purification of Highly Active Nisin

    Directory of Open Access Journals (Sweden)

    André Abts

    2011-01-01

    Full Text Available Nisin is an antimicrobial peptide produced and secreted by several L. lactis strains and is specifically active against Gram-positive bacteria. In previous studies, nisin was purified via cation exchange chromatography at low pH employing a single-step elution using 1 M NaCl. Here, we describe an optimized purification protocol using a five-step NaCl elution to remove contaminants. The obtained nisin is devoid of impurities and shows high bactericidal activity against the nisin-sensitive L. lactis strain NZ9000. Purified nisin exhibits an IC50 of ~3 nM, which is a tenfold improvement as compared to nisin obtained via the one-step elution procedure.

  16. Membranes with Surface-Enhanced Antifouling Properties for Water Purification

    Directory of Open Access Journals (Sweden)

    Nima Shahkaramipour

    2017-03-01

    Full Text Available Membrane technology has emerged as an attractive approach for water purification, while mitigation of fouling is key to lower membrane operating costs. This article reviews various materials with antifouling properties that can be coated or grafted onto the membrane surface to improve the antifouling properties of the membranes and thus, retain high water permeance. These materials can be separated into three categories, hydrophilic materials, such as poly(ethylene glycol, polydopamine and zwitterions, hydrophobic materials, such as fluoropolymers, and amphiphilic materials. The states of water in these materials and the mechanisms for the antifouling properties are discussed. The corresponding approaches to coat or graft these materials on the membrane surface are reviewed, and the materials with promising performance are highlighted.

  17. Purification of the neurotensin receptor from bovine brain

    Energy Technology Data Exchange (ETDEWEB)

    Mills, A.; Demoliou-Mason, C.D.; Barnard, E.A.

    1988-01-05

    The neurotensin receptor protein, solubilized with digitonin/asolectin from bovine cerebral cortex membranes, was purified to apparent homogeneity by affinity chromatography using immobilized neurotensin. The product exhibits saturable and specific binding of (3,11-tyrosyl-3,5-/sup 3/H) neurotensin with an apparent affinity (K/sub d/ = 5.5 nM) comparable to that measured in intact membranes and crude soluble extracts. The affinity-purified material, after reduction with 100 mM dithiothreitol, in denaturing gel electrophoresis showed a single polypeptide of M/sub r/ 72,000. Under nonreducing conditions the apparent M/sub r/, however, was 50,000, suggesting the presence of intramolecular disulfide bonds. The purified neurotensin receptor was judged to be homogenous, in that (i) only a single polypeptide was detectable; and (ii) the overall purification was 30,000-50,000-fold, giving a specific neurotensin-binding activity close to the theoretical maximum.

  18. Stability, purification, and applications of bromelain: A review.

    Science.gov (United States)

    de Lencastre Novaes, Letícia Celia; Jozala, Angela Faustino; Lopes, André Moreni; de Carvalho Santos-Ebinuma, Valéria; Mazzola, Priscila Gava; Pessoa Junior, Adalberto

    2016-01-01

    Bromelain is a cysteine protease found in pineapple tissue. Because of its anti-inflammatory and anti-cancer activities, as well as its ability to induce apoptotic cell death, bromelain has proved useful in several therapeutic areas. The market for this protease is growing, and several studies exploring various properties of this molecule have been reported. This review aims to compile this data, and summarize the main findings on bromelain in the literature to date. The physicochemical properties and stability of bromelain under different conditions are discussed. Several studies on the purification of bromelain from crude extracts using a wide range of techniques such as liquid-liquid extractions by aqueous two-phase system, ultrafiltration, precipitation, and chromatography, have been reported. Finally, the various applications of bromelain are presented. This review therefore covers the main properties of bromelain, aiming to provide an up-to-date compilation of the data reported on this enzyme. © 2015 American Institute of Chemical Engineers.

  19. Fructo-oligosaccharides: Production, Purification and Potential Applications.

    Science.gov (United States)

    Bali, Vandana; Panesar, Parmjit S; Bera, Manab B; Panesar, Reeba

    2015-01-01

    The nutritional and therapeutic benefits of prebiotics have attracted the keen interest of consumers and food processing industry for their use as food ingredients. Fructo-oligosaccharides (FOS), new alternative sweeteners, constitute 1-kestose, nystose, and 1-beta-fructofuranosyl nystose produced from sucrose by the action of fructosyltransferase from plants, bacteria, yeast, and fungi. FOS has low caloric values, non-cariogenic properties, and help gut absorption of ions, decrease levels of lipids and cholesterol and bifidus-stimulating functionality. The purified linear fructose oligomers are added to various food products like cookies, yoghurt, infant milk products, desserts, and beverages due to their potential health benefits. This review is focused on the various aspects of biotechnological production, purification and potential applications of fructo-oligosaccharides.

  20. Surfactin – A Review on Biosynthesis, Fermentation, Purification and Applications

    Directory of Open Access Journals (Sweden)

    Nikhil S. Shaligram

    2010-01-01

    Full Text Available Surfactin, a bacterial cyclic lipopeptide, is produced by various strains of Bacillus subtilis and is primarily recognized as one of the most effective biosurfactants. It has the ability to reduce surface tension of water from 72 to 27 mN/m at a concentration as low as 0.005 %. The structure of surfactin consists of seven amino acids bonded to the carboxyl and hydroxyl groups of a 14-carbon fatty acid. Surfactin possesses a number of biological activities such as the ability to lyse erythrocytes, inhibit clot formation, lyse bacterial spheroplasts and protoplasts, and inhibit cyclic 3',5-monophosphate diesterase. The high cost of production and low yields have limited its use in various commercial applications. Both submerged and solid-state fermentation have been investigated with the mutational approach to improve the productivity. In this review, current state of knowledge on biosynthesis of surfactin, its fermentative production, purification, analytical methods and biomedical applications is presented.

  1. Separation process design for isolation and purification of natural products

    DEFF Research Database (Denmark)

    Malwade, Chandrakant R.

    Natural products are defined as secondary metabolites produced by plants and form a vast pool of compounds with unlimited chemical and functional diversity. Many of these secondary metabolites are high value added chemicals that are frequently used as ingredients in food, cosmetics, pharmaceuticals...... and other consumer products. Therefore, process technology towards industrial scale production of such high value chemicals from plants has significant value. Natural products can be obtained in pure form via synthetic or semi-synthetic route, but due to their complicated nature these methods have not been...... developed to the extent of industrial production for majority of natural products. Thus, isolation and purification of such natural products from plants is the most viable way to obtain natural products in pure form. This PhD project is mainly concerned with the design of separation process to isolate...

  2. Characterization, Preparation, and Purification of Marine Bioactive Peptides

    Directory of Open Access Journals (Sweden)

    Xueqin Wang

    2017-01-01

    Full Text Available Marine bioactive peptides, as a source of unique bioactive compounds, are the focus of current research. They exert various biological roles, some of the most crucial of which are antioxidant activity, antimicrobial activity, anticancer activity, antihypertensive activity, anti-inflammatory activity, and so forth, and specific characteristics of the bioactivities are described. This review also describes various manufacturing techniques for marine bioactive peptides using organic synthesis, microwave assisted extraction, chemical hydrolysis, and enzymes hydrolysis. Finally, purification of marine bioactive peptides is described, including gel or size exclusion chromatography, ion-exchange column chromatography, and reversed-phase high-performance liquid chromatography, which are aimed at finding a fast, simple, and effective method to obtain the target peptides.

  3. Purification and structural characterization of LTP1 polypeptides from beer.

    Science.gov (United States)

    Jégou, S; Douliez, J P; Mollé, D; Boivin, P; Marion, D

    2000-10-01

    We report on the purification of lipid transfer proteins (LTP) from barley seeds and beer with the aim of investigating the chemical modifications that occur during the brewing process. In seeds, the well-known LTP of 9 kDa (LTP1) has been found together with a second form named LTPb that displays comparable amino acid composition but was not fully sequenced. These two forms have been recovered in beer with marked chemical modifications including disulfide bond reduction and rearrangement and especially glycation by Maillard reaction. The glycation is heterogeneous with variable amounts of hexose units bound to LTPs. Circular dichroism shows that glycated LTP1 having all their disulfide bridges reduced are totally unfolded. These results provide a first basis for understanding how barley LTPs become foam-promoting agents during the malting and brewing process.

  4. Influence of Water Salinity on Air Purification from Hydrogen Sulfide

    Directory of Open Access Journals (Sweden)

    Leybovych L.I.

    2015-12-01

    Full Text Available Mathematical modeling of «sliding» water drop motion in the air flow was performed in software package FlowVision. The result of mathematical modeling of water motion in a droplet with diameter 100 microns at the «sliding» velocity of 15 m/s is shown. It is established that hydrogen sulfide oxidation occurs at the surface of phases contact. The schematic diagram of the experimental setup for studying air purification from hydrogen sulfide is shown. The results of the experimental research of hydrogen sulfide oxidation by tap and distilled water are presented. The dependence determining the share of hydrogen sulfide oxidized at the surface of phases contact from the dimensionless initial concentration of hydrogen sulfide in the air has been obtained.

  5. Isolation, purification and antioxidant activities of polysaccharides from Grifola frondosa.

    Science.gov (United States)

    Chen, Gui-tang; Ma, Xue-mei; Liu, Sheng-to; Liao, Yan-li; Zhao, Guo-qang

    2012-06-05

    The crude polysaccharides (GFP) were isolated from the fruiting bodies of Grifola frondosa and purified by DEAE cellulose-52 chromatography and Sephadex G-100 size-exclusion chromatography in that order. Three main fractions, GFP-1, GFP-2 and GFP-3, were obtained through the isolation and purification steps. Then the antioxidant activities of these three fractions were investigated in vitro. The results showed that GFP-1, GFP-2 and GFP-3 possessed significant inhibitory effects on 1,1-diphenyl-2-picrylhydrazyl (DPPH) radical, hydroxyl radical and superoxide radical; their reducing power, ferrous ions chelating effect and the inhibition ability of the rat liver lipid oxidation where also strong. These results suggest that G. frondosa polysaccharides could be a suitable natural antioxidant and may be the functional foods for humans. Copyright © 2012 Elsevier Ltd. All rights reserved.

  6. Purification, biochemical and kinetic properties of recombinant Staphylococcus aureus lipase.

    Science.gov (United States)

    Horchani, Habib; Fendri, Ahmed; Louati, Hanen; Sayari, Adel; Gargouri, Youssef; Verger, Robert

    2012-01-01

    We have compared the purification procedures as well as the biochemical and kinetic properties of wild type (wt-SAL3), untagged recombinant (rec(-His)SAL3), and tagged recombinant (rec(+His)SAL3) purified forms of Staphylococcus aureus lipase (SAL3). We used the pH-stat method (with emulsified tributyrin and olive oil as substrates) and the monomolecular film technique (with the three dicaprin isomers spread in the form of monomolecular films at the air-water interface). The data obtained showed that the recombinant expression process as well as the presence of a his-tag at the N-terminus of recombinant SAL3 affects significantly many biochemical and catalytic properties. The effects of the heterologous expression process on the catalytic properties of the staphylococcal lipases are three times more deleterious than the presence of an N-terminal tag extension.

  7. Purification and characterization of acylation stimulating protein from porcine serum.

    Science.gov (United States)

    Zhang, Hong; Jacobi, Sheila K; Toombs, Candice F; Cianflone, Katherine H; Nersesian, Natalya; Sarath, Gautam; Miner, Jess L

    2002-07-01

    A method for purifying acylation stimulating protein (ASP) from porcine serum is described. The mRNA encoding ASP was cloned by reverse transcriptase-polymerase chain reaction which predicted a 76 residue peptide. Based on this sequence, we generated antisera to a C-terminal peptide (ASP(1-20)) which aided ASP purification. Identity of the purified protein was verified by N-terminal sequencing. The molecular mass of porcine ASP is 8926. Porcine ASP stimulated esterification of fatty acid into triacylglycerol in cultured human cells with potency similar to that of human ASP (twofold at 5 microM). Based on this evidence that ASP exists in porcine blood, and that it has acylation stimulating activity, we propose that ASP may play a role in regulation of energy storage in adipose tissue in the pig.

  8. Membranes with Surface-Enhanced Antifouling Properties for Water Purification

    Science.gov (United States)

    Shahkaramipour, Nima; Tran, Thien N.; Ramanan, Sankara; Lin, Haiqing

    2017-01-01

    Membrane technology has emerged as an attractive approach for water purification, while mitigation of fouling is key to lower membrane operating costs. This article reviews various materials with antifouling properties that can be coated or grafted onto the membrane surface to improve the antifouling properties of the membranes and thus, retain high water permeance. These materials can be separated into three categories, hydrophilic materials, such as poly(ethylene glycol), polydopamine and zwitterions, hydrophobic materials, such as fluoropolymers, and amphiphilic materials. The states of water in these materials and the mechanisms for the antifouling properties are discussed. The corresponding approaches to coat or graft these materials on the membrane surface are reviewed, and the materials with promising performance are highlighted. PMID:28273869

  9. A robust robotic high-throughput antibody purification platform.

    Science.gov (United States)

    Schmidt, Peter M; Abdo, Michael; Butcher, Rebecca E; Yap, Min-Yin; Scotney, Pierre D; Ramunno, Melanie L; Martin-Roussety, Genevieve; Owczarek, Catherine; Hardy, Matthew P; Chen, Chao-Guang; Fabri, Louis J

    2016-07-15

    Monoclonal antibodies (mAbs) have become the fastest growing segment in the drug market with annual sales of more than 40 billion US$ in 2013. The selection of lead candidate molecules involves the generation of large repertoires of antibodies from which to choose a final therapeutic candidate. Improvements in the ability to rapidly produce and purify many antibodies in sufficient quantities reduces the lead time for selection which ultimately impacts on the speed with which an antibody may transition through the research stage and into product development. Miniaturization and automation of chromatography using micro columns (RoboColumns(®) from Atoll GmbH) coupled to an automated liquid handling instrument (ALH; Freedom EVO(®) from Tecan) has been a successful approach to establish high throughput process development platforms. Recent advances in transient gene expression (TGE) using the high-titre Expi293F™ system have enabled recombinant mAb titres of greater than 500mg/L. These relatively high protein titres reduce the volume required to generate several milligrams of individual antibodies for initial biochemical and biological downstream assays, making TGE in the Expi293F™ system ideally suited to high throughput chromatography on an ALH. The present publication describes a novel platform for purifying Expi293F™-expressed recombinant mAbs directly from cell-free culture supernatant on a Perkin Elmer JANUS-VariSpan ALH equipped with a plate shuttle device. The purification platform allows automated 2-step purification (Protein A-desalting/size exclusion chromatography) of several hundred mAbs per week. The new robotic method can purify mAbs with high recovery (>90%) at sub-milligram level with yields of up to 2mg from 4mL of cell-free culture supernatant. Copyright © 2016 Elsevier B.V. All rights reserved.

  10. Nicotinamide riboside phosphorylase from beef liver: purification and characterization.

    Science.gov (United States)

    Imai, T; Anderson, B M

    1987-04-01

    Nicotinamide riboside phosphorylase (NR phosphorylase) from beef liver has been purified to apparent homogeneity at 300-fold purification with a 35% yield. Kinetic constants for the enzyme-catalyzed phosphorolysis were as follows Knicotinamide riboside, 2.5 +/- 0.4 mM; Kinorganic phosphate, 0.50 +/- 0.12 mM; Vmax, 410 +/- 30 X 10(-6) mol min-1 mg protein-1, respectively. The molecular weights of the native enzyme and subunit structure were determined to be 131,000 and 32,000, respectively, suggesting the beef liver NR phosphorylase to be tetrameric in structure and consistent with the presence of identical subunits. The amino acid composition was shown to be very similar to that reported for human erythrocyte purine-nucleoside phosphorylase but differing considerably from that found for rat liver purine-nucleoside phosphorylase. In addition to catalytic activity with nicotinamide riboside, the beef liver enzyme catalyzed a phosphorolytic reaction with inosine and guanosine exhibiting activity ratios, nicotinamide riboside:inosine: guanosine of 1.00:0.35:0.29, respectively. These ratios of activity remained constant throughout purification of the beef liver enzyme and no separation of these activities was detected. Phosphorolysis of nicotinamide riboside was inhibited competitively by inosine (Ki = 75 microM) and guanosine (Ki = 75 microM). Identical rates of thermal denaturation of the beef liver enzyme were observed when determined for the phosphorolysis of either nicotinamide riboside or inosine. These observations coupled with studies of pH and specific buffer effects indicate the phosphorolysis of nicotinamide riboside, inosine, and guanosine to be catalyzed by the same enzyme.

  11. Purification and Characterization of AsES Protein

    Science.gov (United States)

    Chalfoun, Nadia R.; Grellet-Bournonville, Carlos F.; Martínez-Zamora, Martín G.; Díaz-Perales, Araceli; Castagnaro, Atilio P.; Díaz-Ricci, Juan C.

    2013-01-01

    In this work, the purification and characterization of an extracellular elicitor protein, designated AsES, produced by an avirulent isolate of the strawberry pathogen Acremonium strictum, are reported. The defense eliciting activity present in culture filtrates was recovered and purified by ultrafiltration (cutoff, 30 kDa), anionic exchange (Q-Sepharose, pH 7.5), and hydrophobic interaction (phenyl-Sepharose) chromatographies. Two-dimensional SDS-PAGE of the purified active fraction revealed a single spot of 34 kDa and pI 8.8. HPLC (C2/C18) and MS/MS analysis confirmed purification to homogeneity. Foliar spray with AsES provided a total systemic protection against anthracnose disease in strawberry, accompanied by the expression of defense-related genes (i.e. PR1 and Chi2-1). Accumulation of reactive oxygen species (e.g. H2O2 and O2˙̄) and callose was also observed in Arabidopsis. By using degenerate primers designed from the partial amino acid sequences and rapid amplification reactions of cDNA ends, the complete AsES-coding cDNA of 1167 nucleotides was obtained. The deduced amino acid sequence showed significant identity with fungal serine proteinases of the subtilisin family, indicating that AsES is synthesized as a larger precursor containing a 15-residue secretory signal peptide and a 90-residue peptidase inhibitor I9 domain in addition to the 283-residue mature protein. AsES exhibited proteolytic activity in vitro, and its resistance eliciting activity was eliminated when inhibited with PMSF, suggesting that its proteolytic activity is required to induce the defense response. This is, to our knowledge, the first report of a fungal subtilisin that shows eliciting activity in plants. This finding could contribute to develop disease biocontrol strategies in plants by activating its innate immunity. PMID:23530047

  12. ELISA reagent coverage evaluation by affinity purification tandem mass spectrometry.

    Science.gov (United States)

    Henry, Scott M; Sutlief, Elissa; Salas-Solano, Oscar; Valliere-Douglass, John

    2017-10-01

    Host cell proteins (HCPs) must be adequately removed from recombinant therapeutics by downstream processing to ensure patient safety, product quality, and regulatory compliance. HCP process clearance is typically monitored by enzyme-linked immunosorbent assay (ELISA) using a polyclonal reagent. Recently, mass spectrometry (MS) has been used to identify specific HCP process impurities and monitor their clearance. Despite this capability, ELISA remains the preferred analytical approach due to its simplicity and throughput. There are, however, inherent difficulties reconciling the protein-centric results of MS characterization with ELISA, or providing assurance that ELISA has acceptable coverage against all process-specific HCP impurities that could pose safety or efficacy risks. Here, we describe efficient determination of ELISA reagent coverage by proteomic analysis following affinity purification with a polyclonal anti-HCP reagent (AP-MS). The resulting HCP identifications can be compared with the actual downstream process impurities for a given process to enable a highly focused assessment of ELISA reagent suitability. We illustrate the utility of this approach by performing coverage evaluation of an anti-HCP polyclonal against both an HCP immunogen and the downstream HCP impurities identified in a therapeutic monoclonal antibody after Protein A purification. The overall goal is to strategically implement affinity-based mass spectrometry as part of a holistic framework for evaluating HCP process clearance, ELISA reagent coverage, and process clearance risks. We envision coverage analysis by AP-MS will further enable a framework for HCP impurity analysis driven by characterization of actual product-specific process impurities, complimenting analytical methods centered on consideration of the total host cell proteome.

  13. Research of Biogas Purification Using Microalgae Monoraphidium Griffithii Suspension

    Directory of Open Access Journals (Sweden)

    Živilė Bingelytė

    2017-09-01

    Full Text Available Using biogas instead of fossil fuels decreases pollutants such as solid particles, sulphur dioxide, nitrogen oxides concentrations in the environment. Green energy and development of relevant infrastructure improves air quality considerably. Chemical, physical, biological methods are used for biogas purification. The main difficulties using biological methods are selection of suitable microorganisms’ suspensions and making optimal conditions in photobioreactor. Different origin and structure microalgae suspensions are used applying biological treatment methods. Monoraphidium griffithi, which is widespread in fresh water, has relatively high potential. Microalgae’ cultures absorb the main components of biogas – carbon dioxide (CO2 and hydrogen sulphide (H2S. Absorbtion processes are based on photosynthesis. Microalgae absorb specific components of biogas when there are suitable light source and nutrient solvent. The main purposes of the research are to asses emission of biogas using different substrates (chicken manure and wastewater sludge. Also, it was measured main physical and chemical characteristics of both substrates: acidicy, temperature, redox potential, conductivity, biohemical oxygen demand. According results of the research, emission from wastewater sludge is greater than from chicken manure so sludge was chosen in teh next stage of the research. The next stage – asssessment of purification efficienty using Monoraphidium grifftihii suspension. Raw biogas was supplied to photobioreacor (with microalgae suspension. Alterations of methane, carbon dioxide, oxygen, hydrogen sulphide concentrations were measured precisely. According to results concentration of methane in the beginning of the researc was 62%, after 35 days – 69%. Meanwhile carbon dioxide – 37% and 31% by analogy. Experimental research alows to assess Monoraphidium griffithi absorption capacity of ballast components. Results were compared to different scientists

  14. A Heparin Purification Process Removes Spiked Transmissible Spongiform Encephalopathy Agent.

    Science.gov (United States)

    Bett, Cyrus; Grgac, Ksenija; Long, Dianna; Karfunkle, Michael; Keire, David A; Asher, David M; Gregori, Luisa

    2017-05-01

    In 2000, bovine heparin was withdrawn from the US market for fear of contamination with bovine spongiform encephalopathy (BSE) agent, the cause of variant Creutzfeldt-Jakob disease in humans. Thus, US heparin is currently sourced only from pig intestines. Availability of alternative sources of crude heparin, a life-saving drug, would benefit public health. Bovine heparin is an obvious option, but BSE clearance by the bovine heparin manufacturing process should be evaluated. To this end, using hamster 263K scrapie as a surrogate for BSE agent, we applied a four-step bench-scale heparin purification protocol resembling a typical heparin manufacturing process to investigate removal of the spiked scrapie agent. We removed aliquots from each step and analyzed them for residual abnormal prion protein (PrPTSE) using a sensitive in vitro method, real-time quaking-induced conversion (RT-QuIC) assay, and for infectivity using animal bioassays. The purification process reduced infectivity by 3.6 log10 and removed PrPTSE, measured as seeding activity, by 3.4 log10. NaOH treatment was the most effective removal step tested. We also investigated NaOH at different concentrations and pH: the results showed that as much as 5.2 log10 of PrPTSE seeding activity was removed at pH 12.5. Thus, changes to the concentration, treatment time, and temperature of alkaline extraction might further improve removal. Our results, using a basic heparin manufacturing process, inform efforts to reintroduce safe bovine heparin in the USA.

  15. Robotic high-throughput purification of affinity-tagged recombinant proteins.

    Science.gov (United States)

    Wiesler, Simone C; Weinzierl, Robert O J

    2015-01-01

    Affinity purification of recombinant proteins has become the method of choice to obtain good quantities and qualities of proteins for a variety of downstream biochemical applications. While manual or FPLC-assisted purification techniques are generally time-consuming and labor-intensive, the advent of high-throughput technologies and liquid handling robotics has simplified and accelerated this process significantly. Additionally, without the human factor as a potential source of error, automated purification protocols allow for the generation of large numbers of proteins simultaneously and under directly comparable conditions. The delivered material is ideal for activity comparisons of different variants of the same protein. Here, we present our strategy for the simultaneous purification of up to 24 affinity-tagged proteins for activity measurements in biochemical assays. The protocol described is suitable for the scale typically required in individual research laboratories.

  16. Purification of burned gases of domestic wastes; Moderna purificacion de gases quemados de las basuras domesticas

    Energy Technology Data Exchange (ETDEWEB)

    Gottschalk, J.; Buttman, P.; Johansson, T.

    1997-09-01

    The author presents the technology to reduce the emission from the burned gases purification of domestic wastes combustion. The technology was demonstrated in Hobec, Denmark, and developed in Germany. (Author)

  17. Effect of purification followed by solubilization of receptor material on quantitative receptor assays for anticholinergic drugs

    NARCIS (Netherlands)

    Smisterova, J.; Ensing, K; de Zeeuw, R.A

    In order to optimize quantitative receptor assays for anticholinergics, the different receptor preparations resulting from the purification and the solubilization of the P2 pellet From the calf striatum were evaluated. The dissociation constants for two chemically different anticholinergics, the

  18. Use of reverse micellar systems for the extraction and purification of bromelain from pineapple wastes.

    Science.gov (United States)

    Umesh Hebbar, H; Sumana, B; Raghavarao, K S M S

    2008-07-01

    Reverse micellar systems of CTAB/isooctane/hexanol/butanol and AOT/isooctane are used for the extraction and primary purification of bromelain from crude aqueous extract of pineapple wastes (core, peel, crown and extended stem). The effect of forward as well as back extraction process parameters on the extraction efficiency, activity recovery and purification fold is studied in detail for the pineapple core extract. The optimized conditions for the extraction from core resulted in forward and back extraction efficiencies of 45% and 62%, respectively, using reverse micellar system of cationic surfactant CTAB. A fairly good activity recovery (106%) and purification (5.2-fold) of bromelain is obtained under these conditions. Reverse micellar extraction from peel, extended stem and crown using CTAB system resulted in purification folds of 2.1, 3.5, and 1.7, respectively. Extraction from extended stem using anionic surfactant AOT in isooctane did not yield good results under the operating conditions employed.

  19. Technological aspects of de-icing fluids purification and reclaiming process in the airports

    Directory of Open Access Journals (Sweden)

    А. П. Шутько

    2000-12-01

    Full Text Available Presented are the results of de-icing fluids use in Ukraine airports. Model of purification and utilization system also block-module unit for collection and reclaiming of de-icing fluids are developed

  20. Efficient purification of heating surfaces using explosion generators; Effizientere Heizflaechenabreinigung durch Explosionsgeneratoren

    Energy Technology Data Exchange (ETDEWEB)

    Steiner, Christian; Napp, Manfred [Explosion Power GmbH, Lenzburg (Germany); Gablinger, Helen

    2012-11-01

    The purification of steam generators is a permanent task in the operation of waste incinerators in order to enhance the efficiency and boiler availability as well as to reduce the corrosion. Beside the traditional technologies such as soot blowers or rapping gears the manual online explosive purification with gas-filled sacks are well-known. The idea of an automation of this kind of purification has enjoyed much resonance at the market. Since the launching of explosion generators nearly three years ago, 46 generators are put into operation. In 2011, at least additional 37 generators will be put into operation. Thus, practical experiences in the most different types of different steam generators are collected. The purification efficiency is performed against the direction of the flow of the flue gas.

  1. The treatment and purification of wool and mohair scouring wastes- a survey

    CSIR Research Space (South Africa)

    Mozes, TE

    1982-08-01

    Full Text Available in these streams. The aim of this survey is to provide a retrospect of the various processes for the treatment and purification of wool and mohair scouring wastes from 1874 to the present....

  2. Dual-tagging system for the affinity purification of mammalian protein complexes

    Energy Technology Data Exchange (ETDEWEB)

    Giannone, Richard J [ORNL; McDonald, W Hayes [ORNL; Hurst, Gregory {Greg} B [ORNL; Huang, Ying [ORNL; Wu, Jun [ORNL; Liu, Yie [National Institute on Aging, Baltimore; Wang, Yisong [ORNL

    2007-01-01

    Although affinity purification coupled with mass spectrometry (MS) provides a powerful tool to study protein-protein interactions, this strategy has encountered numerous difficulties when adapted to mammalian cells. Here we describe a Gateway{reg_sign}-compatible dual-tag affinity purification system that integrates regulatable expression, tetracysteine motifs, and various combinations of affinity tags to facilitate the cloning, detection, and purification of bait proteins and their interacting partners. Utilizing the human telomere binding protein TRF2 as a benchmark, we demonstrate bait protein recoveries upwards of approximately 16% from as little as 1-7 x 10{sup 7} cells and successfully identify known TRF2 interacting proteins, suggesting that our dual-tag affinity purification approach is a capable new tool for expanding the capacity to explore mammalian proteomic networks.

  3. Development and Performance Evaluation of a Ceramic Filter for Point-of-Use Water Purification

    National Research Council Canada - National Science Library

    Bukola Olalekan Bolaji; Olugbenga Oluseyi Akande

    2013-01-01

    In this work, a ceramic filter for point-of-use water purification was designed, fabricated and tested to evaluate its performance in filtering water to the World Health Organisation (WHO) standards...

  4. The Accompaniment of Purification and Assimilation in the Introduction of Shahnameh

    Directory of Open Access Journals (Sweden)

    hamidreza ardestanirostami

    2017-04-01

    Full Text Available Assimilation-purification is an argument has permanently been considered by theologians. Purification is to purify God from all human characteristics and assimilation is to attach human adjectives to God. In order to purify God, Ferdowsi has called God as 'the God of name' of which the name is only remained and in place of assimilation has called him as 'the God of place' which is the place where creation forms and the signs of God's names and adjectives come into being and God turns to be explicable. As some Zarathustra texts show, God, before creation, was only a name, but after creation changes to the owner of place which makes him identifiable. Ferdowsi, also, in the line ' he is beyond name, signs, and guesses' has had purification in his mind, but in the other line: 'the writer of selected substance' points at assimilation and calls God as a painter. This is how he accompanies assimilation with purification.

  5. MicroRNA-320a and microRNA-4496 attenuate Helicobacter pylori cytotoxin-associated gene A (CagA)-induced cancer-initiating potential and chemoresistance by targeting β-catenin and ATP-binding cassette, subfamily G, member 2.

    Science.gov (United States)

    Kang, Dong Woo; Yang, Eun Sun; Noh, Yu Na; Hwang, Won Chan; Jo, Se-Young; Suh, Young-Ah; Park, Won Sang; Choi, Kang-Yell; Min, Do Sik

    2017-04-01

    Infection with Helicobacter pylori is closely linked to an increased risk of gastric cancer. Although cytotoxin-associated gene A (CagA), a major virulence factor of H. pylori, is known to be a causal factor for gastric carcinogenesis, the molecular link between CagA and gastric cancer-initiating cell (CIC)-like properties remains elusive. Here, we demonstrate that CagA is required for increased expression of β-catenin and its target CIC markers via downregulation of microRNA (miR)-320a and miR-4496. CagA promoted gastric CIC properties and was responsible for chemoresistance. miR-320a and miR-4496 attenuated the in vitro self-renewal and tumour-initiating capacity of CagA-expressing CICs by targeting β-catenin. Moreover, miR-320a and miR-4496 decreased CagA-induced chemoresistance by targeting ATP-binding cassette, subfamily G, member 2 (ABCG2) at the transcriptional and post-transcriptional levels, respectively. Combination therapy with 5-fluorouracil and miR-320a/miR-4496 suppressed gastric tumourigenesis and metastatic potential in an orthotopic mouse model, probably via suppression of CagA-induced CIC properties and chemoresistance. Our results provide novel evidence that CIC properties, chemoresistance and tumourigenesis associated with H. pylori are linked to CagA-induced upregulation of β-catenin and ABCG2. These data provide novel insights into the molecular mechanisms of CagA-induced carcinogenisis and the therapeutic potential of of miR-320a and miR-4496. Copyright © 2016 Pathological Society of Great Britain and Ireland. Published by John Wiley & Sons, Ltd. Copyright © 2016 Pathological Society of Great Britain and Ireland. Published by John Wiley & Sons, Ltd.

  6. Production, purification and characterization of bacteriocin from Lactobacillus murinus AU06 and its broad antibacterial spectrum

    OpenAIRE

    Elayaraja, Sivaramasamy; Annamalai, Neelamegam; Mayavu, Packiyam; Balasubramanian, Thangavel

    2014-01-01

    Objective: To study the production, purification and characterization of bacteriocin from Lactobacillus murinus AU06 isolated from marine sediments and its broad spectrum of inhibition against fish pathogens. Methods: The selected strain was used in production, purification and characterized of bacteriocin. In addition, purified bacteriocin was tested for its antimicrobial activity against fish pathogens. Results: In the present study, the bacteriocin production was found to be higher a...

  7. A three step approach for the purification of alkaline phosphatase from non-pasteurized milk

    OpenAIRE

    Upadhyay, Lata Sheo Bachan; Verma, Nishant

    2014-01-01

    In this study, a three step purification of alkaline phosphatase from non-pasteurized milk has been described. It included cream extraction, n-butanol treatment and acetone precipitation. Different parameters such as buffer concentration, temperature, pH, substrate concentration, acetone and n-butanol treatment were optimized to maximize the enzyme activity. The enzyme was fruitfully purified up to homogeneity from the milk, with percentage recovery and fold purification of 56.17 and 17.67 re...

  8. An improved strategy for tandem affinity purification-tagging of Schizosaccharomyces pombe genes

    OpenAIRE

    Cipak, Lubos; Spirek, Mario; Novatchkova, Maria; Chen, Zhiming; Rumpf, Cornelia; Lugmayr, Wolfgang; Mechtler, Karl; Ammerer, Gustav; Csaszar, Edina; Gregan, Juraj

    2009-01-01

    Tandem affinity purification (TAP) is a method that allows rapid purification of native protein complexes. We developed an improved technique to fuse the fission yeast genes with a TAP tag. Our technique is based on tagging constructs that contain regions homologous to the target gene cloned into vectors carrying a TAP tag. We used this technique to design strategies for TAP-tagging of predicted Schizosaccharomyces pombe genes (http://mendel.imp.ac.at/Pombe_tagging/). To validate the approach...

  9. Optimization of Parameters for Purification of Jatropha Curcas Based Biodiesel Using Organic Adsorbents

    OpenAIRE

    Banga, sangita; Varshney, Pradeep; Kumar, Naveen; Pal, Madan

    2016-01-01

    This paper discusses purification of biodiesel by using organic adsorbents instead of traditional water washing technique. The efficiency of different organic adsorbents under different conditions was compared with each other as well as traditional water washed biodiesel for the purification of Jatropha Curcas based transesterified biodiesel. The proposed methodologies were based on the use of Amberlite BD10 DRY, Purolite PD 206and Tulison T-45BD as adsorbents. The response of each adsorbent ...

  10. Relationship Between Growth of AIgae and Water Purification in a Slow Sand Filter in Summe

    OpenAIRE

    中本, 信忠; 池田, 大介; 田口, 香代; 山本, 満寿夫; 松田, 卓也

    1995-01-01

    The effects of water depth on the growth of algae and on the purification capacity of water in slow sand filters in summer were studied. Filamentous algae grew well in a shallow filter pond. The higher removal rates of available nutrients and dissolved organic carbon in a raw water were observed in the filteration of a shallow filter pond. Importance of algae as a nutrient assimilator and as an oxygen producer in the purification process was discussed.

  11. Undulative induction electron accelerator for the waste and natural water purification systems

    CERN Document Server

    Kulish, Victor V; Gubanov, I V

    2001-01-01

    The project analysis of Undulative Induction Accelerator (EH - accelerator) for the waste and natural water purification systems is accomplished. It is shown that the use of the four-channel design of induction block and the standard set of auxiliary equipment (developed earlier for the Linear Induction Accelerators - LINACs) allow to construct commercially promising purification systems. A quality analysis of the accelerator is done and the optimal parameters are chosen taking into account the specific sphere of its usage.

  12. Engineering a recyclable elastin-like polypeptide capturing scaffold for non-chromatographic protein purification.

    Science.gov (United States)

    Liu, Fang; Chen, Wilfred

    2013-01-01

    Previously, we reported a non-chromatographic protein purification method exploiting the highly specific interaction between the dockerin and cohesin domains from Clostridium thermocellum and the reversible aggregation property of elastin-like polypeptide (ELP) to provide fast and cost-effective protein purification. However, the bound dockerin-intein tag cannot be completely dissociated from the ELP-cohesin capturing scaffold due to the high binding affinity, resulting in a single-use approach. In order to further reduce the purification cost by recycling the ELP capturing scaffold, a truncated dockerin domain with the calcium-coordinating function partially impaired was employed. We demonstrated that the truncated dockerin domain was sufficient to function as an effective affinity tag, and the target protein was purified directly from cell extracts in a single binding step followed by intein cleavage. The efficient EDTA-mediated dissociation of the bound dockerin-intein tag from the ELP-cohesin capturing scaffold was realized, and the regenerated ELP capturing scaffold was reused in another purification cycle without any decrease in the purification efficiency. This recyclable non-chromatographic based affinity method provides an attractive approach for efficient and cost-effective protein purification. © 2013 American Institute of Chemical Engineers.

  13. A chimeric affinity tag for efficient expression and chromatographic purification of heterologous proteins from plants

    Directory of Open Access Journals (Sweden)

    Frank eSainsbury

    2016-02-01

    Full Text Available The use of plants as expression hosts for recombinant proteins is an increasingly attractive option for the production of complex and challenging biopharmaceuticals. Tools are needed at present to marry recent developments in high-yielding gene vectors for heterologous expression with routine protein purification techniques. In this study we designed the Cysta-tag, a new purification tag for immobilized metal affinity chromatography (IMAC of plant-made proteins based on the protein-stabilizing fusion partner SlCYS8. We show that the Cysta-tag may be used to rapidly purify proteins under native conditions, and then be removed enzymatically to isolate the protein of interest. We also show that commonly used protease recognition sites for linking purification tags are differentially stable in leaves of the commonly used expression host Nicotiana benthamiana, with those linkers susceptible to cysteine proteases being less stable then serine protease-cleavable linkers. As an example we describe a Cysta-tag experimental scheme for the one-step purification of a clinically useful protein, human α1-antitrypsin, transiently expressed in N. benthamiana. With potential applicability to the variety of chromatography formats commercially available for IMAC-based protein purification, the Cysta-tag provides a convenient means for the efficient and cost-effective purification of recombinant proteins from plant tissues.

  14. Automated small‐scale protein purification and analysis for accelerated development of protein therapeutics

    Science.gov (United States)

    LeSaout, Xavier; Costioli, Matteo; Jordan, Lynn; Lambert, Jeremy; Beighley, Ross; Provencher, Laurel; McGuire, Kevin; Verlinden, Nico; Barry, Andrew

    2015-01-01

    Small‐scale protein purification presents opportunities for accelerated process development of biotherapeutic molecules. Miniaturization of purification conditions reduces time and allows for parallel processing of samples, thus offering increased statistical significance and greater breadth of variables. The ability of the miniaturized platform to be predictive of larger scale purification schemes is of critical importance. The PerkinElmer JANUS BioTx Pro and Pro‐Plus workstations were developed as intuitive, flexible, and automated devices capable of performing parallel small‐scale analytical protein purification. Preprogrammed methods automate a variety of commercially available ion exchange and affinity chromatography solutions, including miniaturized chromatography columns, resin‐packed pipette tips, and resin‐filled microtiter vacuum filtration plates. Here, we present a comparison of microscale chromatography versus standard fast protein LC (FPLC) methods for process optimization. In this study, we evaluated the capabilities of the JANUS BioTx Pro‐Plus robotic platform for miniaturized chromatographic purification of proteins with the GE ӒKTA Express system. We were able to demonstrate predictive analysis similar to that of larger scale purification platforms, while offering advantages in speed and number of samples processed. This approach is predictive of scale‐up conditions, resulting in shorter biotherapeutic development cycles and less consumed material than traditional FPLC methods, thus reducing time‐to‐market from discovery to manufacturing. PMID:27774045

  15. Superparamagnetic poly(methyl methacrylate) beads for nattokinase purification from fermentation broth.

    Science.gov (United States)

    Yang, Chengli; Xing, Jianmin; Guan, Yueping; Liu, Huizhou

    2006-09-01

    An effective method for purification of nattokinase from fermentation broth using magnetic poly(methyl methacrylate) (PMMA) beads immobilized with p-aminobenzamidine was proposed in this study. Firstly, magnetic PMMA beads with a narrow size distribution were prepared by spraying suspension polymerization. Then, they were highly functionalized via transesterification reaction with polyethylene glycol. The surface hydroxyl-modified magnetic beads obtained were further modified with chloroethylamine to transfer the surface amino-modified magnetic functional beads. The morphology and surface functionality of the magnetic beads were examined by scanning electron microscopy and Fourier transform infrared. An affinity ligand, p-aminobenzamidine was covalently immobilized to the amino-modified magnetic beads by the glutaraldehyde method for nattokinase purification directly from the fermentation broth. The purification factor and the recovery of the enzyme activity were found to be 8.7 and 85%, respectively. The purification of nattokinase from fermentation broth by magnetic beads only took 40 min, which shows a very fast purification of nattokinase compared to traditional purification methods.

  16. The effect of water purification systems on fluoride content of drinking water.

    Science.gov (United States)

    Prabhakar, A R; Raju, O S; Kurthukoti, A J; Vishwas, T D

    2008-03-01

    The purpose of the present study was to determine the effect of different water purification systems on the fluoride content of drinking water and to compare the efficacy of these water purification systems in reducing the fluoride content. Five different water purification systems were tested in this study. They were reverse osmosis, distillation, activated carbon, Reviva , and candle filter. The water samples in the study were of two types, viz, borewell water and tap water, these being commonly used by the people of Davangere City, Karnataka. The samples were collected before and after purification, and fluoride analysis was done using fluoride ion-specific electrode. The results showed that the systems based on reverse osmosis, viz, reverse osmosis system and Reviva showed maximum reduction in fluoride levels, the former proving to be more effective than the latter; followed by distillation and the activated carbon system, with the least reduction being brought about by candle filter. The amount of fluoride removed by the purification system varied between the system and from one source of water to the other. Considering the beneficial effects of fluoride on caries prevention; when drinking water is subjected to water purification systems that reduce fluoride significantly below the optimal level, fluoride supplementation may be necessary. The efficacy of systems based on reverse osmosis in reducing the fluoride content of water indicates their potential for use as defluoridation devices.

  17. High-throughput protein production and purification at the Seattle Structural Genomics Center for Infectious Disease.

    Science.gov (United States)

    Bryan, Cassie M; Bhandari, Janhavi; Napuli, Alberto J; Leibly, David J; Choi, Ryan; Kelley, Angela; Van Voorhis, Wesley C; Edwards, Thomas E; Stewart, Lance J

    2011-09-01

    The establishment of an efficient and reliable protein-purification pipeline is essential for the success of structural genomic projects. The SSGCID Protein Purification Group at the University of Washington (UW-PPG) has established a robust protein-purification pipeline designed to purify 400 proteins per year at a rate of eight purifications per week. The pipeline was implemented using two ÄKTAexplorer 100 s and four ÄKTAprimes to perform immobilized metal-affinity chromatography (IMAC) and size-exclusion chromatography. Purifications were completed in a period of 5 d and yielded an average of 53 mg highly purified protein. This paper provides a detailed description of the methods used to purify, characterize and store SSGCID proteins. Some of the purified proteins were treated with 3C protease, which was expressed and purified by UW-PPG using a similar protocol, to cleave non-native six-histidine tags. The cleavage was successful in 94% of 214 attempts. Cleaved proteins yielded 2.9% more structures than uncleaved six-histidine-tagged proteins. This 2.9% improvement may seem small, but over the course of the project the structure output from UW-PPG is thus predicted to increase from 260 structures to 318 structures. Therefore, the outlined protocol with 3C cleavage and subtractive IMAC has been shown to be a highly efficient method for the standardized purification of recombinant proteins for structure determination via X-ray crystallography.

  18. Sequential purification and crystal growth for the production of low cost silicon substrates

    Science.gov (United States)

    Liaw, M.; Daragona, F. S.

    The objective of this program is to identify and develop low cost processing for fabricating large grain size polycrystalline silicon substrates. Metallurigical grade silicon (MG-Si) which is low cost and abundant for industrial usage was chosen as starting material. However, MG-Si cannot be used directly as substrates for solar cell fabrication for the following reasons: (1) it contains 1 to 2 percent metallic impurities, and (2) it is produced as irregular shapes with a fine grain structure. Various purification techniques have been reported. The techniques being studied under this program use direct methods for the purification of MG-Si. The process uses sequential steps of purification followed by crystal growth. The steps of sequential purification include: (1) leaching of MG-Si charge, (2) phase separation of nonsoluble impurities from molten silicon, (3) reactive gas treatment of molten silicon, (4) liquid liquid extraction (called slagging), and (5) impurity redistribution using ingot pulling. All the purification steps, with the exception of step (1), are performed in a consecutive manner using a crystal puller. The purified ingots will be produced in a desired ingot dimension and further recrystallization is not necessary. The theory and experimental results for each purification technique are presented. The relative effectiveness of the various steps are assessed and the most import step(s) are recommended. Finally the electrical characteristics of solar cells built on a thin epitaxial layer deposited on single pulled MG-Si substrates are discussed and compared to single crystal substrates.

  19. Plasmid Vectors for Proteomic Analyses in Giardia: Purification of Virulence Factors and Analysis of the Proteasome

    Science.gov (United States)

    Stadelmann, Britta; Birkestedt, Sandra; Hellman, Ulf; Svärd, Staffan G.

    2012-01-01

    In recent years, proteomics has come of age with the development of efficient tools for purification, identification, and characterization of gene products predicted by genome projects. The intestinal protozoan Giardia intestinalis can be transfected, but there is only a limited set of vectors available, and most of them are not user friendly. This work delineates the construction of a suite of cassette-based expression vectors for use in Giardia. Expression is provided by the strong constitutive ornithine carbamoyltransferase (OCT) promoter, and tagging is possible in both N- and C-terminal configurations. Taken together, the vectors are capable of providing protein localization and production of recombinant proteins, followed by efficient purification by a novel affinity tag combination, streptavidin binding peptide–glutathione S-transferase (SBP-GST). The option of removing the tags from purified proteins was provided by the inclusion of a PreScission protease site. The efficiency and feasibility of producing and purifying endogenous recombinant Giardia proteins with the developed vectors was demonstrated by the purification of active recombinant arginine deiminase (ADI) and OCT from stably transfected trophozoites. Moreover, we describe the tagging, purification by StrepTactin affinity chromatography, and compositional analysis by mass spectrometry of the G. intestinalis 26S proteasome by employing the Strep II-FLAG–tandem affinity purification (SF-TAP) tag. This is the first report of efficient production and purification of recombinant proteins in and from Giardia, which will allow the study of specific parasite proteins and protein complexes. PMID:22611020

  20. The effect of water purification systems on fluoride content of drinking water

    Directory of Open Access Journals (Sweden)

    Prabhakar A

    2008-03-01

    Full Text Available Objective: The purpose of the present study was to determine the effect of different water purification systems on the fluoride content of drinking water and to compare the efficacy of these water purification systems in reducing the fluoride content. Materials and Methods: Five different water purification systems were tested in this study. They were reverse osmosis, distillation, activated carbon, Reviva ® , and candle filter. The water samples in the study were of two types, viz, borewell water and tap water, these being commonly used by the people of Davangere City, Karnataka. The samples were collected before and after purification, and fluoride analysis was done using fluoride ion-specific electrode. Results: The results showed that the systems based on reverse osmosis, viz, reverse osmosis system and Reviva ® showed maximum reduction in fluoride levels, the former proving to be more effective than the latter; followed by distillation and the activated carbon system, with the least reduction being brought about by candle filter. The amount of fluoride removed by the purification system varied between the system and from one source of water to the other. Interpretation and Conclusion: Considering the beneficial effects of fluoride on caries prevention; when drinking water is subjected to water purification systems that reduce fluoride significantly below the optimal level, fluoride supplementation may be necessary. The efficacy of systems based on reverse osmosis in reducing the fluoride content of water indicates their potential for use as defluoridation devices.

  1. Replacement of trifluoroacetic acid with HCl in the hydrophobic purification steps of pediocin PA-1: a structural effect.

    Science.gov (United States)

    Gaussier, Hélène; Morency, Hélène; Lavoie, Marc C; Subirade, Muriel

    2002-10-01

    Trifluoroacetic acid (TFA) is a purification contaminant associated with pediocin PA-1 that interferes with Fourier transform infrared spectroscopy structural analysis. As revealed by circular dichroism, its presence affects the structural folding of pediocin. Consequently, we propose a new pediocin PA-1 purification procedure using HCl instead of TFA in all of the hydrophobic steps. This procedural change does not affect the purification yield or the amount of pediocin PA-1 purified. Furthermore, removing HCl, as opposed to TFA, after purification is an easier procedure to carry out. In fact, the removal of TFA requires more experimentation and results in protein loss. Thus, HCl is a good alternative to TFA in pediocin PA-1 purification and can be extended to the purification of other proteins. We also show that TFA-induced structural modifications do not significantly affect the antimicrobial activity of pediocin PA-1.

  2. Influence of a water purification unit on the contamination level of salmonella in outcoming water and sludge

    OpenAIRE

    Jacob, Benoit; Korsak Koulagenko, Nicolas; Grooven, Bénédicte; Flament, Etienne; Daube, Georges

    2002-01-01

    Foodborne pathogens occasionally harboured in the gastro-intestinal tract of some domestic animals may be retrieved in slaughterhouses waste water and in sludge of water purification units. Salmonella, athogen common to man and Animals, is often used as a biological risk indicator. The aim of the present study was to assess effectiveness of a recent water purification unit by rapid and semi-quantitative detection of this micro-organism. The water purification unit collects waste water ...

  3. Organic hydrogels as potential sorbent materials for water purification

    Science.gov (United States)

    Linardatos, George; Bekiari, Vlasoula; Bokias, George

    2014-05-01

    Hydrogels are three-dimensional, hydrophilic, polymeric networks capable to adsorb large amounts of water or biological fluids. The networks are composed of homopolymers or copolymers and are insoluble due to the presence of chemical or physical cross-links. Depending on the nature of the structural units, swelling or shrinking of these gels can be activated by several external stimuli, such as solvent, heat, pH, electric stimuli. As a consequence, these materials are attractive for several applications in a variety of fields: drug delivery, muscle mimetic soft linear actuators, hosts of nanoparticles and semiconductors, regenerative medicine etc. Of special interest is the application of hydrogels for water purification, since they can effectively adsorb several water soluble pollutants such as metal ions, inorganic or organic anions, organic dyestaff, etc. In the present work, anionic hydrogels bearing negatively charged -COO- groups were prepared and investigated. These are based on the anionic monomer sodium acrylate (ANa) and the nonionic one N,N-dimethylacrylamide (DMAM). A series of copolymeric hydrogels (P(DMAM-co-ANax) were synthesized. The molar content x of ANa units (expressing the molar charged content of the hydrogel) varies from 0 (nonionic poly(N,N-dimethylacrylamide), PDMAM, hydrogel) up to 1 (fully charged poly(sodium acrylate), PANa, hydrogel). The hydrogels were used to extract organic or inorganic solutes from water. Cationic and anionic model dyes, as well as multivalent inorganic ions, have been studied. It is found that cationic dyes are strongly adsorbed and retained by the hydrogels, while adsorbance of anionic dyes was negligible. Both maximum adsorption and equilibrium binding constant depend on the chemical structure of the dye, the presence of functional chemical groups and the hydrophobic-hydrophilic balance. In the case of metal cations, adsorption depends mostly on the charge of the cation. In addition, crucial factors controlling

  4. New downstream processing strategy for the purification of monoclonal antibodies from transgenic tobacco plants.

    Science.gov (United States)

    Platis, D; Drossard, J; Fischer, R; Ma, J K-C; Labrou, N E

    2008-11-21

    Affinity chromatography on immobilized Protein A is the current method of choice for the purification of monoclonal antibodies (mAbs). Despite its widespread use it presents certain drawbacks, such as ligand instability, leaching, toxicity and high cost. In the present work, we report a new procedure for the purification of two human monoclonal anti-HIV (human immunodeficiency virus) antibodies (mAbs 2G12 and 4E10) from transgenic tobacco plants using stable and low cost chromatographic materials. The first step of the mAb 2G12 purification procedure is comprised of an aqueous two-phase partition system (ATPS) for the removal of polyphenols while providing an essential initial purification boost (2.01-fold purification). In the second step, mAb 2G12 was purified using cation-exchange chromatography (CEX) on S-Sepharose FF, by elution with 20mM sodium phosphate buffer pH 7.5, containing 0.1M NaCl. The eluted mAb was directly loaded onto an immobilized metal affinity chromatography column (IMAC, Zn(2+)-iminodiacetic acid-Sepharose 6B) and eluted by stepwise pH gradient. The proposed method offered 162-fold purification with 97.2% purity and 63% yield. Analysis of the antibody preparation by sodium dodecyl sulphate-polyacrylamide gel electrophoresis (SDS-PAGE), enzyme immunosorbent assay (ELISA) and western blot showed that the mAb 2G12 was fully active and free of degraded variants, polyphenols and alkaloids. The effectiveness of the present purification protocol was evaluated by using a second transgenic human monoclonal anti-HIV mAb 4E10. The results showed that the same procedure can be successfully used for the purification of mAb 4E10. In the case of mAb 4E10, the proposed method offered 148-fold purification with 96.2% purity and 36% yield. Therefore, the proposed protocol may be of generic use for the purification of mAbs from transgenic tobacco plants.

  5. An automated microscale chromatographic purification of virus-like particles as a strategy for process development.

    Science.gov (United States)

    Wenger, Marc D; Dephillips, Peter; Price, Colleen E; Bracewell, Daniel G

    2007-06-01

    The development of fermentation processes for recombinant vaccines requires optimizing expression while maintaining high product quality. Changes to cell fermentation conditions are typically evaluated following cell disruption, with expression levels quantified by immunoassay, liquid chromatography or enzyme activity. However, assay titres do not always predict the effects that intracellular aggregation, proteolysis, post-translational modifications and differences in relative impurity levels can have on purification yield and product purity. Furthermore, heterogeneity in the size and surface properties inherent in viral particles makes unit operations such as chromatography less predictable. In these cases, the purification procedure (or a mimic thereof) must be carried out to give accurate information on the impact of changes in fermentation conditions on purification process performance. This was demonstrated for the development of a recombinant vaccine against human papillomavirus produced in Saccharomyces cerevisiae, where the most informative feedback on fermentation variables was obtained by completing a multistep chromatographic purification to evaluate process yield and product purity. To increase the purification throughput and reduce labour, the chromatography was miniaturized 1000-fold from the laboratory scale using microlitre volumes of adsorbent in a pipette tip and automated on a robotic workstation. The microscale purification is shown to be predictive of the laboratory-scale purification in terms of yield and purity, while providing over a 10-fold increase in throughput and allowing for increased monitoring of fermentation processes. In addition, by reducing the volume of cells needed for this assessment, the fermentation can be correspondingly reduced in scale and carried out in parallel for additional throughput gains.

  6. Microbial polyhydroxyalkanote synthesis repression protein PhaR as an affinity tag for recombinant protein purification

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    Chen Guo

    2010-05-01

    Full Text Available Abstract Background PhaR which is a repressor protein for microbial polyhydroxyalkanoates (PHA biosynthesis, is able to attach to bacterial PHA granules in vivo, was developed as an affinity tag for in vitro protein purification. Fusion of PhaR-tagged self-cleavable Ssp DnaB intein to the N-terminus of a target protein allowed protein purification with a pH and temperature shift. During the process, the target protein was released to the supernatant while PhaR-tagged intein was still immobilized on the PHA nanoparticles which were then separated by centrifugation. Results Fusion protein PhaR-intein-target protein was expressed in recombinant Escherichia coli. The cell lysates after sonication and centrifugation were collected and then incubated with PHA nanoparticles to allow sufficient absorption onto the PHA nanoparticles. After several washing processes, self-cleavage of intein was triggered by pH and temperature shift. As a result, the target protein was released from the particles and purified after centrifugation. As target proteins, enhanced green fluorescent protein (EGFP, maltose binding protein (MBP and β-galactosidase (lacZ, were successfully purified using the PhaR based protein purification method. Conclusion The successful purification of EGFP, MBP and LacZ indicated the feasibility of this PhaR based in vitro purification system. Moreover, the elements used in this system can be easily obtained and prepared by users themselves, so they can set up a simple protein purification strategy by themselves according to the PhaR method, which provides another choice instead of expensive commercial protein purification systems.

  7. Purification and Partial characterization of Trypanosoma cruzi triosephosphate isomerase.

    Science.gov (United States)

    Bourguignon, S C; Meirelles, M N; Pacheco, R S; De Simone, S G

    1998-01-01

    The enzyme triosephosphate isomerase (TPI, EC 5.3.1.1) was purified from extracts of epimastigote forms of Trypanosoma cruzi. The purification steps included: hydrophobic interaction chromatography on phenyl-Sepharose, CM-Sepharose, and high performance liquid gel filtration chromatography. The CM-Sepharose material contained two bands (27 and 25 kDa) with similar isoelectric points (pI 9.3-9.5) which could be separated by gel filtration in high performance liquid chromatography. Polyclonal antibodies raised against the porcine TPI detected one single polypeptide on western blot with a molecular weight (27 kDa) identical to that purified from T. cruzi. These antibodies also recognized only one band of identical molecular weight in western blots of several other trypanosomatids (Blastocrithidia culicis, Crithidia desouzai, Phytomonas serpens, Herpertomonas samuelpessoai). The presence of only one enzymatic form of TPI in T. cruzi epimastigotes was confirmed by agarose gel activity assay and its localization was established by immunocytochemical analysis. The T. cruzi purified TPI (as well as other trypanosomatid' TPIs) is a dimeric protein, composed of two identical subunits with an approximate mw of 27,000 and it is resolved on two dimensional gel electrophoresis with a pI of 9.3. Sequence analysis of the N-terminal portion of the 27 kDa protein revealed a high homology to Leishmania mexicana and T. brucei proteins.

  8. Purification and Some Biological Properties of Asparaginase from Azotobacter vinelandii

    Science.gov (United States)

    Gaffar, S. A.; Shethna, Y. I.

    1977-01-01

    Asparaginase was found in the soluble fraction of cells of Azotobacter vinelandii, and its activity remained the same during growth of the organism in a nitrogen-free medium. The specific activity and the yield of A. vinelandii increased twofold in the presence of ammonium sulfate. Within limits, the temperature (30 to 37°C) and pH (6.5 to 8.0) of the medium showed little effect on the levels of enzyme activity. The enzyme was purified to near homogeneity by standard methods of enzyme purification, including affinity chromatography, and had optimum activity at pH 8.6 and 48°C. The approximate molecular weight was 84,000. The apparent Km value for the substrate was 1.1 × 10-4 M. Metal ions or sulfhydryl reagents were not required for enzyme activity. Cu2+, Zn2+, and Hg2+ showed concentration-dependent inhibition, whereas amino and keto acids had no effect on the enzyme activity. Asparaginase was stable when incubated with rat serum and ascites fluid. The enzyme had no effect on the membrane of sheep erythrocytes and did not inhibit the incorporation of radioactive precursors into deoxyribonucleic acid, ribonucleic acid, and protein in Yoshida ascites sarcoma cells. Asparaginase activity was not detected in the tumor cells. Images PMID:16345199

  9. Isolation, purification and characterization of 5'-phosphodiesterase from Aspergillus fumigatus.

    Directory of Open Access Journals (Sweden)

    Zhiting Luo

    Full Text Available 5'-Phosphodiesterase (5'-PDE catalyzes the hydrolysis of ribonucleic acid to obtain a mixture of ribonucleotides, such as 5'-guanosine monophosphate and 5'-adenosine monophosphate. In this study, a 5'-PDE was newly isolated and purified from Aspergillus fumigatus. Following purification, this enzyme exhibited a specific activity of 1036.76 U/mg protein, a molecular weight of 9.5 kDa, and an optimal temperature and pH for enzyme activity of 60°C and 5.0, respectively. However, its activity was partially inhibited by Fe3+, Cu2+, and Zn2+, but slightly improved by the presence of K+ and Na+. Additionally, chemical-modification experiments were also applied to investigate the structural information of 5'-PDE, in which the residues containing carboxyl and imidazole groups were essential for enzyme activity based on their localization in the 5'-PDE active site. Furthermore, purified 5'-PDE could specifically catalyze the synthesis of ribonucleotides with a Vmax 0.71 mmol/mg·min and a KM of 13.60 mg/mL.

  10. [Purification and properties of NAG A from human kidney].

    Science.gov (United States)

    Yoshida, K

    1992-08-01

    In the present paper, we have reported the purification procedures of N-acetyl-beta, D-glucosaminidase (NAG) A from human renal tissue as well as the enzymatic properties of NAG A. NAG A was purified to homogeneity by gel filtration methods using Sephacry S-400 and S-200, followed by affinity chromatography with TSK DEAE 5-PW. The final activity of the enzyme was 1001 U/ml protein which was 506.6-fold that of the crude extract (supernatant of 20,000 x G of the homogenate). The molecular weight of NAG A was 140 kDa, consisting of two subunits of 30 kDa and 57 kDa. The isoelectric point of the enzyme was 5.60. The optimal pH of the enzyme was between 4.7 and 4.9. The Km value of the enzyme for sodio-m-cresol sulfophtaleinyl-N-acetyl-beta, D-glucosaminide was found 0.177 x 10(-3) mol/l. Lectin affinity chromatographies using concanavalin A and wheat germagglutinin have demonstrated that major sugar-chains of the enzyme were the high mannose type and hybrid type with a fucose residue, and that a small amount of the complex type was contained.

  11. Expression, Purification, and Characterisation of Dehydroquinate Synthase from Pyrococcus furiosus

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    Leonardo Negron

    2011-01-01

    Full Text Available Dehydroquinate synthase (DHQS catalyses the second step of the shikimate pathway to aromatic compounds. DHQS from the archaeal hyperthermophile Pyrococcus furiosus was insoluble when expressed in Escherichia coli but was partially solubilised when KCl was included in the cell lysis buffer. A purification procedure was developed, involving lysis by sonication at 30∘C followed by a heat treatment at 70∘C and anion exchange chromatography. Purified recombinant P. furiosus DHQS is a dimer with a subunit Mr of 37,397 (determined by electrospray ionisation mass spectrometry and is active over broad pH and temperature ranges. The kinetic parameters are KM (3-deoxy-D-arabino-heptulosonate 7-phosphate 3.7 μM and kcat 3.0 sec-1 at 60∘C and pH 6.8. EDTA inactivates the enzyme, and enzyme activity is restored by several divalent metal ions including (in order of decreasing effectiveness Cd2+, Co2+, Zn2+, and Mn2+. High activity of a DHQS in the presence of Cd2+ has not been reported for enzymes from other sources, and may be related to the bioavailability of Cd2+ for P. furiosus. This study is the first biochemical characterisation of a DHQS from a thermophilic source. Furthermore, the characterisation of this hyperthermophilic enzyme was carried out at elevated temperatures using an enzyme-coupled assay.

  12. Design and optimisation of purification procedure for biodiesel washing

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    S.B. Glišić

    2009-09-01

    Full Text Available Almost complete methanolysis of triglycerides is usually not enough to fulfil the strict standards of biodiesel quality. A key step in this process is neutralization of alkali (catalyst followed by the washing procedure necessary for removing different impurities such as traces of catalyst and methanol and removal of soaps and glycerol from esters phase. The washing with hot water is still widely used in many industrial units for the biodiesel production. In this study, different procedures of biodiesel washing using hot water were investigated. The orto-phosphoric acid was suggested as the best compound for alkali catalyst (sodium hydroxide neutralization. The main goal of the performed analysis was to minimize the water usage in the washing-neutralization step during the biodiesel production. Such solution would make the process of biodiesel synthesis more economical taking into account the decrease of energy consumed for evaporation of water during the final product purification, as well as more acceptable procedure related to the impact on environment (minimal waste water release. Results of the performed simulation of the washing process supported by original experimental data suggested that neutralization after the optimized washing process of the methyl ester layer could be the best solution. The proposed washing procedure significantly decreases the amount of waste water giving at the same time the desired purity of final products (biodiesel and glycerol. The simulation of the process was performed using ASPEN plus software supported by ELCANTREL and UNIQUAC procedure of required properties calculation

  13. Large-scale crystallization of proteins for purification and formulation.

    Science.gov (United States)

    Hekmat, Dariusch

    2015-07-01

    Since about 170 years, salts were used to create supersaturated solutions and crystallize proteins. The dehydrating effect of salts as well as their kosmotropic or chaotropic character was revealed. Even the suitability of organic solvents for crystallization was already recognized. Interestingly, what was performed during the early times is still practiced today. A lot of effort was put into understanding the underlying physico-chemical interaction mechanisms leading to protein crystallization. However, it was understood that already the solvation of proteins is a highly complex process not to mention the intricate interrelation of electrostatic and hydrophobic interactions taking place. Although many basic questions are still unanswered, preparative protein crystallization was attempted as illustrated in the presented case studies. Due to the highly variable nature of crystallization, individual design of the crystallization process is needed in every single case. It was shown that preparative crystallization from impure protein solutions as a capture step is possible after applying adequate pre-treatment procedures like precipitation or extraction. Protein crystallization can replace one or more chromatography steps. It was further shown that crystallization can serve as an attractive alternative means for formulation of therapeutic proteins. Crystalline proteins can offer enhanced purity and enable highly concentrated doses of the active ingredient. Easy scalability of the proposed protein crystallization processes was shown using the maximum local energy dissipation as a suitable scale-up criterion. Molecular modeling and target-oriented protein engineering may allow protein crystallization to become part of a platform purification process in the near future.

  14. Extraction purification and characterization of trypsin inhibitors from Andean seeds

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    Patricio Castillo

    2017-09-01

    Full Text Available This work established the conditions of covalent immobilization of trypsin on a Sepharose matrix, which could be applied for the purification of trypsin inhibitors. The higher values of retention of enzymatic activity and immobilized enzymatic activity were obtained with a Sepharose 6B-CL matrix, at room temperature, a pH value of 10.5, an enzymatic load of 25 mg/mL, and a minimum immobilization time of 12 hours, in order to obtain a stable immobilization. The most active trypsin inhibitors were selected through the comparison of, extracts obtained from the seeds of amaranth (Amaranthus caudatus L., pea (Pisum sativum, lupine or “chocho” (Lupinus mutabilis, bean (Phaseolus vulgaris and “sangorache” (Amaranthus hybridus L.. The inhibitors were partially purified using centrifugal ultrafiltration, heat treatment, and TCA precipitation. The permeated and retained fractions of “sangorache” were selected as the most active trypsin inhibitors, and they were selectively purified using affinity chromatography in a Trypsin - Glyoxyl - Sepharose 6B-CL matrix. The kinetic characterization showed the presence of two inhibitors; the first one corresponded to a competitive inhibitor, while the second one behaved as a mixed inhibitor.

  15. Purification and renaturation of membrane neuraminidase from Haemophilus parasuis.

    Science.gov (United States)

    Lichtensteiger, Carol A; Vimr, Eric R

    2003-05-02

    Haemophilus parasuis, which causes polyserositis, polysynovitis, meningitis, septicemia, and pneumonia in pigs, has emerged as an increasing problem in modern swine production systems. Co-factors for and the pathogenesis of H. parasuis disease are not defined. One of the potential virulence factors of H. parasuis is its neuraminidase (sialidase). While purifying the H. parasuis neuraminidase from the membrane fraction, we developed a protocol to renature enzymatic activity after enzyme preparations were resolved electrophorectically in denaturing polyacrylamide gels. The H. parasuis neuraminidase co-resolved with recombinant neuraminidase of Vibrio cholera; thus its apparent molecular mass is 82 kilodalton (kDa). The H. parasuis neuraminidase was associated with the membrane fraction and the purification protocol removed over 99% of the H. parasuis cell protein while retaining over 90% of the neuraminidase activity. Purified protein will provide another avenue to clone the neuraminidase gene that has been refractory to cloning and the protocol will be a means to purify recombinant protein. Copyright 2003 Elsevier Science B.V.

  16. Introducing glycophage arrays: facile production, purification and patterning of glycophages.

    Science.gov (United States)

    Lajoie, Jason M; Shusta, Eric V

    2015-01-01

    Glycosylation is a widespread post-translational modification that plays important roles in health and disease. As glycan sequence and structure are not directly coded into the genome, our understanding of glycans and their functions in biological systems is much more primitive than that of DNA and proteins.Recently, printed glycan microarrays (glycoarrays) have emerged as powerful, high-throughput tools for screening glycan-protein interactions[1,2], and have been applied in disease detection [3], drug discovery [4], the study of immunity [5], and host-pathogen interactions [1, 2], among others.Unfortunately, glycoarray applications are currently limited by the expensive and complex methods available to synthesize glycans or alternatively, by the challenges in identifying and tagging glycans from natural sources [6, 7]. In this issue of Biotechnology Journal, Çelik et al. [8] introduce a potentially powerful new method for facile, scalable production, and purification of glycans compatible with microarray patterning. Çelik et al.’s [8] approach is based on innovative deployment of filamentous phage display so that the displayed proteins can be tagged with specific glycans of interest (glycophages) and subsequently patterned in array format.

  17. Extraction and partial purification of mouse uterine alkaline phosphatase.

    Science.gov (United States)

    Murdoch, R N; Kay, D J; Capper, W J

    1979-04-01

    Alkaline phosphatase in uterine homogenates from day 7 pregnant mice was solubilized using 0.2% (v/v) Triton X-100 and extracted wtih 20% (v/v) n-butanol. The procedure, which resulted in 182-fold purification, included ammonium sulfate precipitation, DEAE-cellulose anion exchange chromatography and Sephadex G200 gel filtration. Solubilization with Triton X-100 was an important step in the procedure since extraction with n-butanol alone only partially solubilized the enzyme and gave low extraction yields, much of the enzyme activity remaining in association with negatively charged residues. However, butanol extraction of Triton X-100-treated homogenates gave high yields of enzyme and eliminated p-nitrophenyl phosphatases which displayed activity in the pH range 3.0--7.5, together with a large proportion of inactive protein. The activity of the purified enzyme preparations was electrophoretically homogeneous on cellulose acetate membranes, suggesting that the alkaline phosphatase in the mouse uterus exists in a single isozymic form. Polyacrylamide-gel electrophoresis revealed that the purified preparations contained at least one protein as an impurity. Attempts to further purify the alkaline phosphatase by isoelectric focusing were unsuccessful since the enzyme was found to have an isoelectric point of about 5.0 and at this pH it was rapidly inactivated.

  18. Purification and properties of Cellvibrio gilvus cellobiose phosphorylase.

    Science.gov (United States)

    Sasaki, T; Tanaka, T; Nakagawa, S; Kainuma, K

    1983-03-01

    The cellobiose phosphorylase (EC 2.4.1.20) of Cellvibrio gilvus, which is an endocellular enzyme, has been purified 196-fold with a recovery of 11% and a specific activity of 27.4 mumol of glucose 1-phosphate formed/min per mg of protein. The purification procedure includes fractionation with protamine sulphate, and hydroxyapatite and DEAE-Sephadex A-50 chromatography. The enzyme appears homogeneous on polyacrylamide-gel electrophoresis, and a molecular weight of 280 000 was determined by molecular-sieve chromatography. Sodium dodecyl sulphate/polyacrylamide-gel electrophoresis revealed a single band and mol.wt. 72 000, indicating that cellobiose phosphorylase consists of four subunits. The enzyme had a specificity for cellobiose, requiring Pi and Mg2+ for phosphorylation, but not for cellodextrin, gentibiose, laminaribiose, lactose, maltose, kojibiose and sucrose. The enzyme showed low thermostability, an optimum pH of 7.6 and a high stability in the presence of 2-mercaptoethanol or dithiothreitol. The Km values for cellobiose and Pi were 1.25 mM and 0.77 mM respectively. Nojirimycin acted as a powerful pure competitive inhibitor (with respect to cellobiose) of the enzyme (Ki = 45 microM). Addition of thiol-blocking agents to the enzyme caused 56% inhibition at 500 microM-N-ethylmaleimide and 100% at 20 microM-p-chloromercuribenzoate.

  19. Synthesis, isolation and purification of [11C]-choline

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    Marcin Szydło

    2016-08-01

    Full Text Available [11C]-choline is an effective PET tracer used for imaging of neoplastic lesions and metastases of the prostate cancer. However, its production can be a challenge for manufacturers, as it has not yet been described in Polish or European pharmacopoeia. In this study the technical aspects of [11C]-choline production are described and detailed process parameters are provided. The quality control procedures for releasing [11C]-choline as solutio iniectabilis are also presented. The purity and quality of the radiopharmaceutical obtained according to the proposed method were find to be high enough to safely administrate the radiopharmaceutical to patients. Application of an automated synthesizer makes it possible to carry out the entire process of [11C]-choline production, isolation and purification within 20 minutes. It is crucial to maintain all aspects of the process as short as possible, since the decay half-time of carbon-11 is 20.4 minutes. The resulting radiopharmaceutical is sterile and pyrogen-free and of a high chemical, radiochemical, and radionuclide purity proved by chromatographic techniques. The yield of the process is up to 20%. [11C]-choline PET scanning can be used as accurate and effective diagnostic tool in all centers equipped with [11C]-target containing cyclotron.

  20. Purification of dihydropyridine receptor from rabbit skeletal muscle

    Energy Technology Data Exchange (ETDEWEB)

    Nakayama, N.; Vaghy, P.; Schwartz, A.

    1986-05-01

    Dihydropyridine (DHP)receptor was purified from T-tubules isolated from freeze-thawed rabbit skeletal muscle after French press treatment of microsomal membranes. DHP receptor was labeled with 25 nM (/sup 3/H)-(+)-PN-200-110 (PN, one of the most potent Ca-antagonists) and solubilized with 1% digitonin. The solubilized receptor was purified in the presence of protease inhibitors (0.1 mM PMSF, 1 mM iodoacetamide, 1..mu..M pepstatin A, 1 mg/l antipain and 0.2 mM o-phenanthroline) using WGA-Sepharose and DEAE-Biogel A column chromatography as well as sucrose density gradient (SDG) centrifugation. The pooled fractions of the SDG had a maximum binding of 590 pmol/mg protein even without correcting for dissociation of PN from the receptors during purification. On SDS-PAGE, a single major band (191 K dalton) was shown both in presence and absence of 20 mM N-ethyl maleimide. However, two major (145 and 103 K dalton) a few minor bands (55,46,32 and 31K dalton) were obtained if the fraction was treated with 20 mM dithiothreitol prior to electrophoresis. The authors data suggest that 191 and 145 k dalton proteins correspond to the ..cap alpha..-subunit of the DHP receptor as reported by Curtis and Catterall.

  1. Simplified, enhanced protein purification using an inducible, autoprocessing enzyme tag.

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    Aimee Shen

    2009-12-01

    Full Text Available We introduce a new method for purifying recombinant proteins expressed in bacteria using a highly specific, inducible, self-cleaving protease tag. This tag is comprised of the Vibrio cholerae MARTX toxin cysteine protease domain (CPD, an autoprocessing enzyme that cleaves exclusively after a leucine residue within the target protein-CPD junction. Importantly, V. cholerae CPD is specifically activated by inositol hexakisphosphate (InsP(6, a eukaryotic-specific small molecule that is absent from the bacterial cytosol. As a result, when His(6-tagged CPD is fused to the C-terminus of target proteins and expressed in Escherichia coli, the full-length fusion protein can be purified from bacterial lysates using metal ion affinity chromatography. Subsequent addition of InsP(6 to the immobilized fusion protein induces CPD-mediated cleavage at the target protein-CPD junction, releasing untagged target protein into the supernatant. This method condenses affinity chromatography and fusion tag cleavage into a single step, obviating the need for exogenous protease addition to remove the fusion tag(s and increasing the efficiency of tag separation. Furthermore, in addition to being timesaving, versatile, and inexpensive, our results indicate that the CPD purification system can enhance the expression, integrity, and solubility of intractable proteins from diverse organisms.

  2. Isolation and purification of beta-lactoglobulin from cow milk

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    Ranjit Aich

    2015-05-01

    Full Text Available Aim: The present study was undertaken to standardize a convenient method for isolation and purification of β-lactoglobulin (β-lg from cow milk keeping its antigenicity intact, so that the purified β-lg can be used for detection of cow milk protein intolerance (CMPI. Materials and Methods: Raw milk was collected from Gir breed of cattle reared in Haringhata Farm, West Bengal. Milk was then converted to skimmed milk by removing fat globules and casein protein was removed by acidification to pH 4.6 by adding 3 M HCl. β-lg was isolated by gel filtration chromatography using Sephacryl S-200 from the supernatant whey protein fraction. Further, β-lg was purified by anion-exchange chromatography in diethylaminoethyl-sepharose. Molecular weight of the purified cattle β-lg was determined by 15 percent one-dimensional sodium dodecyl sulfate-polyacrylamide gel electrophoresis and was analyzed by gel documentation system using standard molecular weight marker. Results: The molecular weight of the purified cattle β-lg was detected as 17.44 kDa. The isolated β-lg was almost in pure form as the molecular weight of purified β-lg monomer is 18kDa. Conclusion: The study revealed a simple and suitable method for isolation of β-lg from whey protein in pure form which may be used for detection of CMPI.

  3. Purification and Partial Characterization of Trypanosoma cruzi Triosephosphate Isomerase

    Directory of Open Access Journals (Sweden)

    Bourguignon SC

    1998-01-01

    Full Text Available The enzyme triosephosphate isomerase (TPI, EC 5.3.1.1 was purified from extracts of epimastigote forms of Trypanosoma cruzi. The purification steps included: hydrophobic interaction chromatography on phenyl-Sepharose, CM-Sepharose, and high performance liquid gel filtration chromatography. The CM-Sepharose material contained two bands (27 and 25 kDa with similar isoelectric points (pI 9.3-9.5 which could be separated by gel filtration in high performance liquid chromatography. Polyclonal antibodies raised against the porcine TPI detected one single polypeptide on western blot with a molecular weight (27 kDa identical to that purified from T. cruzi. These antibodies also recognized only one band of identical molecular weight in western blots of several other trypanosomatids (Blastocrithidia culicis, Crithidia desouzai, Phytomonas serpens, Herpertomonas samuelpessoai. The presence of only one enzymatic form of TPI in T. cruzi epimastigotes was confirmed by agarose gel activity assay and its localization was established by immunocytochemical analysis. The T. cruzi purified TPI (as well as other trypanosomatid' TPIs is a dimeric protein, composed of two identical subunits with an approximate mw of 27,000 and it is resolved on two dimensional gel electrophoresis with a pI of 9.3. Sequence analysis of the N-terminal portion of the 27 kDa protein revealed a high homology to Leishmania mexicana and T. brucei proteins

  4. Purification and characterization of xylanase from Aspergillus ficuum AF-98.

    Science.gov (United States)

    Lu, Fengxia; Lu, Mei; Lu, Zhaoxin; Bie, Xiaomei; Zhao, Haizhen; Wang, Yi

    2008-09-01

    The purification and characterization of xylanase from Aspergillus ficuum AF-98 were investigated in this work. The extracellular xylanase from this fungal was purified 32.6-fold to homogeneity throughout the precipitation with 50-80% (NH(4))(2)SO(4), DEAE-Sephadex A-50 ion exchange chromatography and Sephadex G-100 chromatography. The purified xylanase (specific activity at 288.7 U/ mg protein) was a monomeric protein with a molecular mass of 35.0 kDa as determined by SDS-PAGE. The optimal temperature and pH for the action of the enzyme were at 45 degrees C and 5.0, respectively. The xylanase was activated by Cu(2+) up to 115.8% of activity, and was strongly inhibited by Hg(2+), Pb(2+) up to 52.8% and 89%, respectively. The xylanase exhibited K(m) and V(max) values of 3.267 mg/mL, 18.38 M/min/mg for beechwood xylan and 3.747 mg/mL, 11.1M/min/mg for birchwood xylan, respectively.

  5. Purification and detoxification of petroleum refinery wastewater by electrocoagulation process.

    Science.gov (United States)

    Gousmi, N; Sahmi, A; Li, H Z; Poncin, S; Djebbar, R; Bensadok, K

    2016-09-01

    The treatment of synthetic oily wastewater having the characteristics of a typical petroleum refinery wastewater (PRW) by electrocoagulation (EC) using iron and aluminum electrodes was conducted in an electrolytic reactor equipped with fluid recirculation. During the treatment, the emulsion stability was followed by the measurement of Zeta potential and particle sizes. Effects of some operating conditions such as electrodes material, current density and electrolysis time on removal efficiencies of turbidity, and chemical oxygen demand (COD) were investigated in detail. The PRW purification by the EC process was found to be the most effective using aluminum as the anode and cathode, current density of 60 A/m(2) and 30 min of electrolysis time. Under these conditions, the process efficiencies were 83.52% and 99.94%, respectively, for COD and turbidity removals which correspond to final values of 96 mg O2/L and 0.5 NTU. A moderate energy consumption (0.341 kWh) was needed to treat 1 m(3) of PRW. Besides, the ecotoxicity test proved that toxic substances presented in the PRW, and those inhibiting the germination growth of whet, were eliminated by the EC technique.

  6. Purification and stability characterization of a cell regulatory sialoglycopeptide inhibitor

    Science.gov (United States)

    Moos, P. J.; Fattaey, H. K.; Johnson, T. C.; Spooner, B. S. (Principal Investigator)

    1995-01-01

    Previous attempts to physically separate the cell cycle inhibitory and protease activities in preparations of a purified cell regulatory sialoglycopeptide (CeReS) inhibitor were largely unsuccessful. Gradient elution of the inhibitor preparation from a DEAE HPLC column separated the cell growth inhibitor from the protease, and the two activities have been shown to be distinct and non-overlapping. The additional purification increased the specific biological activity of the CeReS preparation by approximately two-fold. The major inhibitory fraction that eluted from the DEAE column was further analyzed by tricine-SDS-PAGE and microbore reverse phase HPLC and shown to be homogeneous in nature. Two other fractions separated by DEAE HPLC, also devoid of protease activity, were shown to be inhibitory to cell proliferation and most likely represented modified relatives of the CeReS inhibitor. The highly purified CeReS was chemically characterized for amino acid and carbohydrate composition and the role of the carbohydrate in cell proliferation inhibition, stability, and protease resistance was assessed.

  7. WATER PURIFICATION BY COAGULATION UNDER THE INFLUENCE OF ULTRASONIC FIELD

    Directory of Open Access Journals (Sweden)

    Vikulina Vera Borisovna

    2016-03-01

    Full Text Available The authors carried out experiments on the in-fluence of ultrasound on the subsidence of suspended materials. The efficiency of coagulation process in wa-ter purification in ultrasound field is estimated. The influence of ultrasound on the water with suspended materials before introducing coagulant was a condition of the experiment. The magnetostriction method for obtaining ultrasound oscillations with the help of ultra-sound generator of batch production was applied. The samples were chosen and the coagulation process was controlled using standard procedures. The experimental data was obtained which estimate the efficiency in-crease in the subsidence of suspended materials de-pending on the duration of ultrasound processing. Dur-ing one minute of ultrasound processing the following results were obtained: the subsidence efficiency in-creased by 25.83 % in case of coagulant share Al2O3 2.5 mg/l; the subsidence efficiency increased by 23.70 % in case of coagulant share Al2O3 5.0 mg/l.

  8. Purification and characterization of polyphenol oxidase from purslane

    Directory of Open Access Journals (Sweden)

    Reyhan GUL GUVEN

    Full Text Available Abstract The polyphenol oxidase (PPO is an enzyme that is responsible for the enzymatic browning of fruits and vegetables. This is generally undesired process and need to be prevented in food technology. PPO from purslane was purified, characterised and the kinetic parameters for three substrates namely, catechol, L-Dopa and 4-methylcatechol were determined. The optimum pH and temperature values were found to be pH 7.0 and 50 °C, respectively using the catechol as substrate. The apparent molecular weight of the PPO from purslane was determined as high as 163 kDa by partially denaturing SDS-PAGE. Moreover, the inhibition kinetics of the purified PPO were determined, using both synthetic and natural inhibitors. Among inhibitors tested, ascorbic acid was the most effective inhibitor with the lowest Ki value of 0.36 mM. This is the first study on the purification and characterisation of PPO from purslane (Portulaca oleracea that may provide new insight into how to overcome the enzymatic browning.

  9. About the Purification Route of Ionic Liquid Precursors

    Directory of Open Access Journals (Sweden)

    Massimo De Francesco

    2017-03-01

    Full Text Available In this work a purification route of precursors for ionic liquids tailored to electrochemical energy storage systems is reported and described. The study was carried out on the N-butyl-N-methylpyrrolidinium bromide (PYR14Br precursor, which represents the intermediate product of the synthesis process of the N-butyl-N-methylpyrrolidinium bis(trifluoromethanesulfonylimide (PYR14TFSI hydrophobic ionic liquid. The target is to develop an easy and cost-effective approach for efficiently purifying several kinds of ionic liquid precursors and determining their purity content. The PYR14Br precursor was synthesized through an eco-friendly preparation procedure, which requires water as the only processing solvent, and purified through sorbent materials, such as activated charcoal and alumina. The effect of the treatment/nature/content of sorbents and processing temperature/time was investigated. The impurity content was detected by UV-VIS spectrophotometry measurements. Additionally, a correlation between the measured absorbance and the content of impurities within the precursor was obtained. The purity level of the precursor was seen to play a key role in the electrochemical performance of the ionic liquids.

  10. Deep liquid-chromatographic purification of uranium extract from technetium

    Energy Technology Data Exchange (ETDEWEB)

    Volk, V.; Dvoeglazov, K; Podrezova, L.; Vidanov, V.; Pavlyukevich, E. [OAO State Research Center - VNIINM, Rogov str., bld. 5, Moscow (Russian Federation)

    2013-07-01

    The recycling of uranium in the nuclear fuel cycle requires the removal of a number of radioactive and stable impurities like {sup 99}Tc from spent fuels. In order to improve the grade of uranium extract purification from technetium the method of liquid chromatography and the apparatus for its performance have been developed. Process of technetium extraction and concentrating in aqueous solution containing reducing agent has been studied on simulated solutions (U-Tc-HNO{sub 3}-30% TBP-isoparM). The dynamic tests of the method have been carried out on the laboratory unit. Solution of diformyl-hydrazine in nitric acid was used as a stationary phase. Silica gel with specific surface of 186 m{sup 2}/g was used as a carrier of the stationary phase. It is shown that the volume of purified extract increases as the solution temperature increases, concentration of reducing agent increases and extract flow rate decreases. It is established that the technetium content in uranium by this method could achieve a value below 0.3 ppm. Some variants of overload and composition of the stationary phase containing the extracted technetium have been offered and tested. It is defined that the method provides reduction of processing medium-active wastes by more than 10 times during finish refining process. (authors)

  11. Isolation, Purification, and Characterization of Fungal Laccase from Pleurotus sp.

    Directory of Open Access Journals (Sweden)

    Sunil S. More

    2011-01-01

    Full Text Available Laccases are blue copper oxidases (E.C. 1.10.3.2 benzenediol: oxygen oxidoreductase that catalyze the one-electron oxidation of phenolics, aromatic amines, and other electron-rich substrates with the concomitant reduction of O2 to H2O. They are currently seen as highly interesting industrial enzymes because of their broad substrate specificity. A positive strain was isolated and characterized as nonspore forming Basidiomycetes Pleurotus sp. Laccase activity was determined using ABTS as substrate. Laccase was purified by ionexchange and gel filtration chromatography. The purified laccase was a monomer showed a molecular mass of 40±1 kDa as estimated by SDS-PAGE and a 72-fold purification with a 22% yield. The optimal pH and temperature were 4.5 and 65°C, respectively. The Km and Vmax⁡ values are 250 (mM and 0.33 (μmol/min, respectively, for ABTS as substrate. Metal ions like CuSO4, BaCl2, MgCl2, FeCl2, ZnCl2 have no effect on purified laccase whereas HgCl2 and MnCl2 moderately decrease enzyme activity. SDS and sodium azide inhibited enzyme activity, whereas Urea, PCMB, DTT, and mercaptoethanol have no effect on enzyme activity. The isolated laccase can be used in development of biosensor for detecting the phenolic compounds from the effluents of paper industries.

  12. New Structured Packing CUB for Purification of Exhaust Gases

    Directory of Open Access Journals (Sweden)

    Irina Novikova

    2016-10-01

    Full Text Available New structured packing for heat and mass transfer processes named CUB is presented in our article. The packing can be applied in packed towers for exhaust gas cleaning instead random packing, for example, rings type that are the most used in such processes. The advantages of the new packing over random packing are lower pressure drop, capability of purification and as a consequence long-term service of the packing. The researches of intensity of liquid-phase mass-transfer in packed bed depending on liquid spray rate and gas velocity were carried out. Obtained data show that packing CUB is more effective than the most popular type of structured packing under all other conditions being equal. As experimental data shown heat transfer coefficient was up by 17% and mass transfer coefficient was up by 51%. We found out optimal geometry of cross section of the new packing, namely, number of elements and parameters of one element. The new construction of structured packing is applicable for both type of column cross-section round and square.

  13. Natural water purification and water management by artificial groundwater recharge.

    Science.gov (United States)

    Balke, Klaus-Dieter; Zhu, Yan

    2008-03-01

    Worldwide, several regions suffer from water scarcity and contamination. The infiltration and subsurface storage of rain and river water can reduce water stress. Artificial groundwater recharge, possibly combined with bank filtration, plant purification and/or the use of subsurface dams and artificial aquifers, is especially advantageous in areas where layers of gravel and sand exist below the earth's surface. Artificial infiltration of surface water into the uppermost aquifer has qualitative and quantitative advantages. The contamination of infiltrated river water will be reduced by natural attenuation. Clay minerals, iron hydroxide and humic matter as well as microorganisms located in the subsurface have high decontamination capacities. By this, a final water treatment, if necessary, becomes much easier and cheaper. The quantitative effect concerns the seasonally changing river discharge that influences the possibility of water extraction for drinking water purposes. Such changes can be equalised by seasonally adapted infiltration/extraction of water in/out of the aquifer according to the river discharge and the water need. This method enables a continuous water supply over the whole year. Generally, artificially recharged groundwater is better protected against pollution than surface water, and the delimitation of water protection zones makes it even more save.

  14. Purification of Piscirickettsia salmonis and partial characterisation of antigens

    Science.gov (United States)

    Barnes, M.N.; Landolt, M.L.; Powell, D.B.; Winton, J.R.

    1998-01-01

    Piscirickettsia salmonis is the etiological agent of salmonid rickettsial septicemia, an economically significant disease affecting the salmon aquaculture industry. As with other rickettsial pathogens, antigenic analysis of P. salmonis has been limited by the inherent difficulties of purifying an intracellular organism away from host cell material. In this report, we describe the use of diatrizoate meglumine and diatrizoate sodium (DMDS) density gradient centrifugation to purify P. salmonis grown in chinook salmon embryo (CHSE-214) cells. Plaque assay titers and total protein assays confirmed that viable P. salmonis was consistently concentrated in a visible band within the DMDS density gradient at a density of 1.15 to 1.16 g ml-1. Recovery of purified, viable organisms from DMDS density gradients varied from 0.6 to 3%. Preparations of uninfected CHSE-214 cells, CHSE-214 cells infected with P. salmonis, and gradient-purified P. salmonis were compared using sodium dodecyl sulfate polyacrylamide gel electrophoresis to assess the degree of purification and to identify P. salmonis-specific proteins. Although gradient-purified P. salmonis preparations were not completely free of host cell material, 8 bacterial proteins were identified. Polyclonal rabbit antiserum was used in an immunoblot of proteins from purified P. salmonis to identify 3 major and 5 minor antigens. The major antigens of 56, 30 and 20 kDa were potential candidates for experimental vaccines and development of novel diagnostic assays.

  15. The isolation and purification of a caribbean maitotoxin

    Energy Technology Data Exchange (ETDEWEB)

    Davis, S.E.; Knoepp, S.M.; Lanoue, B.A. [and others

    1994-12-31

    The phenomenon known as red tide has been a topic of great interest in that there is concern that the scale and complexity of this natural phenomenon are expanding. It is known that the benthic dinoflagellate, Gambierdiscus toxicus, produces a variety of polyether toxins that contaminate seafood and result in human illness. Maitotoxin (MTX) is one of the toxins that have been implicated in ciguatera seafood poisoning. There is a need for the development of a much broader understanding of the nature of the poisoning toxins. MTX cogeners can be difficult to isolate due to its size and chemical nature. A major goal is to obtain a purified standard of a Caribbean MTX so that more efficient assays can be developed to test seafood for the presence of toxins and thus avoid human harm. The primary goal of this project is to obtain large amounts of pure maitotoxin. The procedure described is also useful as a starting point for the purification of other toxins.

  16. Recombinant allergen Lol p II: expression, purification and characterization.

    Science.gov (United States)

    Tamborini, E; Brandazza, A; De Lalla, C; Musco, G; Siccardi, A G; Arosio, P; Sidoli, A

    1995-05-01

    Pollen from perennial rye grass (Lolium perenne) is a major cause of type I allergies worldwide. It contains complex mixtures of proteins, among which Lol p II is a major allergen. Previously, we have reported the cloning and sequencing of Lol p II and its expression in fusion with the heavy chain of human ferritin as carrier polypeptide (Sidoli et al., 1993, J. biol. Chem. 268, 21819-21825). Here, we describe the expression, purification and characterization of a recombinant Lol p II overproduced as a non-fusion protein in the periplasm of E. coli. The recombinant allergen was expressed in high yields and was easily purified in milligram amounts. It competed with the natural Lol p II for binding to specific IgE, and it induced allergic responses in skin prick tests, indicating to be immunologically analogous to the natural protein. Biochemical analyses indicate that recombinant Lol p II is a highly stable and soluble monomeric molecule which behaves like a small globular protein.

  17. Purification and proteomics of pathogen-modified vacuoles and membranes

    Directory of Open Access Journals (Sweden)

    Jo-Ana eHerweg

    2015-06-01

    Full Text Available Certain pathogenic bacteria adopt an intracellular lifestyle and proliferate in eukaryotic host cells. The intracellular niche protects the bacteria from cellular and humoral components of the mammalian immune system, and at the same time, allows the bacteria to gain access to otherwise restricted nutrient sources. Yet, intracellular protection and access to nutrients comes with a price, i.e. the bacteria need to overcome cell-autonomous defense mechanisms, such as the bactericidal endocytic pathway. While a few bacteria rupture the early phagosome and escape into the host cytoplasm, most intracellular pathogens form a distinct, degradation-resistant and replication-permissive membranous compartment. Intracellular bacteria that form unique pathogen vacuoles include Legionella, Mycobacterium, Chlamydia, Simkania and Salmonella species. In order to understand the formation of these pathogen niches on a global scale and in a comprehensive and quantitative manner, an inventory of compartment-associated host factors is required. To this end, the intact pathogen compartments need to be isolated, purified and biochemically characterized. Here, we review recent progress on the isolation and purification of pathogen-modified vacuoles and membranes, as well as their proteomic characterization by mass spectrometry and different validation approaches. These studies provide the basis for further investigations on the specific mechanisms of pathogen-driven compartment formation.

  18. Advanced Water Purification System for In Situ Resource Utilization Project

    Science.gov (United States)

    Anthony, Stephen M.

    2014-01-01

    A main goal in the field of In Situ Resource Utilization is to develop technologies that produce oxygen from regolith to provide consumables to an extratrrestrial outpost. The processes developed reduce metal oxides in the regolith to produce water, which is then electrolyzed to produce oxygen. Hydrochloric and hydrofluoric acids are byproducts of the reduction processes, which must be removed to meet electrolysis purity standards. We previously characterized Nation, a highly water selective polymeric proton-exchange membrane, as a filtrtion material to recover pure water from the contaminated solution. While the membranes successfully removed both acid contaminants, the removal efficiency of and water flow rate through the membranes were not sufficient to produce large volumes of electrolysis-grade water. In the present study, we investigated electrodialysis as a potential acid removable technique. Our studies have show a rapid and significant reduction in chloride and fluoride concentrations in the feed solution, while generating a relatively small volume of concentrated waste water. Electrodialysis has shown significant promise as the primary separation technique in ISRU water purification processes.

  19. Purification and characterization of deoxyribonuclease from earthworm Eisenia foetida.

    Science.gov (United States)

    Zhang, Jian-lin; Liu, Zhi-zhen; Wang, Xiao-yuan; Yao, Jian-qiang; Luo, Jia; Wang, Jian-hua; Yang, Li-jun; Yang, Qi; Niu, Bo

    2008-10-18

    To purify a kind of deoxyribonuclease from earthworm Eisenia foetida (named earthworm DNase, EDNase) and study its characteristics. Acetone precipitation, ion-exchange chromatography, high performance liquid chromatography, SDS-PAGE, Capillary electrophoresis isoelectric focusing and MALDI-TOP MS were used for the study. This purified protocol improved 137-fold purification and 45.6% recovery of enzyme activity. The molecular mass of EDNase was estimated to be 63,000. Mg2+, Mn2+ and Ca2+ were strong inhibitors of EDNase, while Na+ slightly increased the enzyme activity. The enzyme was completely stable in the pH range from 4.4 to 5.2 and had a pH optimum of 4.8. The optimum temperature was 37 degree C and the enzyme was stable up to 40 degree C. The pI of the enzyme was 6.20. Km and Vmax for the enzyme were 1.52 g/L and 4.89 mg/(mL.min), respectively, with calf thymus DNA as substrate. The enzyme was able to degrade chromosomal DNA, linear lambda-bacteriophage DNA as well as supercoiled plasmid DNA, but didn't display any RNase activity. This kind of deoxyribonuclease possesses unique characteristics, which is different from the deoxyribonucleases which we have known before.

  20. Gelation and fodrin purification from rat brain extracts.

    Science.gov (United States)

    Levilliers, N; Péron-Renner, M; Coffe, G; Pudles, J

    1986-06-03

    Extracts from rat brain tissue have been shown to give rise to a gel which exhibits the following features. It is mainly enriched in actin and in a high-molecular-weight protein with polypeptide chains of 235 and 240 kDa, which we identified as fodrin. Tubulin is also a major component of the gel but it appears to be trapped non-specifically during the gelation process. Gelation is pH-, ionic strength- and Ca2+-concentration-dependent, and is optimal under the conditions which promote the interaction between polymerized actin and fodrin. In a similar way to that described for the purification of rat brain actin (Levilliers, N., Péron-Renner, M., Coffe, G. and Pudles, J. (1984) Biochimie 66, 531-537), we used the gelation system as a selective means of recovering fodrin from the mixture of a low-ionic-strength extract from whole rat brain and a high-ionic-strength extract of the particulate fraction. From this gel, fodrin was purified with a good yield by a simple procedure involving gel dissociation in 0.5 M KCl and depolymerization in 0.7 M KI, Bio-Gel A-15m chromatography, followed by ammonium sulfate precipitation.

  1. Prediction of purification of biopharmeceuticals with molecular dynamics

    Science.gov (United States)

    Ustach, Vincent; Faller, Roland

    Purification of biopharmeceuticals remains the most expensive part of protein-based drug production. In ion exchange chromatography (IEX), prediction of the elution ionic strength of host cell and target proteins has the potential to reduce the parameter space for scale-up of protein production. The complex shape and charge distribution of proteins and pores complicates predictions of the interactions in these systems. All-atom molecular dynamics methods are beyond the scope of computational limits for mass transport regimes. We present a coarse-grained model for proteins for prediction of elution pH and ionic strength. By extending the raspberry model for colloid particles to surface shapes and charge distributions of proteins, we can reproduce the behavior of proteins in IEX. The average charge states of titratatable amino acid residues at relevant pH values are determined by extrapolation from all-atom molecular dynamics at pH 7. The pH specific all-atom electrostatic field is then mapped onto the coarse-grained surface beads of the raspberry particle. The hydrodynamics are reproduced with the lattice-Boltzmann scheme. This combination of methods allows very long simulation times. The model is being validated for known elution procedures by comparing the data with experiments. Defense Threat Reduction Agency (Grant Number HDTRA1-15-1-0054).

  2. Water transport and purification in nanochannels controlled by asymmetric wettability.

    Science.gov (United States)

    Chen, Qinwen; Meng, Lingyi; Li, Qikai; Wang, Dong; Guo, Wei; Shuai, Zhigang; Jiang, Lei

    2011-08-08

    Biomimetic asymmetric nanochannels have recently attracted increasing attention from researchers, especially in the aspect of the asymmetric wettability (a hydrophilic-hydrophobic system), which can be utilized to control the wetting behavior of aqueous media and to offer a means for guiding water motion. By using molecular dynamics simulations, a design for a potentially efficient water filter is presented based on (n, n) single-walled carbon nanotubes, where n = 6, 8, 10 and 12, asymmetrically modified with hydrophilic groups (carboxyl, -COOH) at one tip and hydrophobic groups (trifluoromethyl, -CF(3) ) at the other. The reduced water density on the hydrophobic sides of the functionalized nanotubes are observed in both pure water and aqueous electrolyte solution, except for the functionalized (6, 6) tube, due to the change of dipole orientation of the single-file water wire within it. The functionalized (8, 8) tube can significantly maintain the low water density on the hydrophobic side. Both (6, 6) and (8, 8) tubes have relatively high energy barriers at their tips for ion permeation, which can be obtained by calculating the potential of mean force. Such tip functionalization of a nanotube therefore suggests the great possibilities of water transport and filtration, dominated by asymmetric wettability. The functionalized (8, 8) tube could act as a nanofluidic channel for water purification, not only for ion exclusion but also as a stable water column structure. Copyright © 2011 WILEY-VCH Verlag GmbH & Co. KGaA, Weinheim.

  3. Purification and properties of soluble actin from sea urchin eggs.

    Science.gov (United States)

    Mabuchi, I; Spudich, J A

    1980-03-01

    Unfertilized eggs of the sea urchin, Strongylocentrotus purpuratus, were homogenized in a buffer containing 0.1 M KCl and 2 mM MgCl2 at pH 6.85. About 50% of the actin was recovered in the high-speed supernate of the homogenate. More than 80% of the actin in this supernate was found to be monomeric upon gel filtration chromatography through a Sephadex G-150 column or by a DNase I inhibition assay. The critical concentration for polymerization of this actin prior to further purification was 0.3-0.9 mg/ml under various conditions. Actin was purified to near homogeneity from the Sephadex G-150 pool with high yield. The purified actin had a critical concentration for polymerization of 0.02-0.03 mg/ml. The isoelectric point of the crude actin and the purified actin was the same. Indeed, we found that there is only one isoelectric focusing species of actin in the sea urchin egg, and it has an isoelectric point more basic than rabbit skeletal muscle actin. The discrepancy between the polymerizability of the crude and purified actin may be due to the presence of factors in the crude fraction which inhibit the polymerization of actin.

  4. Purification of nanogram-range immunoprecipitated DNA in ChIP-seq application.

    Science.gov (United States)

    Zhong, Jian; Ye, Zhenqing; Lenz, Samuel W; Clark, Chad R; Bharucha, Adil; Farrugia, Gianrico; Robertson, Keith D; Zhang, Zhiguo; Ordog, Tamas; Lee, Jeong-Heon

    2017-12-21

    Chromatin immunoprecipitation-sequencing (ChIP-seq) is a widely used epigenetic approach for investigating genome-wide protein-DNA interactions in cells and tissues. The approach has been relatively well established but several key steps still require further improvement. As a part of the procedure, immnoprecipitated DNA must undergo purification and library preparation for subsequent high-throughput sequencing. Current ChIP protocols typically yield nanogram quantities of immunoprecipitated DNA mainly depending on the target of interest and starting chromatin input amount. However, little information exists on the performance of reagents used for the purification of such minute amounts of immunoprecipitated DNA in ChIP elution buffer and their effects on ChIP-seq data. Here, we compared DNA recovery, library preparation efficiency, and ChIP-seq results obtained with several commercial DNA purification reagents applied to 1 ng ChIP DNA and also investigated the impact of conditions under which ChIP DNA is stored. We compared DNA recovery of ten commercial DNA purification reagents and phenol/chloroform extraction from 1 to 50 ng of immunopreciptated DNA in ChIP elution buffer. The recovery yield was significantly different with 1 ng of DNA while similar in higher DNA amounts. We also observed that the low nanogram range of purified DNA is prone to loss during storage depending on the type of polypropylene tube used. The immunoprecipitated DNA equivalent to 1 ng of purified DNA was subject to DNA purification and library preparation to evaluate the performance of four better performing purification reagents in ChIP-seq applications. Quantification of library DNAs indicated the selected purification kits have a negligible impact on the efficiency of library preparation. The resulting ChIP-seq data were comparable with the dataset generated by ENCODE consortium and were highly correlated between the data from different purification reagents. This study provides

  5. Isolation of Mannheimia (Pasteurella) haemolytica serotypes A2 and ...

    African Journals Online (AJOL)

    v23i2.4572 · AJOL African Journals Online. HOW TO USE AJOL... for Researchers · for Librarians · for Authors. OTHER RESOURCES... Journal Quality · for Researchers · for Journals · for Authors · for Policy Makers · about Open Access · FAQ's ...

  6. Prevalence and Biotyping of Pasteurella Haemolytica Isolates from ...

    African Journals Online (AJOL)

    In all, biotype A was 60.75%, biotype T 22.43% and untypable isolates were 16.82%. Overall, biotype A occurred more significantly than biotype T (P<0.05) and appeared to be the predominant biotype in the area. Gross pathological appearance of pneumonic lungs was characterized by pulmonary oedema, congestion and ...

  7. Shear Dependent LC Purification of an Engineered DNA Nanoswitch and Implications for DNA Origami.

    Science.gov (United States)

    Halvorsen, Ken; Kizer, Megan E; Wang, Xing; Chandrasekaran, Arun Richard; Basanta-Sanchez, Maria

    2017-06-06

    As DNA nanotechnology matures, there is increasing need for fast, reliable, and automated purification methods. Here, we develop UHPLC methods to purify self-assembled DNA nanoswitches, which are formed using DNA origami approaches and are designed to change conformations in response to a binding partner. We found that shear degradation hindered LC purification of the DNA nanoswitches, removing oligonucleotides from the scaffold strand and causing loss of function. However, proper choice of column, flow rate, and buffers enabled robust and automated purification of DNA nanoswitches without loss of function in under a half hour. Applying our approach to DNA origami structures, we found that ∼400 nm long nanotubes degraded under the gentlest flow conditions while ∼40 nm diameter nanospheres remained intact even under aggressive conditions. These examples show how fluid stresses can affect different DNA nanostructures during LC purification and suggest that shear forces may be relevant for some applications of DNA nanotechnology. Further development of this approach could lead to fast and automated purification of DNA nanostructures of various shapes and sizes, which would be an important advance for the field.

  8. MAP Tag: A Novel Tagging System for Protein Purification and Detection.

    Science.gov (United States)

    Fujii, Yuki; Kaneko, Mika K; Kato, Yukinari

    2016-12-01

    Protein purification is an essential procedure in fields such as biochemistry, molecular biology, and biophysics. Acquiring target proteins with high quality and purity is still difficult, although several tag systems have been established for protein purification. Affinity tag systems are excellent because they possess high affinity and specificity for acquiring the target proteins. Nevertheless, further affinity tag systems are needed to compensate for several disadvantages of the presently available affinity tag systems. Herein, we developed a novel affinity tag system designated as the MAP tag system. This system is composed of a rat anti-mouse podoplanin monoclonal antibody (clone PMab-1) and MAP tag (GDGMVPPGIEDK) derived from the platelet aggregation-stimulating domain of mouse podoplanin. PMab-1 possesses high affinity and specificity for the MAP tag, and the PMab-1/MAP tag complex dissociates in the presence of the epitope peptide, indicating that the MAP tag system is suitable for protein purification. We successfully purified several proteins, including a nuclear protein, soluble proteins, and a membrane protein using the MAP tag system. The MAP tag system is very useful not only for protein purification but also in protein detection systems such as western blot and flow cytometric analyses. Taken together, these findings indicate that the MAP tag system could be a powerful tool for protein purification and detection.

  9. Iodixanol Density Gradient Preparation in University of Wisconsin Solution for Porcine Islet Purification

    Directory of Open Access Journals (Sweden)

    Michael P.M. Van der Burg

    2003-01-01

    Full Text Available Previously published as Graham, J.M. (2002 Purification of Islets of Langerhans from porcine pancreas. TheScientificWorldJOURNAL 2, 1657–1661. ISSN 1537-744X; DOI 10.1100/tsw.2002.847.Generally, prior to the purification of isolated pancreatic islets, the collagenase-digested tissue is incubated in the University of Wisconsin solution (UWS; ~320 mOsm for osmotic stabilization to preserve or improve the density differences between islets and acinar fragments. The adverse effects arising from the subsequent pelleting and resuspension of the islets in a second, different (often highly hyperosmotic purification solution are avoided in the protocol described here; preparation of the purification medium is simply achieved by mixing the UWS preincubated islets with a second UWS containing the inert impermeant iodixanol. Flotation of the islets isolated from juvenile porcine pancreases through this mildly hypertonic (~380 mOsm gradient of iodixanol-UWS achieves a much higher recovery of islets of an improved viability than the customary method using a Ficoll gradient. The method has been extended to human islet purification.

  10. Development and Operation of a Liquid Scintillator Purification System for a Solar Neutrino Detector

    Science.gov (United States)

    Chen, M.; Benziger, J. B.; Calaprice, F. P.; Darnton, N.; Johnson, M.; Loeser, F.; Vogelaar, R. B.

    1996-10-01

    An on-line purification system for a large-scale, liquid scintillator detector has been developed for the Counting Test Facility (CTF), a five-ton prototype of the Borexino solar neutrino detector at Gran Sasso. This purification system was operated to remove radioactive impurities from the pseudocumene-based scintillator in the CTF. Counter-current water extraction was performed to remove ionic impurities from the scintillator. Notably, the radon daughters ^210Bi and ^210Po were identified prior to purification and were successfully removed by water extraction. Vacuum distillation of the entire scintillator mixture allowed high radiopurity and chemical purity to be maintained; in addtion, it enabled a test of the origin of ^14C in the scintillator mixture to be performed. Finally, nitrogen stripping was utilized to remove noble gas radioactive isotopes, such as ^85Kr and ^222Rn. The results of the CTF purification activities and an overview of the purification scheme for the Borexino solar neutrino experiment will be presented.

  11. Automated genomic DNA purification options in agricultural applications using MagneSil paramagnetic particles

    Science.gov (United States)

    Bitner, Rex M.; Koller, Susan C.

    2002-06-01

    The automated high throughput purification of genomic DNA form plant materials can be performed using MagneSil paramagnetic particles on the Beckman-Coulter FX, BioMek 2000, and the Tecan Genesis robot. Similar automated methods are available for DNA purifications from animal blood. These methods eliminate organic extractions, lengthy incubations and cumbersome filter plates. The DNA is suitable for applications such as PCR and RAPD analysis. Methods are described for processing traditionally difficult samples such as those containing large amounts of polyphenolics or oils, while still maintaining a high level of DNA purity. The robotic protocols have ben optimized for agricultural applications such as marker assisted breeding, seed-quality testing, and SNP discovery and scoring. In addition to high yield purification of DNA from plant samples or animal blood, the use of Promega's DNA-IQ purification system is also described. This method allows for the purification of a narrow range of DNA regardless of the amount of additional DNA that is present in the initial sample. This simultaneous Isolation and Quantification of DNA allows the DNA to be used directly in applications such as PCR, SNP analysis, and RAPD, without the need for separate quantitation of the DNA.

  12. PURIFICATION AND CHARACTERIZATION OF POLY-HYDROXYBUTYRATE (PHB IN CUPRIAVIDUS NECATOR

    Directory of Open Access Journals (Sweden)

    Sergio Leon De Rooy

    2010-06-01

    Full Text Available Purification and characterization of biodegradable plastic namely Polyhydroxybutyrate (PHB in Cupriavidus necator have been carried out. C. necator was grown on a Ramsay medium with fixed substrate conditions and optimized for time. Stepwise purification of PHB was carried out, by using hydrogen peroxide and chloroform. The effect of temperature, time, and hydrogen peroxide concentration on the purification were also evaluated. The extracted PHB was studied with XRD, FTIR and 1H-NMR and 13C-NMR to determine its structure and purity. Yield and crystallinity were also studied with HPLC and XRD, respectively. The results of the research showed that higher concentrations of hydrogen peroxide gave better yields, whereas higher temperatures and longer lysis times led to different results. Higher crystallinity was observed when purification temperatures were elevated, but higher hydrogen peroxide concentration and longer extraction time gave varying crystallinity. The highest yield ca 66.10 % DCW was reached by purification using H2O2 20 %, at 100 oC for 2 h. The results of   TGA analysis indicated that the purity of the PHB obtained was about 75 % and by using DSC, it was found that the PHB showed good thermal properties.   Keywords:  PHB, recovery, hydrogen peroxide, characterization

  13. Purification of SUMO-1 modified IκBα and complex formation with NF-κB.

    Science.gov (United States)

    Lens, Zoé; Dewitte, Frédérique; Van Lint, Carine; de Launoit, Yvan; Villeret, Vincent; Verger, Alexis

    2011-12-01

    Covalent modification of proteins with SUMO (Small Ubiquitin-like MOdifier) affects many cellular processes, including transcriptional regulation, DNA repair and signal transduction. Although hundreds of SUMO targets have been identified, many biological outcomes of protein sumoylation remain poorly understood. In particular, biochemical and structural analysis can only be easily conducted if highly pure sumoylated substrates are available. Purification of sumoylated substrates in vitro or in bacteria have been previously reported but separating the sumoylated protein from the undesired unmodified fraction is often technically challenging, inefficient and time consuming. Here we develop a new vector system for in vivo sumoylation in Escherichia coli which improves purification of sumoylated proteins. We describe the purification of IκBα, its sumoylation, the subsequent separation and purification of the modified and the unmodified forms and the purification of the complex IκBα-SUMO-1/NF-κB. After a first GST affinity chromatography and GST-tag removal, a unique metal-ion affinity chromatography using a 6xHis-SUMO-1 tag results in mgs of highly pure SUMO-1 modified IκBα. Our pure SUMO-1 modified IκB/NF-κB complex could be a useful tool to identify new interaction partner specific of the SUMO-1 modified IκBα form. This approach may be extended to other SUMO substrates not isolable by classical chromatography techniques. Copyright © 2011 Elsevier Inc. All rights reserved.

  14. Rescue purification maximizes the use of human islet preparations for transplantation.

    Science.gov (United States)

    Ichii, Hirohito; Pileggi, Antonello; Molano, R Damaris; Baidal, David A; Khan, Aisha; Kuroda, Yoshikazu; Inverardi, Luca; Goss, John A; Alejandro, Rodolfo; Ricordi, Camillo

    2005-01-01

    The relative inefficiency of the islet purification process may hamper obtaining enough islets for transplantation even with adequate pre-purification counts. In this study, we determined the effect of an additional purification step on total islet yields and pancreas utilization at our center. Twenty-five pancreata were processed using the automated method followed by continuous gradient purification (CGP), and the less pure islet fractions were subjected to additional rescue gradient purification (RGP). CGP and RGP islets were combined and transplanted into patients with type 1 diabetes. CGP and RGP islets showed no significant differences in cell viability, insulin secretion in vitro and function when transplanted into chemically diabetic mice. Mean RGP contribution to the final preparation was 27.9 +/- 19.9%. In 12 of 25 preparations, CGP yielded <5000 IEQ/kg of recipient body weight, and inclusion of RGP islets to the final preparation allowed to obtain the minimal islet number required for transplantation. Transplanted islets resulted in sustained C-peptide production, HbA1(C) normalization and insulin-independence or reduced insulin requirements. Taken together, our data suggest that RGP islets are comparable in terms of viability and potency to CGP islets. RGP may be of assistance in maximizing the number of islet preparations successfully used in transplant protocols.

  15. Chemoenzymatic reversible immobilization and labeling of proteins without prior purification.

    Science.gov (United States)

    Rashidian, Mohammad; Song, James M; Pricer, Rachel E; Distefano, Mark D

    2012-05-23

    Site-specific chemical modification of proteins is important for many applications in biology and biotechnology. Recently, our laboratory and others have exploited the high specificity of the enzyme protein farnesyltransferase (PFTase) to site-specifically modify proteins through the use of alternative substrates that incorporate bioorthogonal functionality including azides and alkynes. In this study, we evaluate two aldehyde-containing molecules as substrates for PFTase and as reactants in both oxime and hydrazone formation. Using green fluorescent protein (GFP) as a model system, we demonstrate that the purified protein can be enzymatically modified with either analogue to yield aldehyde-functionalized proteins. Oxime or hydrazone formation was then employed to immobilize, fluorescently label, or PEGylate the resulting aldehyde-containing proteins. Immobilization via hydrazone formation was also shown to be reversible via transoximization with a fluorescent alkoxyamine. After characterizing this labeling strategy using pure protein, the specificity of the enzymatic process was used to selectively label GFP present in crude E. coli extract followed by capture of the aldehyde-modified protein using hydrazide-agarose. Subsequent incubation of the immobilized protein using a fluorescently labeled or PEGylated alkoxyamine resulted in the release of pure GFP containing the desired site-specific covalent modifications. This procedure was also employed to produce PEGylated glucose-dependent insulinotropic polypeptide (GIP), a protein with potential therapeutic activity for diabetes. Given the specificity of the PFTase-catalyzed reaction coupled with the ability to introduce a CAAX-box recognition sequence onto almost any protein, this method shows great potential as a general approach for selective immobilization and labeling of recombinant proteins present in crude cellular extract without prior purification. Beyond generating site-specifically modified proteins, this

  16. Boronic acid-modified magnetic materials for antibody purification.

    Science.gov (United States)

    Dhadge, Vijaykumar L; Hussain, Abid; Azevedo, Ana M; Aires-Barros, Raquel; Roque, Ana C A

    2014-02-06

    Aminophenyl boronic acids can form reversible covalent ester interactions with cis-diol-containing molecules, serving as a selective tool for binding glycoproteins as antibody molecules that possess oligosaccharides in both the Fv and Fc regions. In this study, amino phenyl boronic acid (APBA) magnetic particles (MPs) were applied for the magnetic separation of antibody molecules. Iron oxide MPs were firstly coated with dextran to avoid non-specific binding and then with 3-glycidyloxypropyl trimethoxysilane to allow further covalent coupling of APBA (APBA_MP). When contacted with pure protein solutions of human IgG (hIgG) and bovine serum albumin (BSA), APBA_MP bound 170 ± 10 mg hIgG g(-1) MP and eluted 160 ± 5 mg hIgG g(-1) MP, while binding only 15 ± 5 mg BSA g(-1) MP. The affinity constant for the interaction between hIgG and APBA_MP was estimated as 4.9 × 10(5) M(-1) (Ka) with a theoretical maximum capacity of 492 mg hIgG adsorbed g(-1) MP (Qmax), whereas control particles bound a negligible amount of hIgG and presented an estimated theoretical maximum capacity of 3.1 mg hIgG adsorbed g(-1) MP (Qmax). APBA_MPs were also tested for antibody purification directly from CHO cell supernatants. The particles were able to bind 98% of IgG loaded and to recover 95% of pure IgG (purity greater than 98%) at extremely mild conditions.

  17. Purification and some properties of rose (Fructus cynosbati) hips invertase.

    Science.gov (United States)

    Sacan, Ozlem; Yanardag, Refiye

    2012-04-01

    Invertase was purified from rose (Fructus cynosbati) hips by ammonium sulfate fractionation and hydroxyapatite column chromatography. The enzyme was obtained with a yield of 4.25% and about 10.48-fold purification and had a specific activity of 8.59 U/mg protein. The molecular mass of invertase was estimated to be 66.51 kDa by PAGE and 34 kDa by SDS-PAGE, indicating that the native enzyme was a homodimer. The enzyme was a glycoprotein and contained 5.86% carbohydrate. The K(m) for sucrose was 14.55 mM and the optimum pH and temperature of the enzyme were 4.5 and 40 degrees C, respectively. Sucrose was the most preferred substrate of the enzyme. The enzyme also hydrolyzed D(+) raffinose, D(+) trehalose and inulin (activity 39.88, 8.12 and 4.94%, respectively of that of sucrose), while D(+) lactose, cellobiose and D(+) maltose showed no effect on the enzyme. The substrate specificity was consistent with that for a beta-fructofuranoside, which is the most popular type in the higher plants. The enzyme was completely inhibited by HgCl2, MnCl2, MnSO4, FeCl3, Pb(NO3)2, ammonium heptamolybdate, iodoacetamide and pyridoxine hydrochloride. It was also inhibited by Ba(NO3)2 (86.32%), NH4Cl (84.91%), MgCl2 (74.45%), urea (71.63%), I2 (69.64%), LiCl (64.99%), BaCl2 (50.30%), Mg(NO3)2 (49.90%), CrCl3 (31.90%) and CuSO4 (21.45%) and but was activated by Tris (73.99%) and methionine (12.47%).

  18. Gas purification by use of hot metal getter beds

    Energy Technology Data Exchange (ETDEWEB)

    Albrecht, H.

    1992-11-01

    An experimental program is described which was performed in the frame of a tritium technology task for the NET/ITER fusion fuel cycle. The aim was to investigate commercial gas purifiers containing metallic getters for the purification of gas streams such as the plasma exhaust gas. Five purifiers with up to 3000g of getter material were tested in the PEGASUS facility mainly with respect to the removal of methane, which is known to be much more difficult to remove than other impurities like O{sub 2}, N{sub 2}, or CO. A proposal for a fuel cleanup method based on a combination of getter beds and Pd/Ag diffusors is presented as the main conclusion of the test program. The discussion of this method includes the aspects of flow rates, tritium inventory, and consumption of getter material. (orig.) [Deutsch] Im Rahmen einer Tritium Technology Task fuer den NET/ITER Brennstoff-Kreislauf wurde ein experimentelles Vorhaben durchgefuehrt mit dem Ziel, kommerzielle Gasreiniger, die nach dem Prinzip der Rueckhaltung von Verunreinigungen an heissen Metall-Gettern arbeiten, auf ihre Eignung zur Reinigung von inerten Gasstroemen, wie z.B. dem Plasma Exhaust Gas, zu untersuchen. An der zu diesem Zweck gebauten PEGASUS-Anlage wurden fuenf Gasreiniger mit bis zu 3 kg Gettermaterial eingesetzt, um vor allem die Rueckhaltung von Methan zu bestimmen, das sich wesentlich schwerer abtrennen laesst als etwa O{sub 2}, N{sub 2} oder CO. Als Schlussfolgerung aus dem Versuchsprogramm wird ein Brennstoff-Reinigungsverfahren vorgeschlagen, das aus einer Kombination von Getterbetten und Pd/Ag-Permeatoren besteht. In der Diskussion dieses Verfahrens werden u.a. die Aspekte des Gasdurchsatzes, des Tritium Inventares und des Verbrauchs an Gettermaterial angesprochen. (orig.)

  19. Partial purification and properties of thiamine pyrophosphokinase from pig brain.

    Science.gov (United States)

    Peterson, J W; Gubler, C J; Kuby, S A

    1975-08-26

    Pig brain thiamine pyrophosphokinase (ATP: thiamine pyrophosphotransferase, EC 2.7.6.2) was purified 260-fold over extracts of brain acetone powder. A direct, radiometric assay was used to follow the purification. By isoelectric focusing, the purified enzyme appeared to have an isoionic point of approx. pH 4.2, but these preparations were still not homogeneous by disc-gel electrophoresis nor by analytical ultracentrifugation. The purified enzyme has a broad pH optimum extending from pH 8.3 to 9.3 in 0.028 M phosphate/glycylglycine buffers. For optimal enzymatic activity, the ratio of magnesium to ATP must be fixed at 0.6, which suggests that for this ATP-pyrophosphoryl transfer reaction, the enzymatically preferred reactant may be Mg(ATP)6-/2. A preliminary study of the kinetics of the reaction reveals that the enzyme may function via a partial "ping-pong" mechanism; on this basis, dissociation constants for ATPt and for thiamine were evaluated. Pyrithiamine, butylthiamine, ethylthiamine, and oxythiamine appeared to be competitive inhibitors with respect to thiamine as the variable substrate, and their inhibitor dissociation constants were calculated. The relatively poor affinity of oxythiamine to the enzyme emphasizes the 4-amino group in the pyrimidine ring as one of the specificity requirements for thiamine pyrophosphokinase. Preliminary values for the apparent equilibrium coefficient of the thiamine pyrophosphokinase-catalyzed reaction, in terms of total species, has been approximated at several initial concentrations of reactants: e.g. K'eq,app = (see article) 9.66 - 10(-3) M; and [Th]initial - 1 - 10(-6) and 2 - 10(-6) M, respectively, where TDP, Th, t and eq represent thiamine diphosphate, thiamine, total concentration and equilibrium concentration, respectively.

  20. Biotechnology applications of amino acids in protein purification and formulations.

    Science.gov (United States)

    Arakawa, T; Tsumoto, K; Kita, Y; Chang, B; Ejima, D

    2007-11-01

    Amino acids are widely used in biotechnology applications. Since amino acids are natural compounds, they can be safely used in pharmaceutical applications, e.g., as a solvent additive for protein purification and as an excipient for protein formulations. At high concentrations, certain amino acids are found to raise intra-cellular osmotic pressure and adjust to the high salt concentrations of the surrounding medium. They are called "compatible solutes", since they do not affect macromolecular function. Not only are they needed to increase the osmotic pressure, they are known to increase the stability of the proteins. Sucrose, glycerol and certain amino acids were used to enhance the stability of unstable proteins after isolation from natural environments. The mechanism of the action of these protein-stabilizing amino acids is relatively well understood. On the contrary, arginine was accidentally discovered as a useful reagent for assisting in the refolding of recombinant proteins. This effect of arginine was ascribed to its ability to suppress aggregation of the proteins during refolding, thereby increasing refolding efficiency. By the same mechanism, arginine now finds much wider applications than previously anticipated in the research and development of proteins, in particular in pharmaceutical applications. For example, arginine solubilizes proteins from loose inclusion bodies, resulting in efficient production of active proteins. Arginine suppresses protein-protein interactions in solution and also non-specific adsorption to gel permeation chromatography columns. Arginine facilitates elution of bound proteins from various column resins, including Protein-A or dye affinity columns and hydrophobic interaction columns. This review covers various biotechnology applications of amino acids, in particular arginine.