WorldWideScience
 
 
1

Serum alkaline phosphatase screening for vitamin D deficiency states  

International Nuclear Information System (INIS)

Objective: To determine whether serum vitamin D levels are correlated with serum levels of alkaline phosphatase or not. Study Design: Cross-sectional, observational study. Place and Duration of Study: Multi-centre study, conducted at Liaquat National Hospital and Medical College, National Medical Centre and Medicare Hospital, Karachi, from January to October 2009. Methodology: Patients attending the Orthopaedic OPDs with complaints of pain in different body regions and serum vitamin D/sub 3/ levels of greater or equal to 30 ng/ml were included in the study. Patients with vitamin D deficiency were further categorized into mild deficiency or insufficiency (vit. D/sub 3/ = 20-29 ng/ml), moderate deficiency (vit. D/sub 3/ = 5 - 19 ng/ml) and severe deficiency forms (vit. D/sub 3/ < 5 ng/ml). Pearson correlation was applied to test the correlation of serum alkaline phosphatase levels with serum vitamin D/sub 3/ levels. P-value < 0.05 was considered to be significant. Results: Out of 110 samples, 26 had mild (23%), 61 had moderate (55%) and 21 had severe (19.1%) vitamin D deficiencies. All of the patients in the three groups had alkaline phosphatase with in normal limits and the total mean value of the enzyme was 135.97 +- 68.14I U/L. The inter group comparison showed highest values of alkaline phosphatase in the moderate vitamin D deficiency group. The correlation coefficient of alkaline phosphatase and serum vitamin D/sub 3/ levels was r =0.05 (p =0.593). Conclusion: Serus was r =0.05 (p =0.593). Conclusion: Serum vitamin D/sub 3/ levels may not be correlated with increased serum alkaline phosphatase levels. Therefore, alkaline phosphatase may not be used as a screening test to rule out vitamin D deficiency. (author)

2

Serum Alkaline Phosphatase Predicts Mortality among Maintenance Hemodialysis Patients  

Digital Repository Infrastructure Vision for European Research (DRIVER)

Several observational studies have demonstrated that serum levels of minerals and parathyroid hormone (PTH) have U- or J-shaped associations with mortality in maintenance hemodialysis patients, but the relationship between serum alkaline phosphatase (AlkPhos) and risk for all-cause or cardiovascular death is unknown. In this study, a 3-yr cohort of 73,960 hemodialysis patients in DaVita outpatient dialysis were studied, and the hazard ratios for all-cause and cardiovascular death were higher ...

Regidor, Deborah L.; Kovesdy, Csaba P.; Mehrotra, Rajnish; Rambod, Mehdi; Jing, Jennie; Mcallister, Charles J.; Wyck, David; Kopple, Joel D.; Kalantar-zadeh, Kamyar

2008-01-01

3

Probable levetiracetam-related serum alkaline phosphatase elevation  

Directory of Open Access Journals (Sweden)

Full Text Available Abstract Background Levetiracetam (LEV is an antiepileptic drug with a favorable tolerability and safety profile with little or no effect on liver function. Case presentation Here, we reported an epileptic pediatric patient who developed a significant elevation in serum alkaline phosphatase level (ALP during LEV monotherapy. Moreover, the serum ALP level was surprisingly decreased to normal after LEV discontinuation. The Naranjo Adverse Drug Reaction Probability Scale score was 6, indicating firstly LEV was a probable cause for the increased serum ALP. Conclusions Cautious usage and concerns of the LEV-associated potential ALP elevation should be considered when levetiracetam is prescribed to epilepsy patients, especially pediatric patients.

Xiong Nian

2012-09-01

4

Serum alkaline phosphatases as indicators of radiation damage in rats  

International Nuclear Information System (INIS)

The effects of whole-body, single-dose irradiations (100, 300, 500, 800 R 60Co ? rays) on the activities of isoenzymes of alkaline phosphatase in the serum of rats have been studied. The degree of changes in the intestinal and liver isoenzyme activity was directly dependent on the radiation dose. A marked, time-dependent decrease in the bone isoenzyme activity in serum was found only after 800 R. A computer processing of biochemical data by stepwise discriminant analysis enabled an early diagnosis of postradiation injury of the gut and bone and to classify the irradiated rats according to the radiation dose

5

Serum Alkaline Phosphatase Levels and Mortality of Chronic Hemodialysis Patients  

Directory of Open Access Journals (Sweden)

Full Text Available Objective: Alkaline phosphatase (ALP is considered a biomarker of high bone turnover in hemodialysis (HD patients with secondary hyperparathyroidism. This study was conducted to determine whether high serum ALP levels are associated with increased all-cause mortality of HD patients. Patients and Methods: This was a retrospective cohort study conducted at a single center. The subjects were 195 patients on chronic HD therapy who were followed up for a 5 years, and relationships between their baseline data and outcomes were assessed statistically. The serum ALP level was used as the predictor, and the primary end point was all-cause mortality. Results: Based on the median serum ALP of 236 IU/L, the subjects were divided into a low-ALP group (<236 IU/l and a high-ALP group (?236 IU/l. The high-ALP group was older and had a longer dialysis vintage, lower serum phosphorus concentrations, and higher serum parathyroid hormone levels, and they also had lower serum albumin levels and higher C-reactive protein values. In a multivariate Cox model in which the baseline serum ALP levels were used adjusted for age, gender, HD vintage, comorbidity, bone metabolism parameters, and serum liver enzyme levels, each doubling of the serum ALP level was associated with a significant increase in the hazard of all-cause mortality (hazard ratio 10.70, 95% CI 1.53 - 74.24. Conclusion: High baseline serum ALP levels are associated with increased mortality of HD patients, independent of bone metabolism parameters and serum liver enzyme levels. ALP is a potential target for the treatment of HD patients.

Tetsuri Yamashita

2011-09-01

6

Quantitative method for determining serum alkaline phosphatase isoenzyme activity I. Guanidine hydrochloride: new reagent for selectively inhibiting major serum isoenzymes of alkaline phosphatase.  

Digital Repository Infrastructure Vision for European Research (DRIVER)

The potential use of the protein denaturant guanidine hydrochloride to inhibit selectively the enzyme activity of serum alkaline phosphatase isoenzymes from liver, bone, intestine, and placenta was investigated. Inhibition of each isoenzyme was shown to be dependent on time and concentration of inhibitor. In the presence of 0.3 mol/l (28.7 g/l) guanidine hydrochloride for 170 seconds 14%, 47%, and 90% of the total alkaline phosphatase activity remained in samples of bone, liver, and intestina...

Shephard, M. D.; Peake, M. J.

1986-01-01

7

Changes in lactate dehydrogenase and alkaline phosphatase in serum of mice after x-irradiation  

International Nuclear Information System (INIS)

Changes in the activities of LDH and alkaline phosphatase in the serum of mice were investigated in detail from the 2nd day to the 30th day after whole-body X-irradiation of 400 R, a dose which produces 13%, 30-day-mortality. Serum LDH levels were significantly decreased during the first 6 days after irradiation, but subsequently returned to a normal range by the 14th day. Serum alkaline phosphatase levels were decreased to a minimum on the 12th day. They returned gradually to a level slightly below control level by the 22nd day. Serum LDH and alkaline phosphatase levels seem to be good indicators of radiation injury in mice during the 2-3 weeks after irradiation, even if they have been exposed to a sublethal dose. (auth.)

8

Effects of Transport Stress on Serum Alkaline Phosphatase Activity in Beagle Dogs  

Digital Repository Infrastructure Vision for European Research (DRIVER)

Here, to determine the effects of transport stress on blood parameters in dogs, we investigated the changes in hematologic and serum chemical parameters in healthy beagle dogs transported from Beijing, China, to Osaka, Japan, to obtain the background data. Only the activity of serum alkaline phosphatase increased clearly upon arrival, a change attributed to transport stress, but the activity gradually reduced afterward. No marked changes in levels of other blood parameters were...

Ochi, Takehiro; Nishiura, Ippei; Tatsumi, Mitsuyoshi; Hirano, Yoshimi; Yahagi, Kouichi; Sakurai, Yasuhiro; Sudo, Yuji; Koyama, Hironari; Hagita, Yuichi; Fujimoto, Yoshikatsu; Kitamura, Shinji; Hashimoto, Hideki; Nakamura, Tomoya; Yamada, Asobi; Tanimoto, Masayoshi

2013-01-01

9

Canine corticosteroid-induced alkaline phosphatase in serum was solubilized by phospholipase activity in vivo.  

Science.gov (United States)

In this study, gas chromatography/mass spectrometry revealed the presence of stoichiometric amounts of myo-inositol in association with serum corticosteroid-induced isozyme of alkaline phosphatase (CALP) in canine serum. Such remnants are consistent with prior membrane attachment of serum CALP and its release into serum by endogenous phospholipase activity. Serum CALP was further shown to behave similarly to CALP released from hepatocyte membranes by glycosyl phosphatidylinositol phospholipase D (GPI-PLD) and differently from CALP solubilized by GPI-phospholipase C (PLC) on both native polyacrylamide gel electrophoresis and Western blot analysis using anti-cross-reacting determinant antibody. In addition to bile canalicular surfaces, CALP activity was found over hepatocyte sinusoidal surfaces by histochemical staining of canine liver sections. A significantly higher ratio of CALP to total alkaline phosphatase activity was observed in serum as opposed to bile in 10 of 11 paired serum and bile samples from dogs. This suggested that bile is not likely to be the source of serum CALP and is consistent with the release of CALP from hepatocyte basolateral surfaces directly into serum. It was concluded that serum CALP was once membrane bound and was released by phospholipase activity into serum. Our findings are consistent with release of CALP from the sinusoidal surfaces of hepatocytes into serum either by endogenous GPI-PLD activity or release by GPI-PLC followed by modification of the phosphatidylinositol remnant in vivo. PMID:7653569

Solter, P F; Hoffmann, W E

1995-08-01

10

High-performance liquid chromatographic assay for acid and alkaline phosphatase in serum.  

Science.gov (United States)

High-performance liquid chromatography was used to assay serum acid and alkaline phosphatase. Samples were incubated with adenosine-5'-monophosphoric acid (AMP) in a buffer of required pH, 5'-nucleotidase was inhibited with Ni2+ ions, and the phosphatase activity was determined by measuring the concentration of the reaction product, adenosine. The analysis time, after the incubation is terminated, is short (7 min), and the assay is quantitative and reproducible. Complete separation of the reaction product from the substrate and the naturally occurring serum constituents and the high sensitivity of the ultraviolet detection system eliminate some of the problems commonly encountered in spectrophotometric assays. PMID:231044

Krstulovic, A M; Hartwick, R A; Brown, P R

1979-05-01

11

Serum Alkaline Phosphatase Levels in Healthy Children and Evaluation of Alkaline Phosphatasez-scores in Different Types of Rickets  

Digital Repository Infrastructure Vision for European Research (DRIVER)

Objective: Serum alkaline phosphatase (ALP) levels show great variation with age and sex in children and adolescents. Additionally, different buffers used even in the same method cause variable results. This detail is not usually taken into account in the evaluation. We aimed to study pediatric age- and sex-specific reference ranges for ALP by colorimetric assay using p-nitrophenyl phosphate as substrate and diethanolamine as buffer and also to compare the ALP levels in patients with differen...

Turan, Serap; Topcu, Burcu; Go?kc?e, Ibrahim; Gu?ran, Tu?lay; Atay, Zeynep; Omar, Anjumanara; Akc?ay, Teoman; Bereket, Abdullah

2011-01-01

12

Peripartal changes in serum alkaline phosphatase activity and lactate dehydrogenase activity in dairy cows.  

Digital Repository Infrastructure Vision for European Research (DRIVER)

Peripartal serum alkaline phosphatase activity and lactate dehydrogenase activity were measured in 30 dairy cows in order to examine the association between retained fetal membranes and enzyme activity. Daily blood samples were obtained from pregnant cows, starting 15 days before the expected day of calving until eight days after parturition. Sera from 15 cows which retained fetal membranes longer than 24 hours and 15 cows which shed fetal membranes within six hours after parturition were ana...

Peter, A. T.; Bosu, W. T.; Macwilliams, P.; Gallagher, S.

1987-01-01

13

ALP (Alkaline Phosphatase) Test  

Science.gov (United States)

... Also known as: ALK PHOS; Alkp Formal name: Alkaline Phosphatase Related tests: AST ; ALT ; GGT ; Bilirubin ; Liver Panel ; Bone Markers ; Alkaline Phosphatase Isoenzymes; Bone Specific ALP At a Glance Test ...

14

[Assays of placenta-iso-enzyme of alkaline serum-phosphatase with disk-elektrophoresis (author's transl)].  

Science.gov (United States)

Alkaline serum-phosphatase may be subdivided with disk-electrophoresis in 7% poly-acrylamid gel into the iso-enzymes: liver-cell, bone- and ileum-phosphatase. In the 7% gel liver-cell and placenta phosphatase move together. Separation of these two can be achieved by raising the gel-concentration to 10%. The method is suitable for quantitative densitometric evaluation. PMID:1220356

Liebe, S; Schlegel, C

1975-12-01

15

Correlation Between Serum Level Parathormone, Alkaline Phosphatase, Calcium and Phosphorus of Patients Hemodialysis in Zahedan  

Directory of Open Access Journals (Sweden)

Full Text Available Secondary hyperparathyroidism and its effects on bone tissue are among the most important complication of end-stage renal disease. In the present study, we investigated correlation between the serum parathormone level (PTH of hemodialysis men and women with calcium (Ca, phosphorus (Pi and alkaline phosphatase (ALP. We studied 30 chronic renal failure hemodialysis patients 16 men and 14 women, aged 22-66 years old (average 44 years old with dialysis duration of 5 months to 14 years. We measured the serum Ca., Pi and ALP in intervals of 3 months and serum PTH levels was measured in 3 month. Data analysis was performed using SPSS software. Our results showed that serum PTH and ALP levels were higher in women than in men (90% versus 70%, but abnormal range of serum Ca and Pi levels were higher in men then women (Ca : 8% versus 2%, Pi :58% versus 50%. Hemodialysis patients showed correlation between PTH and ALP (p<0.05, but the correlation of PTH with Ca and Pi levels was not statistically significant. No correlation was observed between PTH and ALP and Pi in men, however it was significant between PTH and Ca (p<0.08, r = - 0.63. The women showed correlation between PTH and ALP (p<0.05, but not between Ca and Pi levels. Based on the findings of this study, Secondary hyperparathyroidism and its effects on bone tissue were greater in women than men hemodialysis.

R. Saravani

2007-01-01

16

Pesticide-induced Changes in Serum Levels of Acid Phosphatase, Alkaline Phosphates and Glutamate Oxaloacetate Transaminase in Rats  

Directory of Open Access Journals (Sweden)

Full Text Available Three insecticides (Monitor, Talstar and Benzenehexachloride {BHC}, orally administered at the doses of 0.06, 0.5 and 0.2 mg kg 1 body weight respectively for 21 days affected the body weight and biochemical environment of blood and liver in the rats. Talstar treatment resulted in an increase in the body weight while Monitor and BHC treatment reduced the body weight. After 21 days of the Monitor treatment, activities of acid phosphatases increased (55% and alkaline phosphates decreased (2% in the serum while slight inhibition was observed in the serum glutamate oxaloacetate transaminase activity. Benzenehexachloride treatment resulted in increased serum levels of acid phosphatase, alkaline phosphatase and glutamate oxaloacetate transaminase (42, 2 and 18% respectively. Talstar treatment decreased the activities of acid phosphatase and alkaline phosphatase (76 and 5%, respectively while glutamate oxaloacetate transaminase activity was increased (25%. These data suggest that chronic exposure to Monitor, Talstar and BHC causes hepatocyte necrosis and increases the liver enzyme synthesis.

Muhammad Nasim Khan

2003-01-01

17

[A mathematical model for the study of the kinetics of serum alkaline phosphatase inhibition by sodium diethyldithiocarbamate (NaDDTC)].  

Science.gov (United States)

A mathematical model for the kinetic of the serum alkaline phosphatase inhibition by NaDDTC is described in this paper. The model is tested over an experimental data set consisting on measured % residual activity of enzyme in human serum, after inhibition. The Bessey method is used. The solution of the differential equations for the model is found to be the best fitting for the observed time-dependent phenomena. The rate constant, K, for the inhibition reaction is then computed. PMID:6264531

Jovine, R; Ghezzo, F; Orecchio, F; Savarese, N; Ficarra, M G

1980-03-01

18

Serum Alkaline Phosphatase and Phosphate and Risk of Mortality and Hospitalization  

Digital Repository Infrastructure Vision for European Research (DRIVER)

Background and objectives: Elevated alkaline phosphatase (AlkPhos) and phosphate levels are associated with cardiovascular morbidity and mortality in patients receiving dialysis. A retrospective cohort study was conducted to test these associations in outpatients with an estimated GFR ?60 ml/min/1.73 m2.

Abramowitz, Matthew; Muntner, Paul; Coco, Maria; Southern, William; Lotwin, Irwin; Hostetter, Thomas H.; Melamed, Michal L.

2010-01-01

19

ESTIMATION OF SERUM ALKALINE PHOSPHATASE, CHOLESTEROL, CALCIUM AND PHOSPHORUS DURING PRE-LA YING AND LAYING CONDITIONS IN DIFFERENT STRAINS OF CHICKENS  

Digital Repository Infrastructure Vision for European Research (DRIVER)

In order to estimate serum alkaline phosphatase, cholesterol, calcium and phosphorus during pre-laying and laying reproductive conditions, 60 hens of Desi, Fayoumi, Cross (Rhode Island Red X Fayoumi} and Nick Chick strains were maintained for one year. Five random blood samples from each strain were collected and analyzed during both pre egg laying and egg laying physiological conditions. It was observed that alkaline phosphatase activity increased significantly (P<0.05) during laying conditi...

Bashir Mahmood Bhatti, Tanzeela Talat And Rozina Sardar

2002-01-01

20

Multicenter evaluation of Iso-ALP test kit for measurement of bone alkaline phosphatase activity in serum and plasma.  

Science.gov (United States)

A test kit (Iso-ALP, Boehringer Mannheim) for measuring human bone alkaline phosphatase activity in serum or plasma was evaluated in five laboratories in three countries. The assay is based on the principle described by Rosalki and Foo (Clin Chem 1984;30:1182-6) and uses wheat germ lectin to precipitate bone alkaline phosphatase. Residual ALP in the supernate in comparison with total ALP is used to quantify the bone fraction. The imprecision of residual ALP measurement was low (median between-run CV 4.9%) and comparable with that of total ALP. Linearity of precipitation was demonstrable up to a bone ALP activity (diethanolamine buffer 37 degrees C) of 2000 U/L, though a matrix effect was observed for dilutions of high-activity sera in saline or bovine serum albumin. For assaying patients' samples, different batches of lectin demonstrated excellent comparability. Taking electrophoresis as a basis for standardization, we determined that the lectin precipitated approximately 90% of bone ALP, but < 5% of nonbone ALP. From this we derived serum/plasma upper reference limits for bone ALP activity in adults and children. PMID:8472360

Rosalki, S B; Foo, A Y; Burlina, A; Prellwitz, W; Stieber, P; Neumeier, D; Klein, G; Poppe, W A; Bodenmüller, H

1993-04-01

 
 
 
 
21

The activity of bone and intestinal isoenzymes of serum alkaline phosphatase as a measure of radioprotective efficiency  

International Nuclear Information System (INIS)

The activity of serum alkaline phosphatase was investigated in 3 month-old Wistar rats, weighing 200 - 220 g. In addition to the total activity the activity of bone and intestinal isoenzymes was determined in the serum by the heat inactivation-inhibition method. Determination was performed in control rats, in rats irradiated with a whole-body dose of 650 R (60Co) and in those treated with cystamine (i.p. 50 mg/kg) or with mexamine (i.m. 10 mg/kg). The activity of alkaline phosphatase was followed up to the 15th day after irradiation and administration of radioprotective substances. In spite of a pronounced decrease of the activity, mainly of bone isoenzyme, there was a distinct turn in rats treated with radioprotecte substances from the 6th day on to values found in control rats. The results can be used for the evaluation of the efficiency of radioprotective compounds and as a simple biochemical criterion of the radioprotective effect during the development of both intestinal and bone radiation syndrome. (author)

22

Influence of radioprotective agents on the activities of isoenzymes of blood serum alkaline phosphatase in irradiated dogs  

International Nuclear Information System (INIS)

Changes of the total serum activity of alkaline phosphatase (AP) and its bone and intestinal isoenzymes were studied in dogs ?-irradiated by 3.0 Gy. The activities of AP isoenzymes were determined by means of a heat inactivation-inhibition method. After irradiation the serum AP activities were lower in general. In case of the bone isoenzyme the decrease was most pronounced. The changes of the total AP activity and of the intestinal isoenzyme were less distinct. The i.m. administration of the radioprotective mixture cystamine with mexamine (24 mg/kg + 4 mg/kg) before irradiation led only to a moderate reduction of the radiation-altered AP values in the dogs. The present results indicate that the radioprotective way used here renders only an unsatisfactory protection for large laboratory animals against ionizing radiation. (orig.)

23

Influence of radioprotective agents on the activities of isoenzymes of blood serum alkaline phosphatase in irradiated dogs.  

Science.gov (United States)

Changes of the total serum activity of alkaline phosphatase (AP) and its bone and intestinal isoenzymes were studied in dogs gamma-irradiated by 3.0 Gy. The activities of AP isoenzymes were determined by means of a heat inactivation-inhibition method. After irradiation the serum AP activities were lower in general. In case of the bone isoenzyme the decrease was most pronounced. The changes of the total AP activity and of the intestinal isoenzyme were less distinct. The i.m. administration of the radioprotective mixture cystamine with mexamine (24 mg/kg + 4 mg/kg) before irradiation led only to a moderate reduction of the radiation-altered AP values in the dogs. The present results indicate that the radioprotective way used here renders only an unsatisfactory protection for large laboratory animals against ionizing radiation. PMID:6930747

Krízala, J; Kuna, P; Stoklasová, A; Ledvina, M; Dostál, M

1980-05-01

24

[Biological effects of dithiocarbamates: changes in serum alkaline phosphatase activity "in vitro" by sodium diethyldithiocarbamate (NADDTC) (author's transl)].  

Science.gov (United States)

The Authors demonstrate that serum alkaline phosphatase (s-ALP) activity in vitro is inhibited by NaDDTC. This may be related to the NaDDTC chelating action on zinc enzyme. The trend of the phenomenon follows an exponential pattern. The activity of the isozyme pool was restored after removal from medium of NaDDTC by dialysis. The NaDDTC concentration able to inhibit in vitro enzymatic activity was nearly one thousand times higher than that found in vivo, in experimental animals (rabbit). The Authors conclude that the in vivo activity of NaDDTC in different enzymatic systems, such as ALP and other metallo-dependent enzymes, cannot be explained by its chelating action alone, but also by its influence on other systems. At present investigations in this field are in progress in our laboratory. PMID:6264534

Orecchio, F; Maggiore, A; Savarese, N; Ficarra, M G

1980-12-01

25

ESTIMATION OF SERUM ALKALINE PHOSPHATASE, CHOLESTEROL, CALCIUM AND PHOSPHORUS DURING PRE-LA YING AND LAYING CONDITIONS IN DIFFERENT STRAINS OF CHICKENS  

Directory of Open Access Journals (Sweden)

Full Text Available In order to estimate serum alkaline phosphatase, cholesterol, calcium and phosphorus during pre-laying and laying reproductive conditions, 60 hens of Desi, Fayoumi, Cross (Rhode Island Red X Fayoumi} and Nick Chick strains were maintained for one year. Five random blood samples from each strain were collected and analyzed during both pre egg laying and egg laying physiological conditions. It was observed that alkaline phosphatase activity increased significantly (P<0.05 during laying condition. Serum cholesterol remained unaffected by both the strain difference and laying condition. Serum calcium and phosphorus levels increased (P<0.05 during laying condition. The interaction of strain and stage of laying condition was found to exert significant (P<0.05 effect on serum calcium levels. The study showed that the availability of calcium and phosphorus in requisite quantities be provided in diet of laying hens to ensure sustained and quality egg production.

Bashir Mahmood Bhatti, Tanzeela Talat and Rozina Sardar

2002-04-01

26

Changes in the activity of bone and intestinal isoenzymes of serum alkaline phosphatases in irradiated rabbits treated with radioprotective substances  

International Nuclear Information System (INIS)

The influence of radioprotective substances on post-irradiation changes of the activity of total alkaline phosphatase (AP, E.C. 3.1.3.1.) and its bone and intestine isoenzymes in the blood serum was studied in rabbits. After whole-body exposure to 8 Gy (800 R), a marked decrease of the AP activity occurred (both total and isoenzyme activity) as early as the 1st day, and persists over the entire period investigated (from the 1st to the 21st day). 5-methoxytryptamine given i.p. or s.c. administration of cystamine caused only a moderate effect on the radiation-induced changes in AP levels. However, a slow i.v. infusion of cystamine reversed the decrease more effectively and the activity values were similar to the irradiated controls. The results indicate that the investigation of the activity changes in isoenzymes of serum AP may serve not only as an indicator of radiation damage, but also as a biochemical criterion of the radioprotective efficiency. (author)

27

Alkaline phosphatase and transaminase activity in rat liver and blood serum at delayed periods following external ?-irradiation combined with internal exposure to plutonium 239  

International Nuclear Information System (INIS)

A study was made of activity of alkaline phosphatase and alanine- and aspartate aminotransferase in rat liver and blood serum at remote times after external ?-irradiation combined with internal exposure to 239Pu nitrate delivered in two chronically effective doses. The radionuclide was shown to be mainly responsible for the changes observed in activity of the enzymes under study. The degree to which the changes were manifest depended upon dose of plutonium administered

28

Profile of serum alkaline phosphatase after inoculation of mononuclear cells and bone morphogenetic protein in the repair of osteochondral defects in rabbits  

Digital Repository Infrastructure Vision for European Research (DRIVER)

In this study, serum alkaline phosphatase activity was measured in response to the repair of osteochondral defects in twenty-four New Zealand rabbits. The animals were divided into three groups: a control (GC), those treated with bone marrow mononuclear cells (GCM) and those that received mononuclear cells with autologous bone morphogenetic protein (BMP + GCM). After exposing the trochlear groove of the left stifle joint, a wedge-shaped segment was removed. Later, the defect was filled with a...

Luiz Augusto Souza; Benito Juarez Nunes Alves de Oliveira; Duvaldo Eurides; Ednaldo Carvalho Guimarães; Luiz Antônio Franco da Silva; Lorena Borges Alves; Ana Flávia Delben Pereira Arruda; Taís Andrade Dias

2011-01-01

29

Serum Proteins and Alkaline Phosphatase Levels in Patients with Tuberous Sclerosis  

Science.gov (United States)

Six 4- to 37-year-old patients with tuberosis sclerosis (a chronic condition characterized by siezures, intercranial calcification, a reddish-yellow sebaceous glandular mass on the face, and frequent crises in early years), did not exhibit an elevation of the (alpha + beta) globulin fraction in their serum. (Author/MC)

Fischer, M. H.; And Others

1974-01-01

30

Pesticide-induced Changes in Serum Levels of Acid Phosphatase, Alkaline Phosphates and Glutamate Oxaloacetate Transaminase in Rats  

Digital Repository Infrastructure Vision for European Research (DRIVER)

Three insecticides (Monitor, Talstar and Benzenehexachloride {BHC}), orally administered at the doses of 0.06, 0.5 and 0.2 mg kg 1 body weight respectively for 21 days affected the body weight and biochemical environment of blood and liver in the rats. Talstar treatment resulted in an increase in the body weight while Monitor and BHC treatment reduced the body weight. After 21 days of the Monitor treatment, activities of acid phosphatases increased (55%) and alkaline phosphates dec...

Muhammad Nasim Khan; Tahira Sarwar

2003-01-01

31

21 CFR 862.1050 - Alkaline phosphatase or isoenzymes test system.  

Science.gov (United States)

...isoenzymes test system is a device intended to measure alkaline phosphatase or its isoenzymes (a group of enzymes with similar biological activity) in serum or plasma. Measurements of alkaline phosphatase or its isoenzymes are used in the...

2010-04-01

32

Profile of serum alkaline phosphatase after inoculation of mononuclear cells and bone morphogenetic protein in the repair of osteochondral defects in rabbits  

Directory of Open Access Journals (Sweden)

Full Text Available In this study, serum alkaline phosphatase activity was measured in response to the repair of osteochondral defects in twenty-four New Zealand rabbits. The animals were divided into three groups: a control (GC, those treated with bone marrow mononuclear cells (GCM and those that received mononuclear cells with autologous bone morphogenetic protein (BMP + GCM. After exposing the trochlear groove of the left stifle joint, a wedge-shaped segment was removed. Later, the defect was filled with an osteochondral autograft preserved in 98% glycerin. For the GC group, only the bone graft was performed. For the GCM, in addition to the graft, 2x106 seed mononuclear cells were implanted. For the GCM + BMP, the same number of cells, associated with 1?g of bone morphogenetic protein, were intraarticularly administered. The osteoblastic response was measured by analyzing the serum alkaline phosphatase on day 0 (preoperative 3, 15, 30, and 45 after surgery, and by radiographic examinations. Analysis of variance in randomized blocks, factorial and Tukey’s test (p = 0.05 were made. The overall mean GCM was superior to the other groups and the highest rates were among the 15th and 45th days postoperatively. The discrepancy in values between individuals of the same group casts doubts on the veracity of the test.

Luiz Augusto de Souza

2011-12-01

33

21 CFR 864.7660 - Leukocyte alkaline phosphatase test.  

Science.gov (United States)

...2010-04-01 false Leukocyte alkaline phosphatase test. 864.7660 Section... § 864.7660 Leukocyte alkaline phosphatase test. (a) Identification. A leukocyte alkaline phosphatase test is a device used...

2010-04-01

34

Relationship between serum heat-stable alkaline phosphatase activity and blood pressure in patients with pre-eclampsia and eclampsia  

Directory of Open Access Journals (Sweden)

Full Text Available Background : The objective of this study was to explore the relationship, if any, between theserum heat-stable alkaline phosphatase (HS-ALP activity and the blood pressure (BP of patients with pre-eclampsia and eclampsia. Method : The activity of HS-ALP was measured using the 4– nitrophenyl phosphate (4– NPP method after incubation at a high temperature of 65 0 C for exactly 30 minutes in one hundred normal pregnant women and in another one hundred with pre-eclampsia/eclampsia. The normal pregnant women were used as controls. The blood pressure (BP, systolic as well as diastolic was measured in each of the studied patient using desktop mercury sphygmomanometer. Results :In the patients with pre-eclampsia/eclampsia, it was found that the higher the systolic and diastolic BP, the higher is the activity of the HS-ALP. Conclusion : It can be concluded that the HS-ALP activity in patients with pre-eclampsia/eclampsia is positively related to the severity of the hypertension and therefore this could help in detecting early complication.

Aliyu I

2006-03-01

35

Effects of 60Co gamma-ray local irradiation on rat liver on alkaline phosphatase, lactate dehydrogenase and catalase in the liver and serum  

International Nuclear Information System (INIS)

Rats were given a single exposure of various doses (0, 5, 50, 500, and 5000 rads) to local irradiation of 60Co ?-ray on liver. Activities of alkaline phosphatase (ALP), lactate dehydrogenase (LDH) and catalase in the serum and liver were measured at various time intervals after irradiation. These results were summarized as follows; 1. ALP activity in the serum had no effect on irradiation up to 500 rads, but in the case of 5000 rads irradiation exhibited a marked loss from 4 days after irradiation. ALP activity in the liver to 5000 rads exposure on 7 days after irradiation increased, on the other hand in the serum decreased, and the patterns of ALP activities in the liver and serum to the irradiation doses were opposite. 2. LDH activity in the serum by exposure to 5, 500 and 5000 rads increased at 4 days after irradiation, but at 7 days significantly decreased. LDH activity in the liver to the irradiation doses on 7 days after irradiation did not markedly change, but in the serum it tended to be low in inverse proportion to the irradiation doses. 3. Catalase activity in the serum to 50 and 500 rads exposure increased at 4 days after irradiation and decreased at 7 days, but to 5000 rads exposure it decreased in the course of time. Catalase activity in the liver and serum on 7 days after irradiation were inversely proportional to irradiation doses. It is difficult that catalase activity makes a index of clinical irradiation effects, because catalase activityiation effects, because catalase activity decrease under the various conditions, such as cancer, anemia, infection of bacterias and so on. Since activities of ALP and LDH increase in almost disease, decrease of ALP activity and decrease following temporary increase of LDH activity by irradiation may be able to become a clinical indicator on irradiation effects. (author)

36

Automated and semiautomated analysis of rat alkaline phosphatase isoenzymes.  

Science.gov (United States)

A semiautomated quantitative assay for rat serum alkaline phosphatase (ALP) isoenzyme determination was developed, incorporating selective precipitation of bone alkaline phosphatase (BALP) with wheat germ lectin and differential inhibition with levamisole for determination of intestinal alkaline phosphatase (IALP). The assays for each isoenzyme were linear over a broad range of activities. The within-run and between-run coefficients of variation were less than 11% for all 3 isoenzymes. Dilution of serum with saline results in an artifactual overestimation of BALP activity. Comparison of ALP and ALP isoenzyme activity in rats of various ages showed that BALP activity drops dramatically with increasing age of rats. IALP activity is greater in immature rats compared to that in mature rats. While there was no difference between male and female rats at 4 wk of age with regard to total ALP activity and activity of any of the isoenzymes, total ALP activity and activity of the individual isoenzymes were higher in males than in females at most ages over 4 wk. Gavage with corn oil resulted in increased serum IALP activity, and bile duct ligation resulted in increased liver alkaline phosphatase activity. This combined assay for the 3 ALP isoenzymes in rat serum is an efficient means of analysis of large numbers of samples and should increase markedly the specificity of serum ALP activity in identifying the target organ in toxicologic studies when serum ALP activity is increased. PMID:7732280

Hoffmann, W E; Everds, N; Pignatello, M; Solter, P F

1994-01-01

37

Serum proteins, trace metals and phosphatases in psoriasis  

Directory of Open Access Journals (Sweden)

Full Text Available Serum proteins, zinc, copper, acid phosphatase (AcPase and alkaline phosphatase (AlPase were studied in both active and remission phases of psoriasis. Data were compared with healthy controls, ?1, ? and ? globulins were high in active phase while ?1 and ? globulins were at par in remission phase. Serum copper was low but zinc and alkaline phosphatase were significantly high in both active and remission phases of the disease. Acid phosphatase level was at par in all the experimental groups. Study suggests a positive correlation of globulin, zinc and Alpase in active and remission phase of psoriasis.

Bhatnagar M

1994-01-01

38

[Consideration on the King-Armstrong method and on the kinetic method for the measurement of the alkaline phosphatase activity in serum (author's transl)].  

Science.gov (United States)

The comparative study of a determined time method, according to King-Armstrong, for the measurement of the alkaline phosphatase activity in serum and of a kinetic method consents us to make the following points: 1) the reagents used in the King-Armstrong method are easily prepared and are stable for months, with the sole addition of cloroform to the substrate and to the buffer to avoid the growth of moulds. 2) The substrate used for the kinetic method undergoes spontaneous Xydrolysis in acqueous solution, which makes it possible to keep it refrigerated for a limited number of days. Freezing at - 20 degrees C does not improve the situation. 3) The degree of precision within the series and between the series is the same for the two methods, and very good. 4) From the working point of view the method of "determined time" is advantageous for few samples, while the kinetic method becomes preferable when the number of samples goes over 20 and 30 a day, provided that it is linked with the suitable automatic apparatus. 5) The results of the measurement of the enzyme activity found with the two methods on 120 sera of normal and pathological subjects have shown good correlation, confirming the clinical diagnostic value of the results obtained by the kinetic method, which uses a transphosphorilant buffer and their congruency with those obtained by the classical King-Armstrong method. PMID:1030811

Spandrio, L; Panigada, C

1976-06-01

39

Cationized dextran nanoparticle-encapsulated CXCR4-siRNA enhanced correlation between CXCR4 expression and serum alkaline phosphatase in a mouse model of colorectal cancer  

Directory of Open Access Journals (Sweden)

Full Text Available Fatemeh Abedini,1 Hossein Hosseinkhani,2 Maznah Ismail,1,3 Abraham J Domb,4 Abdul Rahman Omar,1,5 Pei Pei Chong,1,2 Po-Da Hong,3 Dah-Shyong Yu,6 Ira-Yudovin Farber41Laboratory of Molecular Biomedicine, Institute of Bioscience, Universiti Putra Malaysia, Selangor, 2Graduate Institute of Biomedical Engineering, National Taiwan University of Science and Technology, Taipei, Taiwan, 3Faculty of Medicine & Health Sciences, Universiti Putra Malaysia, Selangor, Malaysia, 4Institute of Drug Research, The Center for Nanoscience and Nanotechnology, School of Pharmacy-Faculty of Medicine, The Hebrew University of Jerusalem, Jerusalem, Israel, 5Faculty of Veterinary Medicine, Universiti Putra Malaysia, Selangor, Malaysia, 6Nanomedicine Research Center, National Defense Medical Center, Taipei, TaiwanPurpose: The failure of colorectal cancer treatments is partly due to overexpression of CXCR4 by tumor cells, which plays a critical role in cell metastasis. Moreover, serum alkaline phosphatase (ALP levels are frequently elevated in patients with metastatic colorectal cancer. A polysaccharide, dextran, was chosen as the vector of siRNA. Spermine was conjugated to oxidized dextran by reductive amination process to obtain cationized dextran, so-called dextran-spermine, in order to prepare CXCR4-siRNAs/dextran-spermine nanoparticles. The fabricated nanoparticles were used in order to investigate whether downregulation of CXCR4 expression could affect serum ALP in mouse models of colorectal cancer.Methods: Colorectal cancer was established in BALB/C mice following injection of mouse colon carcinoma cells CT.26WT through the tail vein. CXCR4 siRNA for two sites of the target gene was administered following injection of naked siRNA or siRNA encapsulated into nanoparticles.Results: In vivo animal data revealed that CXCR4 silencing by dextran-spermine nanoparticles significantly downregulated CXCR4 expression compared with naked CXCR4 siRNA. Furthermore, there was correlation between CXCR4 expression and serum ALP.Conclusion: CXCR4 siRNA/dextran-spermine nanoparticles appear to be highly effective, and may be suitable for further in vivo applications. Further research evaluation will be needed to determine the effect of CXCR4 silencing on serum ALP levels, which may be a useful marker to predict liver metastasis in colorectal cancer.Keywords: nanoparticles, cationized dextran, colorectal cancer, serum ALP enzyme, CXCR4, siRNA

Abedini F

2012-07-01

40

The effect of long term anticonvulsant therapy on serum calcium, phosphate and alkaline phosphatase in Nigerian epileptic patients.  

Science.gov (United States)

A survey of calcium metabolism in fifty-two epileptic patients on long term therapy with anti-convulsant drugs, showed that their total serum calcium levels were within normal limits. There was neither clinical nor radiological evidence of osteomalacia in any of them. It is postulated therefore that the osteomalacic effect of antivonculsant drugs reported among Caucasians does not appear to occur in African patients in a tropical environment most probably due to the abundance of vitamin D derived from the sun's ultra-violet light. PMID:1216324

Apantaku, J B; Afonja, O A; Boyo, A E

1975-12-01

 
 
 
 
41

Ratiometric electrochemical detection of alkaline phosphatase.  

Science.gov (United States)

A novel ferrocene-derived substrate for the ratiometric electrochemical detection of alkaline phosphatase (ALP) was designed and synthesised. It was demonstrated to be an excellent electrochemical substrate for the ALP-labelled enzyme-linked immunosorbent assay (ELISA). PMID:25413385

Goggins, Sean; Naz, Christophe; Marsh, Barrie J; Frost, Christopher G

2015-01-11

42

Odontoblast alkaline phosphatases and Ca2+ transport.  

Science.gov (United States)

The same isoenzyme of nonspecific alkaline phosphatase (APase), assayed with p-nitrophenylphosphate (p-NPP), was shown be present in different calcifying tissues, bone, calcifying cartilage, odontoblasts and enamel organ. Indications were also found that the enzymatic degradation of inorganic pyrophosphate (PPi) in calcifying tissues is mediated by APase. By using specific APase inhibitors, it was shown that two enzymes capable of degrading ATP exist. These were characterized in dentinogenically active odontoblasts, and it was concluded that one is the classical APase, the other is a Ca2+ and Mg2+ activated ATPase, named Ca2+-ATPase. The two phosphatases were solubilized from odontoblasts and separated. The localization of APase and Ca2+-ATPase in odontoblasts was investigated by subcellular fractionation and EM histochemistry. Routine methods for fixation were found to almost completely inactivate the enzymes. By using a mild fixation technique that preserved 80% of the enzyme activity, the main localization for both APase and Ca2+-ATPase was found to be in the membranes of intercellular vesicles located in the cell body and odontoblasts process. No activity was found in the cell membranes. It is concluded that there are at least two enzymes able to degrade phosphate compounds at alkaline pH in hard tissue forming cells. One is the nonspecific alkaline phosphatase (APase; EC 3. 1. 3. 1), which is active against p-NPP, PPi, glycerophosphates and ATP among other substrates. The other is a more specific Ca2+-ATPase (EC 3. 6. 1. 3). There seems to be an intimate relation between these two enzymes in the tissue. The function of APase in biological calcification is still obscure. In contrast, the finding of an ATP dependent, intravesicularly directed, transmembranous Ca2+-transport in vesicles derived from the microsomal fraction of odontoblasts may explain the role of Ca2+-ATPase. PMID:153909

Linde, A; Granström, G

1978-12-01

43

Mutation of a single amino acid converts germ cell alkaline phosphatase to placental alkaline phosphatase.  

Science.gov (United States)

Human placental and germ cell alkaline phosphatases (PLAP and GCAP, respectively), are characterized by their differential sensitivities to inhibition by L-leucine, EDTA, and heat. Yet, they differ by only 7 amino acids at positions 15, 67, 68, 84, 241, 254, and 429 within their respective 484 residues. To determine the structural basis and the amino acid(s) involved in these physicochemical differences, we constructed three GCAP mutants by site-directed mutagenesis and six GCAP/PLAP chimeras and then expressed these alkaline phosphatase mutants in COS-1 cells. We report that the differential reactivity of PLAP and GCAP depends critically on a single amino acid at position 429. GCAP with Gly-429 is strongly inhibited by L-leucine, EDTA, and heat, whereas PLAP with Glu-429 is resistant. By substituting Gly-429 of GCAP with a series of amino acids, we demonstrate that the relative sensitivities of these mutants to L-leucine, EDTA, and heat inhibition are, in general, parallel. Mutants in the order of resistance to these treatments are: Glu (most resistant), Asp/Ile/Leu, Gln/Val/Lys, Ser/His, and Arg/Thr/Met/Cys/Phe/Trp/Tyr/Pro/Asn/Ala/Gly (least resistant). However, the Ser-429 and His-429 mutants were more resistant to EDTA and heat inhibition than the wild-type GCAP, but were equally sensitive to L-leucine inhibition. Structural analysis of mammalian alkaline phosphatase modeled on the refined crystal structure of Escherichia coli alkaline phosphatase indicates that the negative charge of Glu-429 of PLAP, which simultaneously stabilizes the protein as a whole and the metal binding specifically, probably acts through interactions with the metal ligand His-320 (His-331 in E. coli alkaline phosphatase). Replacement of codon 429 with Gly in GCAP leads to destabilization and loosening of the metal binding. The data suggest that the natural binding site for L-leucine may be near position 429, with the amino and carboxyl groups of L-leucine interacting with bound phosphate and His-432 (His-412 in E. coli alkaline phosphatase), respectively. PMID:1939159

Watanabe, T; Wada, N; Kim, E E; Wyckoff, H W; Chou, J Y

1991-11-01

44

Human placental alkaline phosphatase in liver and intestine  

International Nuclear Information System (INIS)

Three distinct forms of human alkaline phosphatase, presumably isozymes, are known, each apparently associated with a specific tissue. These are placental, intestinal, and liver (kidney and bone). The authors have used a specific immunoassay and HPLC to show that placental alkaline phosphatase is also present in extracts of liver and intestine in appreciable amounts

45

Effect of alkaline phosphatase on thromboxane mimetic induced platelet activation.  

Science.gov (United States)

Recently it has been reported that alkaline phosphatase selectively inhibited thromboxane mimetic induced platelet aggregation and secretion suggesting that the phosphorylation state on the platelet surface may be important for thromboxane induced platelet activation. We report here studies attempting to elucidate the mechanism of action of alkaline phosphatase. Washed human platelet aggregation induced by the thromboxane mimetic IBOP was completely abolished when incubated with alkaline phosphatase (1 unit/ml) for 5 min. The effect was inhibited by co-incubation with 5mM phosphate. Binding studies using [125I]BOP showed that neither the affinity of IBOP for the receptor (control: 9.2 +/- 2.1 nM, alkaline phosphatase: 7.9 +/- 1.8 nM) nor the Bmax (control: 1780 +/- 320 sites/plt. alkaline phosphatase: 1920 +/- 290 sites/plt) were effected by alkaline phosphatase treatment. GTPase activity was measured in platelet membranes treated with and without alkaline phosphatase as measured by IBOP induced hydrolysis of [gamma-32P]GTP. The EC50 values for IBOP induced GTPase were similar whereas the maximum amount of released Pi in the control membranes was more than two fold greater than in alkaline phosphatase treated membranes. These studies suggest that thromboxane induced platelet activation may be dependent upon the phosphorylation state of the thromboxane receptor and/or closely associated protein. PMID:7878191

Margolin, N; True, T A; Saussy, D L; Mais, D E

1994-10-01

46

21 CFR 862.1050 - Alkaline phosphatase or isoenzymes test system.  

Science.gov (United States)

... 2010-04-01 false Alkaline phosphatase or isoenzymes test system...Systems § 862.1050 Alkaline phosphatase or isoenzymes test system. (a) Identification. An alkaline phosphatase or isoenzymes test...

2010-04-01

47

Phosphatidylinositol anchor of HeLa cell alkaline phosphatase  

Energy Technology Data Exchange (ETDEWEB)

Alkaline phosphatase from cancer cells, HeLa TCRC-1, was biosynthetically labeled with either /sup 3/H-fatty acids or (/sup 3/H)ethanolamine as analyzed by sodium dodecyl sulfate-polyacrylamide gel electrophoresis and fluorography of immunoprecipitated material. Phosphatidylinositol-specific phospholipase C (PI-PLC) released a substantial proportion of the /sup 3/H-fatty acid label from immunoaffinity-purified alkaline phosphatase but had no effect on the radioactivity of (/sup 3/H)ethanolamine-labeled material. PI-PLC also liberated catalytically active alkaline phosphatase from viable cells, and this could be selectively blocked by monoclonal antibodies to alkaline phosphatase. However, the alkaline phosphatase released from /sup 3/H-fatty acid labeled cells by PI-PLC was not radioactive. By contrast, treatment with bromelain removed both the /sup 3/H-fatty acid and the (/sup 3/H)ethanolamine label from purified alkaline phosphatase. Subtilisin was also able to remove the (/sup 3/H)ethanolamine label from the purified alkaline phosphatase. The /sup 3/H radioactivity in alkaline phosphatase purified from (/sup 3/H)ethanolamine-labeled cells comigrated with authentic (/sup 3/H)ethanolamine by anion-exchange chromatography after acid hydrolysis. The data suggest that the /sup 3/H-fatty acid and (/sup 3/H)ethanolamine are covalently attached to the carboxyl-terminal segment since bromelain and subtilisin both release alkaline phosphatase from the membrane by cleavage at that end of the polypeptide chain. The data are consistent with findings for other proteins recently shown to be anchored in the membrane through a glycosylphosphatidylinositol structure and indicate that a similar structure contributes to the membrane anchoring of alkaline phosphatase.

Jemmerson, R.; Low, M.G.

1987-09-08

48

Alkaline phosphatase isoenzyme activities in rheumatoid arthritis: hepatobiliary enzyme dissociation and relation to disease activity.  

Digital Repository Infrastructure Vision for European Research (DRIVER)

OBJECTIVES--Hyperphosphatasaemia has been observed occasionally in patients with rheumatoid arthritis (RA), and it has been suggested that the serum alkaline phosphatase (ALP) level is related to the activity of the disease. Therefore, the relationship between serum ALP and RA was studied. METHODS--The serum activities of hepatobiliary enzymes (ALP isoenzymes, gamma-glutamyltranspeptidase (GTP), leucine aminopeptidase (LAP), aspartate aminotransferase (AST), and alanine aminotransferase (ALT)...

Aida, S.

1993-01-01

49

Nucleotide and amino acid sequences of human intestinal alkaline phosphatase: close homology to placental alkaline phosphatase  

International Nuclear Information System (INIS)

A cDNA clone for human adult intestinal alkaline phosphatase (ALP) [orthophosphoric-monoester phosphohydrolase (alkaline optimum); EC 3.1.3.1] was isolated from a ?gt11 expression library. The cDNA insert of this clone is 2513 base pairs in length and contains an open reading frame that encodes a 528-amino acid polypeptide. This deduced polypeptide contains the first 40 amino acids of human intestinal ALP, as determined by direct protein sequencing. Intestinal ALP shows 86.5% amino acid identity to placental (type 1) ALP and 56.6% amino acid identity to liver/bone/kidney ALP. In the 3'-untranslated regions, intestinal and placental ALP cDNAs are 73.5% identical (excluding gaps). The evolution of this multigene enzyme family is discussed

50

Design and Synthesis of Selective Inhibitors of Placental Alkaline Phosphatase  

Digital Repository Infrastructure Vision for European Research (DRIVER)

Placental Alkaline Phosphatase (PLAP) is a tissue-restricted isozyme of the Alkaline Phosphatase (AP) superfamily. PLAP is an oncodevelopmental enzyme expressed during pregnancy and in a variety of human cancers, but its biological function remains unknown. We report here a series of catechol compounds with great affinity for the PLAP isozyme and significant selectivity over other members of the AP superfamily. These selective PLAP inhibitors will provide small molecule probes for the study o...

Lanier, Marion; Sergienko, Eduard; Sima?o, Ana Maria; Su, Ying; Chung, Thomas; Milla?n, Jose? Luis; Cashman, John R.

2010-01-01

51

Negative modulation of alkaline phosphatase and creatine kinase by homobrassinolide  

Digital Repository Infrastructure Vision for European Research (DRIVER)

Homobrassinolide is a plant hormone implicated in plant growth and development. Its effect on animal metabolism was less known to date. We have investigated its effect on the marker enzymes such as alkaline phosphatase and creatine kinase in selected rat tissues-brain, heart, liver, kidney, skeletal muscle and testis. Homobrassinolide was administered (66 and 330ng/ Kg body weight) intradermally in male albino wistar strain rats and changes in alkaline phosphatase and creatine kinase...

Nirmal Kumar, G.; Lakshmy, S.; Srikumar, K.

2011-01-01

52

Overexpression of Human Bone Alkaline Phosphatase in Pichia Pastoris  

Science.gov (United States)

The Pichiapastoris expression system was utilized to produce functionally active human bone alkaline phosphatase in gram quantities. Bone alkaline phosphatase is a key enzyme in bone formation and biomineralization, yet important questions about its structural chemistry and interactions with other cellular enzymes in mineralizing tissues remain unanswered. A soluble form of human bone alkaline phosphatase was constructed by deletion of the 25 amino acid hydrophobic C-terminal region of the encoding cDNA and inserted into the X-33 Pichiapastoris strain. An overexpression system was developed in shake flasks and converted to large-scale fermentation. Alkaline phosphatase was secreted into the medium to a level of 32mgAL when cultured in shake flasks. Enzyme activity was 12U/mg measured by a spectrophotometric assay. Fermentation yielded 880mgAL with enzymatic activity of 968U/mg. Gel electrophoresis analysis indicates that greater than 50% of the total protein in the fermentation is alkaline phosphatase. A purification scheme has been developed using ammonium sulfate precipitation followed by hydrophobic interaction chromatography. We are currently screening crystallization conditions of the purified recombinant protein for subsequent X-ray diffraction analyses. Structural data should provide additional information on the role of alkaline phosphatase in normal bone mineralization and in certain bone mineralization anomalies.

Karr, Laurel; Malone, Christine, C.; Rose, M. Franklin (Technical Monitor)

2000-01-01

53

Activity of Follicular Fluid Phosphatases and Their Correlation with Levels of Serum Esteroidal Hormones and Gonadotropins  

Digital Repository Infrastructure Vision for European Research (DRIVER)

Introduction: The aim of this study was to evaluate the correlation between serum levels of ovarian and gonadotropin hormones, age and number of follicles with follicular alkaline and acid phosphatase activity in infertile women under controlled ovarian hyperstimulation. Methods: After collection of follicular fluid and calculation of the number of follicles, the specific activity of alkaline (ALP) and acid phosphatase (ACP) was determined according to the total protein in 19 women at the tim...

Sh Byranvand; Rezaieyan, Z.; Saidi, H.; Khademi, A.; Safdarian, L.; Agha Hosseini, M.; Al Yassin, A.; Salehnia, M.

2006-01-01

54

Characterization of Human Bone Alkaline Phosphatase in Pichia Pastoris  

Science.gov (United States)

A soluble form of human bone alkaline phosphatase has been expressed in a recombinant strain of the methylotrophic yeast Pichia pastoris. We constructed a plasmid containing cDNA encoding for human bone alkaline phosphatase, with the hydrophobic carboxyl terminal portion deleted. Alkaline phosphatase was secreted into the medium to a level of 32mg/L when cultured in shake flasks, and enzyme activity was 12U/mg, as measured by a spectrophotometric assay. By conversion to a fermentation system, a yield of 880mg/L has been achieved with an enzyme activity of 968U/mg. By gel electrophoresis analysis, it appears that greater than 50% of the total protein in the fermentation media is alkaline phosphatase. Although purification procedures are not yet completely optimized, they are expected to include filtration, ion exchange and affinity chromatography. Our presentation will focus on the purification and crystallization results up to the time of the conference. Structural data should provide additional information on the role of alkaline phosphatase in normal bone mineralization and in certain bone mineralization anomalies.

Malone, Christine C.; Ciszak, Eva; Karr, Laurel J.

1999-01-01

55

Sensitive optical detection of alkaline phosphatase activity with quantum dots  

International Nuclear Information System (INIS)

A simple method has been developed to detect the activity of alkaline phosphatase (ALP) by the changing of fluorescence intensities of the quantum dots (QDs). In this system, the fluorescence intensities of the QDs were quenched by p-nitrophenol (pNP) which was produced in the process of ALP catalytic reaction. A series of linear calibration curves of the activity of ALP were obtained in different pH buffer solutions. The wide linear range was 3–1000 U L?1 and the detection limit was 3 U L?1 (S/N=3). Furthermore, the experimental conditions of biosensor were optimized, and anti-interference ability was presented. The activity of ALP was also detected in serum and the recovery of ALP in serum samples was more than 95%. The excellent performance of this biosensor indicates that it can be used in practice detection of ALP. -- Highlights: • A sensitive ALP biosensor is constructed based on QDs without complex processes. • The analysis processing is very convenient, simple and rapid. • The detection mechanism of the ALP biosensor is studied by XPS. • The paper proposes a feasible approach for some substrates or enzymes detecting

56

Sensitive optical detection of alkaline phosphatase activity with quantum dots  

Energy Technology Data Exchange (ETDEWEB)

A simple method has been developed to detect the activity of alkaline phosphatase (ALP) by the changing of fluorescence intensities of the quantum dots (QDs). In this system, the fluorescence intensities of the QDs were quenched by p-nitrophenol (pNP) which was produced in the process of ALP catalytic reaction. A series of linear calibration curves of the activity of ALP were obtained in different pH buffer solutions. The wide linear range was 3–1000 U L{sup ?1} and the detection limit was 3 U L{sup ?1} (S/N=3). Furthermore, the experimental conditions of biosensor were optimized, and anti-interference ability was presented. The activity of ALP was also detected in serum and the recovery of ALP in serum samples was more than 95%. The excellent performance of this biosensor indicates that it can be used in practice detection of ALP. -- Highlights: • A sensitive ALP biosensor is constructed based on QDs without complex processes. • The analysis processing is very convenient, simple and rapid. • The detection mechanism of the ALP biosensor is studied by XPS. • The paper proposes a feasible approach for some substrates or enzymes detecting.

Ren, Xiangling [Laboratory of Controllable Preparation and Application of Nanomaterials, Technical Institute of Physics and Chemistry, Chinese Academy of Sciences, No. 29, Zhongguancun East Road, Haidian District, Beijing 100190 (China); The State Key Laboratory of Bioelectronics, Southeast University, Nanjing 210096 (China); Chen, Zhenzhen; Chen, Xiaoying; Liu, Jing [Laboratory of Controllable Preparation and Application of Nanomaterials, Technical Institute of Physics and Chemistry, Chinese Academy of Sciences, No. 29, Zhongguancun East Road, Haidian District, Beijing 100190 (China); Tang, Fangqiong, E-mail: tangfq@mail.ipc.ac.cn [Laboratory of Controllable Preparation and Application of Nanomaterials, Technical Institute of Physics and Chemistry, Chinese Academy of Sciences, No. 29, Zhongguancun East Road, Haidian District, Beijing 100190 (China)

2014-01-15

57

Parameters of evaluation of zinc nutritional status: comparison between zinc hair rates and serum alkaline phosphatase in pre-scholars of the Municipality of João Pessoa, Paraíba Parâmetros de avaliação do estado nutricional de zinco: comparação entre os teores de zinco no cabelo e fosfatase alcalina sérica em pré-escolares do município de João Pessoa, Paraíba  

Digital Repository Infrastructure Vision for European Research (DRIVER)

OBJECTIVES: to evaluate the nutritional status of zinc in children from two to five years old, institutio-nalised in daycare centers in the Municipality of João Pessoa, Paraíba. METHODS: a systematic random sample comprised of 377 children. The nutritional status of zinc was evaluated according to the following parameters: mineral concentrations in the hair and enzymatic activity levels of serum alkaline phosphatase. RESULTS: inadequate zinc concentrations prevalence in the hair was of 61,9...

Sandra Cristina da Silva-Santana; Alcides da Silva Diniz; Margarida Maria de Feitas Lóla; Rejane Santana Oliveira; Solange Maria Miranda Silva; Severino Francisco de Oliveira; Patrick Kolsteren

2002-01-01

58

Diagnostic Utility of Heat Stable Alkaline Phosphatase in Hypertensive Disorders of Pregnancy  

Science.gov (United States)

Background: Hypertensive disorders in pregnancy (HDP) complicate 3-10% of all pregnancies. Though there are several biochemical parameters which aid in predicting hypertension of pregnancy, human placental alkaline phosphatase (PLAP), synthesized in placenta during pregnancy by placental syncytiotrophoblast, assumes diagnostic relevance. The purpose of this study was to compare the total alkaline phosphatase (ALP) and heat stable placental alkaline phosphatase (PLAP) levels in the serum of normotensive and hypertensive disorders of pregnancy and to evaluate the clinical utility of ALP and PLAP as a reliable, sensitive, specific and economical biochemical marker of HDP. Materials and Methods: This was a case control study, carried out on pregnant women with hypertension, of south Indian population. Study included pregnant women, 60 patients with hypertension and 60 controls. Biochemical assays were carried out by the IFCC approved procedures based on spectrophotometric method and using fully automated random access chemistry analyser. Data was compared by using student t-test. ROC was drawn to find out optimum cut off for ALP, PLAP and PLAP/ALP ratio in HDP. Pearson’s correlation was performed to ascertain the association among markers. Results: Serum total ALP, PLAP and PLAP/ALP ratio levels were significantly higher in hypertensive pregnant women when compared to controls (phypertension in pregnancy. Conclusion: Serum heat stable ALP isoenzyme and PLAP/ALP ratio could be useful adjuvant markers in diagnosis of HDP in association with other relevant and economically viable biochemical tests. PMID:25584211

Abu Raghavan, Srinivasan; Ghosh, Seethesh; Basu, Sharbari; Ramasamy, Ramesh; Murugaiyan, Sathish Babu

2014-01-01

59

Placental alkaline phosphatase as a tumour marker in seminoma using the H17 E2 monoclonal antibody assay.  

Digital Repository Infrastructure Vision for European Research (DRIVER)

Serum samples from 62 patients with seminoma were assayed for placental alkaline phosphatase-like activity using the monoclonal antibody H17 E2, in order to evaluate its utility as a serum tumour marker. Fifteen of 16 patients (94%) with active seminoma had elevated serum PLAP levels. Sixteen of 46 (35%) of patients considered to be in remission had elevated PLAP levels (false positive rate 35%). Fifteen false positive results were considered attributable to concomitant smoking, and if these ...

Horwich, A.; Tucker, D. F.; Peckham, M. J.

1985-01-01

60

As fosfatases alcalinas, transaminases e gama-glutamil-transferase séricas em pacientes epilépticos tratados com carbamazepina The serum alkaline phosphatases, transaminases, and gamma-glutamil transferases in epileptic patients treated with carbamazepine  

Directory of Open Access Journals (Sweden)

Full Text Available INTRODUÇÃO: A carbamazepina é a droga utilizada no tratamento de pacientes com epilepsia parcial (ou focal secundariamente generalizada. Apesar do uso terapêutico, este fármaco tem sido implicado no aumento das atividades séricas de algumas enzimas. Alguns autores descreveram valores de prevalência de 7,7%, 13% e 22% para aumento de atividade das fosfatases alcalinas séricas (FA ou EC 3.1.3.1. A divergência de resultados também foi encontrada para as atividades da gama-glutamil-transferase sérica (gama-glutamil transferase ou GGT ou EC 2.3.2.2. OBJETIVO: Assim, a meta desta pesquisa é determinar, dentre outros objetivos, a freqüência de alterações nas atividades das FA, GGT e transaminases (AST, aspartato-amino-transferase, EC 2.6.1.1; e ALT, alanina-amino-transferase, EC 2.6.1.2 de uma amostra de pacientes do ambulatório de epilepsia em Salvador, Bahia. MATERIAL E MÉTODOS: O desenho do estudo é descritivo do tipo série de casos, aprovado pelo Comitê de Ética local, no qual uma amostra de conveniência de 52 pacientes epilépticos de acompanhamento ambulatorial foi obtida sem interferência dos pesquisadores. Estes pacientes foram organizados por faixa etária de 12 a 30 e de 31 a 90 anos e, subdivididos por tempo de monoterapia com carbamazepina. As atividades séricas das enzimas GGT, FA, AST e ALT foram determinadas. RESULTADOS: As proporções de alterações por variáveis foram descritas: 42% para as FA, 18% para as GGT, 2% para as ALT e 12% para as AST, respectivamente. A faixa etária de 12 a 30 anos apresentou 56% de alterações nas FA enquanto que aquela de 31 a 90 anos, apenas 18%. CONCLUSÃO: Nós concluímos que as enzimas FA, GGT, AST e ALT apresentaram maiores freqüências de alterações de suas atividades naqueles pacientes com idade igual ou inferior a 30 anos, sendo que as FA apresentaram maiores valores.INTRODUCTION: Carbamazepine is the drug of choice used in the treatment of patients with partial (or focal epilepsy with secondary generalization. Despite its therapeutical use, this drug has been implicated in the increase of serum activities in some enzymes. Some authors have described prevalence values of 7.7%, 13%, and 22% for the increase of activity of serum alkaline phosphatases (AF or EC 3.1.3.1. A divergence in the results was also found for the activities of the serum g-glutamil transferase (gamma-glutamil transferase or GGT, or EC 2.3.2.2. OBJECTIVE: Hence, among other objectives, the aim of this research is to determine the frequency of sample alterations in serum enzymatic activities of AF, GGT and transaminases (AST, aspartate amino-transferase, EC 2.6.1.1; and ALT, alanine-amino-transferase, EC 2.6.1.2 in epilepsy ward patients in Salvador, Bahia. MATERIAL AND METHODS: The design of the study is descriptive and it is a case series type. It has been approved by the local Ethics Committee. In this study, a convenience sample of 52 epileptic patients who receive ambulatory care was obtained without interference by the researchers. These patients were divided according to age groups of 12 to 30 years and 31 to 90 years, which were then subdivided according to the period of monotherapy with carbamazepine. The serum activities of the enzymes GGT, AF, AST and ALT were determined. RESULTS: The ratios of alterations per variables were described: 42% for the FA, 18% for the GGT, 2% for the ALT, and 12% for the AST respectively. The age group of 12 to 30 years presented 56% of alterations in the AF while the group of 31 to 90 years presented only 18% of alterations. CONCLUSION: We conclude that the enzymes AF, GGT, AST, and ALT presented higher frequencies of alterations of their activities in those patients with age equal to or below 30 years, while the AF presented higher values.

Helder Jacobina Santos

2006-03-01

 
 
 
 
61

Parameters of evaluation of zinc nutritional status: comparison between zinc hair rates and serum alkaline phosphatase in pre-scholars of the Municipality of João Pessoa, Paraíba / Parâmetros de avaliação do estado nutricional de zinco: comparação entre os teores de zinco no cabelo e fosfatase alcalina sérica em pré-escolares do município de João Pessoa, Paraíba  

Scientific Electronic Library Online (English)

Full Text Available SciELO Brazil | Language: English Abstract in portuguese OBJETIVOS: avaliar o estado nutricional de zinco em crianças de dois a cinco de idade, institucionalizadas em creches do município de João Pessoa, Paraíba. MÉTODOS: Utilizou-se uma amostra aleatória sistemática, constituída de 377 crianças. O status nutricional de zinco foi avaliado pelos seguintes [...] parâmetros: concentrações do mineral no cabelo e níveis de atividade enzimática da fosfatase alcalina sérica. RESULTADOS: a prevalência das concentrações inadequadas de zinco no cabelo foi de 61,9% predominando a deficiência na sua forma grave, 38,5%. A prevalência das concentrações inadequadas de níveis de atividade enzimática da fosfatase alcalina sérica foi de 40,1%. As concentrações de zinco no cabelo não mostraram correlação com a fosfatase alcalina sérica (r = 0,01; p = 0,882). A distribuição do zinco no cabelo segundo sexo e idade foi homogênea; comportamento idêntico foi observado com a fosfatase alcalina. CONCLUSÕES: a deficiência de zinco mostrou-se extremamente elevada no município de João Pessoa. Abstract in english OBJECTIVES: to evaluate the nutritional status of zinc in children from two to five years old, institutio-nalised in daycare centers in the Municipality of João Pessoa, Paraíba. METHODS: a systematic random sample comprised of 377 children. The nutritional status of zinc was evaluated according to t [...] he following parameters: mineral concentrations in the hair and enzymatic activity levels of serum alkaline phosphatase. RESULTS: inadequate zinc concentrations prevalence in the hair was of 61,9% with predominance in the form of severe deficiency, 38,5%. Prevalence of inadequate concentrations of levels of serum alkaline phosphatase activity was 40,1%. Zinc concentrations in the hair did not correlatated with serum alkaline phosphatase (r = 0,01; p = 0,882). According to sex and age zinc distribution in the hair was homogenous. The alkaline phosphaase had identical behavior. CONCLUSIONS: zinc deficiency is extremely high in the municipality of João Pessoa.

Sandra Cristina da, Silva-Santana; Alcides da Silva, Diniz; Margarida Maria de Feitas, Lóla; Rejane Santana de, Oliveira; Solange Maria Miranda, Silva; Severino Francisco de, Oliveira; Patrick, Kolsteren.

2002-12-01

62

Parameters of evaluation of zinc nutritional status: comparison between zinc hair rates and serum alkaline phosphatase in pre-scholars of the Municipality of João Pessoa, Paraíba Parâmetros de avaliação do estado nutricional de zinco: comparação entre os teores de zinco no cabelo e fosfatase alcalina sérica em pré-escolares do município de João Pessoa, Paraíba  

Directory of Open Access Journals (Sweden)

Full Text Available OBJECTIVES: to evaluate the nutritional status of zinc in children from two to five years old, institutio-nalised in daycare centers in the Municipality of João Pessoa, Paraíba. METHODS: a systematic random sample comprised of 377 children. The nutritional status of zinc was evaluated according to the following parameters: mineral concentrations in the hair and enzymatic activity levels of serum alkaline phosphatase. RESULTS: inadequate zinc concentrations prevalence in the hair was of 61,9% with predominance in the form of severe deficiency, 38,5%. Prevalence of inadequate concentrations of levels of serum alkaline phosphatase activity was 40,1%. Zinc concentrations in the hair did not correlatated with serum alkaline phosphatase (r = 0,01; p = 0,882. According to sex and age zinc distribution in the hair was homogenous. The alkaline phosphaase had identical behavior. CONCLUSIONS: zinc deficiency is extremely high in the municipality of João Pessoa.OBJETIVOS: avaliar o estado nutricional de zinco em crianças de dois a cinco de idade, institucionalizadas em creches do município de João Pessoa, Paraíba. MÉTODOS: Utilizou-se uma amostra aleatória sistemática, constituída de 377 crianças. O status nutricional de zinco foi avaliado pelos seguintes parâmetros: concentrações do mineral no cabelo e níveis de atividade enzimática da fosfatase alcalina sérica. RESULTADOS: a prevalência das concentrações inadequadas de zinco no cabelo foi de 61,9% predominando a deficiência na sua forma grave, 38,5%. A prevalência das concentrações inadequadas de níveis de atividade enzimática da fosfatase alcalina sérica foi de 40,1%. As concentrações de zinco no cabelo não mostraram correlação com a fosfatase alcalina sérica (r = 0,01; p = 0,882. A distribuição do zinco no cabelo segundo sexo e idade foi homogênea; comportamento idêntico foi observado com a fosfatase alcalina. CONCLUSÕES: a deficiência de zinco mostrou-se extremamente elevada no município de João Pessoa.

Sandra Cristina da Silva-Santana

2002-12-01

63

Asparagine-linked carbohydrate does not determine the cellular location of yeast vacuolar nonspecific alkaline phosphatase.  

Digital Repository Infrastructure Vision for European Research (DRIVER)

The nonspecific alkaline phosphatase of Saccharomyces sp. strain 1710 has been shown by phosphatase cytochemistry to be exclusively located in the vacuole, para-Nitrophenyl phosphate-specific alkaline phosphatase is not detected by this procedure because the activity of this enzyme is sensitive to the fixative agent, glutaraldehyde. To determine whether the oligosaccharide of nonspecific alkaline phosphatase is necessary to transport the enzyme into the vacuole, protoplasts were derepressed i...

Clark, D. W.; Tkacz, J. S.; Lampen, J. O.

1982-01-01

64

Alkaline phosphatase prevents platelet stimulation by thromboxane-mimetics.  

Digital Repository Infrastructure Vision for European Research (DRIVER)

1. The effects of alkaline phosphatase on platelet aggregation, secretion and thromboxane B2 (TxB2) generation induced by the full dose-range of common platelet agonists were studied in human platelet-rich plasma and washed platelets. 2. Platelet aggregation and adenosine 5'-triphosphate (ATP) secretion induced by threshold and supramaximal concentrations of arachidonate and stable TxA2 and prostaglandin endoperoxide-mimetics (compounds U46619 and EP171) were abolished in the presence of alka...

Hatmi, M.; Haye, B.; Gavaret, J. M.; Vargaftig, B. B.; Jacquemin, C.

1991-01-01

65

Interplay between intestinal alkaline phosphatase, diet, gut microbes and immunity  

Digital Repository Infrastructure Vision for European Research (DRIVER)

Intestinal alkaline phosphatase (IAP) plays an essential role in intestinal homeostasis and health through interactions with the resident microbiota, diet and the gut. IAP’s role in the intestine is to dephosphorylate toxic microbial ligands such as lipopolysaccharides, unmethylated cytosine-guanosine dinucleotides and flagellin as well as extracellular nucleotides such as uridine diphosphate. IAP’s ability to detoxify these ligands is essential in protecting the host from sepsis during a...

Estaki, Mehrbod; Decoffe, Daniella; Gibson, Deanna L.

2014-01-01

66

Kinetic determination of alkaline phosphatase activity based on hydrolytic cleavage of the P-F bond in monofluorophosphate and fluoride ion-selective electrode.  

Science.gov (United States)

Alkaline phosphatase catalyzes the hydrolytic cleavage of the P-F bond in monofluorophosphate with the subsequent release of fluoride ions. A kinetic potentiometric method is described in which a fluoride ion-selective electrode is used for the sensitive and selective measurement of the released F- for the determination of alkaline phosphatase activity. It is shown that monofluorophosphate can be used as an alternative substrate for alkaline phosphatase. The reaction demonstrates a well-defined correlation with the hydrolysis of the P-O bond in 4-nitrophenyl phosphate. The serum alkaline phosphatase was determined in human serum samples by the potentiometric technique, and the results obtained compared well with a standard spectrophotometric method. PMID:2077934

Venetz, W P; Mangan, C; Siddiqi, I W

1990-11-15

67

ALKALINE PHOSPHATASE ACTIVITY AS A MARKER OF DOG SEMEN FREEZABILITY  

Directory of Open Access Journals (Sweden)

Full Text Available The investigation was performed to evaluate the dog semen freezability and itsquality after thawing allowing its use for artificial insemination (AI. On the basis ofsperm motility, concentration and alkaline phosphatase (AP activity in semenplasma it was possible to establish that AP activity corresponds with the basic factorof semen examination. Significant statistical differences occurred between thequality of ejaculates which were qualified or disqualified to deep freezing and AI.These results show that AP activity in raw dog semen plasma can be used as amarker for the dog semen qualification for deep freezing and AI with 95%probability of the prognosis of the results.

K. KOSINIAK-KAMYSZ

2013-12-01

68

ALKALINE PHOSPHATASE ACTIVITY AS A MARKER OF DOG SEMEN FREEZABILITY  

Directory of Open Access Journals (Sweden)

Full Text Available The investigation was performed to evaluate the dog semen freezability and itsquality after thawing allowing its use for artificial insemination (AI. On the basis ofsperm motility, concentration and alkaline phosphatase (AP activity in semenplasma it was possible to establish that AP activity corresponds with the basic factorof semen examination. Significant statistical differences occurred between thequality of ejaculates which were qualified or disqualified to deep freezing and AI.These results show that AP activity in raw dog semen plasma can be used as amarker for the dog semen qualification for deep freezing and AI with 95%probability of the prognosis of the results.

KOSINIAK-KAMYSZ K.

2007-01-01

69

As fosfatases alcalinas, transaminases e gama-glutamil-transferase séricas em pacientes epilépticos tratados com carbamazepina / The serum alkaline phosphatases, transaminases, and gamma-glutamil transferases in epileptic patients treated with carbamazepine  

Scientific Electronic Library Online (English)

Full Text Available SciELO Brazil | Language: Portuguese Abstract in portuguese INTRODUÇÃO: A carbamazepina é a droga utilizada no tratamento de pacientes com epilepsia parcial (ou focal) secundariamente generalizada. Apesar do uso terapêutico, este fármaco tem sido implicado no aumento das atividades séricas de algumas enzimas. Alguns autores descreveram valores de prevalência [...] de 7,7%, 13% e 22% para aumento de atividade das fosfatases alcalinas séricas (FA ou EC 3.1.3.1). A divergência de resultados também foi encontrada para as atividades da gama-glutamil-transferase sérica (gama-glutamil transferase ou GGT ou EC 2.3.2.2). OBJETIVO: Assim, a meta desta pesquisa é determinar, dentre outros objetivos, a freqüência de alterações nas atividades das FA, GGT e transaminases (AST, aspartato-amino-transferase, EC 2.6.1.1; e ALT, alanina-amino-transferase, EC 2.6.1.2) de uma amostra de pacientes do ambulatório de epilepsia em Salvador, Bahia. MATERIAL E MÉTODOS: O desenho do estudo é descritivo do tipo série de casos, aprovado pelo Comitê de Ética local, no qual uma amostra de conveniência de 52 pacientes epilépticos de acompanhamento ambulatorial foi obtida sem interferência dos pesquisadores. Estes pacientes foram organizados por faixa etária de 12 a 30 e de 31 a 90 anos e, subdivididos por tempo de monoterapia com carbamazepina. As atividades séricas das enzimas GGT, FA, AST e ALT foram determinadas. RESULTADOS: As proporções de alterações por variáveis foram descritas: 42% para as FA, 18% para as GGT, 2% para as ALT e 12% para as AST, respectivamente. A faixa etária de 12 a 30 anos apresentou 56% de alterações nas FA enquanto que aquela de 31 a 90 anos, apenas 18%. CONCLUSÃO: Nós concluímos que as enzimas FA, GGT, AST e ALT apresentaram maiores freqüências de alterações de suas atividades naqueles pacientes com idade igual ou inferior a 30 anos, sendo que as FA apresentaram maiores valores. Abstract in english INTRODUCTION: Carbamazepine is the drug of choice used in the treatment of patients with partial (or focal) epilepsy with secondary generalization. Despite its therapeutical use, this drug has been implicated in the increase of serum activities in some enzymes. Some authors have described prevalence [...] values of 7.7%, 13%, and 22% for the increase of activity of serum alkaline phosphatases (AF or EC 3.1.3.1). A divergence in the results was also found for the activities of the serum g-glutamil transferase (gamma-glutamil transferase or GGT, or EC 2.3.2.2). OBJECTIVE: Hence, among other objectives, the aim of this research is to determine the frequency of sample alterations in serum enzymatic activities of AF, GGT and transaminases (AST, aspartate amino-transferase, EC 2.6.1.1; and ALT, alanine-amino-transferase, EC 2.6.1.2) in epilepsy ward patients in Salvador, Bahia. MATERIAL AND METHODS: The design of the study is descriptive and it is a case series type. It has been approved by the local Ethics Committee. In this study, a convenience sample of 52 epileptic patients who receive ambulatory care was obtained without interference by the researchers. These patients were divided according to age groups of 12 to 30 years and 31 to 90 years, which were then subdivided according to the period of monotherapy with carbamazepine. The serum activities of the enzymes GGT, AF, AST and ALT were determined. RESULTS: The ratios of alterations per variables were described: 42% for the FA, 18% for the GGT, 2% for the ALT, and 12% for the AST respectively. The age group of 12 to 30 years presented 56% of alterations in the AF while the group of 31 to 90 years presented only 18% of alterations. CONCLUSION: We conclude that the enzymes AF, GGT, AST, and ALT presented higher frequencies of alterations of their activities in those patients with age equal to or below 30 years, while the AF presented higher values.

Helder Jacobina, Santos; Antonio de Souza, Andrade Filho; Olivia Lordelo, Sanches; Tiago, Spolador; Luís Erlon Araújo, Rodrigues.

2006-03-01

70

Excretion of alkaline phosphatase by Escherichia coli K-12 pho constitutive mutants transformed with plasmids carrying the alkaline phosphatase structural gene.  

Digital Repository Infrastructure Vision for European Research (DRIVER)

Escherichia coli alkaline phosphatase constitutive mutants carrying a pst or a phoS mutation and a plasmid-bearing gene phoA+ excreted into the growth medium up to 50% of the total alkaline phosphatase production. This excretion was pH dependent and did not involve drastic modifications of the cell envelope. Alkaline phosphatase accounted for 80% of total released proteins. Amplification of gene phoA+ was a necessary condition for excretion to occur. When the beta-lactamase structural gene bl...

Lazzaroni, J. C.; Atlan, D.; Portalier, R. C.

1985-01-01

71

Study on alkaline and acid phosphatase activity in acute uranium intoxication  

International Nuclear Information System (INIS)

The protective potential of diethyl barbituric acid sodium salt is studied, in comparison with that of acetazolamide, on kidneys under acute uranium intoxication. Experiments involved rats given intraperitoneal injections with uranyl acetate on 12 successive days up to a total dose of 0.5, 2.0 or 7.0 mg/kg. The resulting effects are measured by chemical assays of serum and urine for alkaline and acid phosphatase and histochemical assays for phosphatase activities in kidneys, kinetics being followed over a 30-day period after total dose administration. Protection of kidneys from toxic uranium effects was found to be of about the same degree with sodium diethyl barbiturate as with acetazolamide. (A.B.)

72

Improved double immunohistochemical staining method for cryostat and paraffin wax sections, combining alkaline phosphatase anti-alkaline phosphatase and indirect immunofluorescence.  

Digital Repository Infrastructure Vision for European Research (DRIVER)

Aims - To develop an immunohistochemical staining method for cryostat and paraffin wax sections so that two different antigens in the same section of tissues could be detected by combining immunoenzyme and immunofluorescence techniques. Methods - This double immunohistochemical staining method combines alkaline phosphatase-anti-alkaline phosphatase (APAAP) using New Fuchsin as a chromogen and indirect immunofluorescence. Results - APAAP staining for one antigen of this double immunohistochemi...

Tao, Q.; Srivastava, G.; Loke, S. L.; Chan, E. Y.; Ho, F. C.

1994-01-01

73

Outer membrane protein e of Escherichia coli K-12 is co-regulated with alkaline phosphatase.  

Digital Repository Infrastructure Vision for European Research (DRIVER)

Outer membrane protein e is induced in wild-type cells, just like alkaline phosphatase and some other periplasmic proteins, by growth under phosphatase limitation. nmpA and nmpB mutants, which synthesize protein e constitutively, are shown also to produce the periplasmic enzyme alkaline phosphatase constitutively. Alternatively, individual phoS, phoT, and phoR mutants as well as pit pst double mutants, all of which are known to produce alkaline phosphatase constitutively, were found to be con...

Tommassen, J.; Lugtenberg, B.

1980-01-01

74

Intestinal alkaline phosphatase prevents metabolic syndrome in mice  

Science.gov (United States)

Metabolic syndrome comprises a cluster of related disorders that includes obesity, glucose intolerance, insulin resistance, dyslipidemia, and fatty liver. Recently, gut-derived chronic endotoxemia has been identified as a primary mediator for triggering the low-grade inflammation responsible for the development of metabolic syndrome. In the present study we examined the role of the small intestinal brush-border enzyme, intestinal alkaline phosphatase (IAP), in preventing a high-fat-diet–induced metabolic syndrome in mice. We found that both endogenous and orally supplemented IAP inhibits absorption of endotoxin (lipopolysaccharides) that occurs with dietary fat, and oral IAP supplementation prevents as well as reverses metabolic syndrome. Furthermore, IAP supplementation improves the lipid profile in mice fed a standard, low-fat chow diet. These results point to a potentially unique therapy against metabolic syndrome in at-risk humans. PMID:23569246

Kaliannan, Kanakaraju; Hamarneh, Sulaiman R.; Economopoulos, Konstantinos P.; Nasrin Alam, Sayeda; Moaven, Omeed; Patel, Palak; Malo, Nondita S.; Ray, Madhury; Abtahi, Seyed M.; Muhammad, Nur; Raychowdhury, Atri; Teshager, Abeba; Mohamed, Mussa M. Rafat; Moss, Angela K.; Ahmed, Rizwan; Hakimian, Shahrad; Narisawa, Sonoko; Millán, José Luis; Hohmann, Elizabeth; Warren, H. Shaw; Bhan, Atul K.; Malo, Madhu S.; Hodin, Richard A.

2013-01-01

75

Interfering effect of bilirubin on the determination of alkaline phosphatase  

Science.gov (United States)

Objective: This study was designed to evaluate whether the high concentration of bilirubin is able to interfere the determination of alkaline phosphatase (ALP). Methods: Clinical tests evaluating the interfering substance of bilirubin of various concentrations on the determination of ALP were conducted based upon the Clinical and Laboratory Standards Institute (CLSI) document EP7-A2, the most recent guideline on interference testing approved in 2005. Results: Paired t-test comparing different doses of bilirubin revealed that the concentration of 1 000 ?mmol/L bilirubin negatively interfered the determination of ALP levels. The experiment designed with five different concentrations of bilirubin showed that bilirubin can exert negative interference on the measurement of ALP in a linear pattern. Conclusion: High concentration of bilirubin can cause false measurement of ALP levels, probably interfering with the clinical prognosis of liver diseases. PMID:25550938

Wang, Zhong; Guo, Hongyan; Wang, Ying; Kong, Fanjun; Wang, Rui

2014-01-01

76

Activity of alkaline phosphatase adsorbed and grafted on "polydopamine" films.  

Science.gov (United States)

The oxidation of dopamine in slightly basic solutions and in the presence of oxygen as an oxidant allows for the deposition of dopamine-eumelanin ("polydopamine") films on almost all kinds of materials allowing for an easy secondary functionalization. Molecules carrying nucleophilic groups like thiols and amines can be easily grafted on those films. Herein we show that alkaline phosphatase (ALP), as a model enzyme, adsorbs to "polydopamine" films and part of the adsorbed enzyme is rapidly desorbed in contact with Tris buffer. However a significant part of the enzyme remains irreversibly adsorbed and keeps some enzymatic activity for at least 2 weeks whereas ALP adsorbed on quartz slides is rapidly and quantitatively deactivated. In addition we estimated the Michaelis constant Km of the enzyme irreversibly bound to the "polydopamine" film. The Michaelis constant, and hence the affinity constant between paranitrophenol phosphate and ALP are almost identical between the enzyme bound on the film and the free enzyme in solution. Complementarily, it was found that "polydopamine" films display some phosphatase like catalytic activity. PMID:24935183

Ball, Vincent

2014-09-01

77

Histochemical imaging of alkaline phosphatase using a novel fluorescent substrate.  

Science.gov (United States)

Histochemical visualization of phosphatase is exclusively required for Western immunoblotting and antigen-positive cell staining using an alkaline phosphatase (AP)-labeled secondary antibody. This detection has been performed by several reagents including 5-bromo-4-chloro-3-indolyl-phosphate (X-Phos), nitro blue tetrazolium (NBT), 3-(2'-spiroadamantane)-4-methoxy-4-(3?-phosphoryloxy)phenyl-1,2-dioxetane and 2-(5'-chloro-2'-phosphoryloxyphenyl)-6-chloro-4-[3H]-quinazolinone (ELF® 97 Phosphate). We previously reported that 2-(benzothiazol-2-yl)-4-bromophenol bonded with N-acetylneuraminic acid (BTP3-Neu5Ac), enabled fluorescent histochemical visualization of sialidase activity. 2-(Benzothiazol-2-yl)-4-bromophenol (BTP3), which is formed from BTP3-Neu5Ac by sialidase reaction, is a crystalline, insoluble and stable fluorogenic compound, deposited at the site of enzyme activity. We developed a BTP3 phosphate ester (BTP3-Phos) for the purpose of fluorescent histochemical visualization of phosphatase activity. BTP3-Phos emitted fluorescence in a manner dependent on the concentration of the AP-labeled antibody. BTP3-Phos also enabled fluorescent histochemical visualization of AP-blotted dots in a manner dependent on the concentration of the AP-labeled antibody. The detection sensitivity of BTP3-Phos was estimated to be greater than that of the conventional method using X-Phos and NBT. Influenza A virus-infected cells were fixed and reacted with anti-influenza A virus antibodies and incubated continuously with an AP-labeled secondary antibody. BTP3-Phos stained the infected cells with distinct green fluorescence. These results indicate that BTP3-Phos can enable fluorescent immunohistochemical staining analysis using an AP-labeled antibody. BTP3-Phos would be beneficial for histochemical staining of AP activity, and may be applicable for multi-color staining or a cell sorter. PMID:25109307

Takahashi, Tadanobu; Otsubo, Tadamune; Ikeda, Kiyoshi; Minami, Akira; Suzuki, Takashi

2014-01-01

78

Interplay between intestinal alkaline phosphatase, diet, gut microbes and immunity.  

Science.gov (United States)

Intestinal alkaline phosphatase (IAP) plays an essential role in intestinal homeostasis and health through interactions with the resident microbiota, diet and the gut. IAP's role in the intestine is to dephosphorylate toxic microbial ligands such as lipopolysaccharides, unmethylated cytosine-guanosine dinucleotides and flagellin as well as extracellular nucleotides such as uridine diphosphate. IAP's ability to detoxify these ligands is essential in protecting the host from sepsis during acute inflammation and chronic inflammatory conditions such as inflammatory bowel disease. Also important in these complications is IAP's ability to regulate the microbial ecosystem by forming a complex relationship between microbiota, diet and the intestinal mucosal surface. Evidence reveals that diet alters IAP expression and activity and this in turn can influence the gut microbiota and homeostasis. IAP's ability to maintain a healthy gastrointestinal tract has accelerated research on its potential use as a therapeutic agent against a multitude of diseases. Exogenous IAP has been shown to have beneficial effects when administered during ulcerative colitis, coronary bypass surgery and sepsis. There are currently a handful of human clinical trials underway investigating the effects of exogenous IAP during sepsis, rheumatoid arthritis and heart surgery. In light of these findings IAP has been marked as a novel agent to help treat a variety of other inflammatory and infectious diseases. The purpose of this review is to highlight the essential characteristics of IAP in protection and maintenance of intestinal homeostasis while addressing the intricate interplay between IAP, diet, microbiota and the intestinal epithelium. PMID:25400448

Estaki, Mehrbod; DeCoffe, Daniella; Gibson, Deanna L

2014-11-14

79

Affinity methods with lectins: a tool to identify canine alkaline phosphatase isoenzymes.  

Science.gov (United States)

Affinity methods were used to characterize selective interactions of alkaline phosphatase (ALP) isoenzymes from different dog tissues with lectins. Specific lectins were used to identify liver, intestinal and steroid-induced ALP isoenzymes in serum from dogs with Cushing syndrome or steroid-treated dogs. For the first approach, 12 lectins were assayed by affinity dots. Selective interactions were found among wheat germ agglutinin (WGA), jacalin, con A (concanavalin A) and Helix pomatia agglutinin (HPA) and several ALP-containing samples. These four reactive lectins were assayed by line electrophoresis with lectins in holes. A strong reactivity of con A with all isoenzymes was found, although the patterns were different. WGA interacted with intestinal, bone marrow extracts and Cushing syndrome serum. Jacalin changed the electrophoretic patterns of intestinal and liver ALP, and Cushing serum. Finally, by crossed electrophoresis with lectins in gels, it was possible to distinguish among hepatic or intestinal ALPs and the steroid-induced isoenzyme in serum. Affinity electrophoresis with lectins provided a clear separation and identification of the different dog ALP isoenzymes. PMID:8258637

Calderón de la Barca, A M; Jensen, A L; Bøg-Hansen, T C

1993-10-01

80

Subcellular localization of alkaline phosphatase in Bacillus licheniformis 749/C by immunoelectron microscopy with colloidal gold  

International Nuclear Information System (INIS)

Subcellular distribution of the alkaline phosphatase of Bacillus licheniformis 749/C was determined by an immunoelectron microscopy method. Anti-alkaline phosphatase antibody labeled with 15- to 18-nm colloidal gold particles (gold-immunoglobulin G [IgG] complex) were used for the study. Both the plasma membrane and cytoplasmic material were labeled with the gold-IgG particles. These particles formed clusters in association with the plasma membrane; in contrast, in the cytoplasm the particles were largely dispersed, and only a few clusters were found. The gold-IgG binding was quantitatively estimated by stereological analysis of labeled, frozen thin sections. This estimation of a variety of control samples showed that the labeling was specific for the alkaline phosphatase. Cluster formation of the gold -IgG particles in association with the plasma membrane suggests that existence of specific alkaline phosphatase binding sites (receptors) in the plasma membrane of B. licheniformis 749/C. 27 references, 6 figures, 1 table

 
 
 
 
81

Characterization of the phosphatidylinositol-glycan membrane anchor of human placental alkaline phosphatase  

International Nuclear Information System (INIS)

Placental alkaline phosphatase [orthophosphoric-monoester phosphohydrolase (alkaline optimum), EC 3.1.3.1] is a member of a diverse group of membrane proteins whose attachment to the lipid bilayer is mediated by a phosphatidylinositol-glycan. To investigate structural aspects of the glycolipid anchor, cultured WISH cells were used because, they produce the enzyme in abundant quantities. When cell suspensions were incubated with purified phosphatidylinositol-specific phospholipase C, most of the placental alkaline phosphatase was released from membranes in a hydrophilic form. On incubation of the cells with [14C]ethanolamine, [14C]myristic acid, or myo[3H]inositol, each was incorporated into the phosphatase near the carboxyl terminus, showing that these components, which are found in other phosphatidylinositol membrane-linked proteins, are also present in placental alkaline phosphatase

82

New pleiotropic alkaline phosphatase-negative mutants of Escherichia coli K-12.  

Digital Repository Infrastructure Vision for European Research (DRIVER)

Escherichia coli K-12 mutants showing reduced alkaline phosphatase activity were isolated as 5-fluorouracil-plus-adenosine-resistant derivatives of a upp pho (either phoS or phoT) strain. One class of these mutants displayed a temperature-sensitive alkaline phosphatase-negative phenotype, a pleiotropic defect for growth on some substrates, an increased sensitivity to toxic compounds (e.g., EDTA, mitomycin, and chloramphenicol), and alterations in the expression of some membrane proteins. It p...

Heyde, M.; Portalier, R.

1982-01-01

83

Expression and characterization of recombinant thermostable alkaline phosphatase from a novel thermophilic bacterium Thermus thermophilus XM.  

Science.gov (United States)

A gene (tap) encoding a thermostable alkaline phosphatase from the thermophilic bacterium Thermus thermophilus XM was cloned and sequenced. It is 1506 bp long and encodes a protein of 501 amino acid residues with a calculated molecular mass of 54.7 kDa. Comparison of the deduced amino acid sequence with other alkaline phosphatases showed that the regions in the vicinity of the phosphorylation site and metal binding sites are highly conserved. The recombinant thermostable alkaline phosphatase was expressed as a His6-tagged fusion protein in Escherichia coli and its enzymatic properties were characterized after purification. The pH and temperature optima for the recombinant thermostable alkaline phosphatases activity were pH 12 and 75 degrees C. As expected, the enzyme displayed high thermostability, retaining more than 50% activity after incubating for 6 h at 80 degrees C. Its catalytic function was accelerated in the presence of 0.1 mM Co2+, Fe2+, Mg2+, or Mn2+ but was strongly inhibited by 2.0 mM Fe2+. Under optimal conditions, the Michaelis constant (K(m)) for cleavage of p-nitrophenyl-phosphate was 0.034 mM. Although it has much in common with other alkaline phosphatases, the recombinant thermostable alkaline phosphatase possesses some unique features, such as high optimal pH and good thermostability. PMID:17989875

Li, Jianbo; Xu, Limei; Yang, Feng

2007-11-01

84

Intestinal alkaline phosphatase inhibits the proinflammatory nucleotide uridine diphosphate  

Science.gov (United States)

Uridine diphosphate (UDP) is a proinflammatory nucleotide implicated in inflammatory bowel disease. Intestinal alkaline phosphatase (IAP) is a gut mucosal defense factor capable of inhibiting intestinal inflammation. We used the malachite green assay to show that IAP dephosphorylates UDP. To study the anti-inflammatory effect of IAP, UDP or other proinflammatory ligands (LPS, flagellin, Pam3Cys, or TNF-?) in the presence or absence of IAP were applied to cell cultures, and IL-8 was measured. UDP caused dose-dependent increase in IL-8 release by immune cells and two gut epithelial cell lines, and IAP treatment abrogated IL-8 release. Costimulation with UDP and other inflammatory ligands resulted in a synergistic increase in IL-8 release, which was prevented by IAP treatment. In vivo, UDP in the presence or absence of IAP was instilled into a small intestinal loop model in wild-type and IAP-knockout mice. Luminal contents were applied to cell culture, and cytokine levels were measured in culture supernatant and intestinal tissue. UDP-treated luminal contents induced more inflammation on target cells, with a greater inflammatory response to contents from IAP-KO mice treated with UDP than from WT mice. Additionally, UDP treatment increased TNF-? levels in intestinal tissue of IAP-KO mice, and cotreatment with IAP reduced inflammation to control levels. Taken together, these studies show that IAP prevents inflammation caused by UDP alone and in combination with other ligands, and the anti-inflammatory effect of IAP against UDP persists in mouse small intestine. The benefits of IAP in intestinal disease may be partly due to inhibition of the proinflammatory activity of UDP. PMID:23306083

Hamarneh, Sulaiman R.; Mohamed, Mussa M. Rafat; Ramasamy, Sundaram; Yammine, Halim; Patel, Palak; Kaliannan, Kanakaraju; Alam, Sayeda N.; Muhammad, Nur; Moaven, Omeed; Teshager, Abeba; Malo, Nondita S.; Narisawa, Sonoko; Millán, José Luis; Warren, H. Shaw; Hohmann, Elizabeth; Malo, Madhu S.; Hodin, Richard A.

2013-01-01

85

Studies on dissolved metalloenzymes in lake water. III. Correlation between dissolved alkaline phosphatase and orthophosphate in lake water  

Energy Technology Data Exchange (ETDEWEB)

Alkaline phosphatase activity (both dissolved and total) in Lake Kasumigaura was determined periodically together with concentrations of phosphorus compounds therein. Distinct negative correlation was found between the dissolved alkaline phosphatase activity and the orthophosphate concentration during the phosphorus-limited season. The decrease of the concentration of dissolved alkaline phosphatase was observed at the level of 1 ng ml/sup -1/ of orthophosphate. The present results suggest that both alkaline phosphate in vivo and dissolved alkaline phosphatase in the lake water play important roles in phosphorus cycles in natural water. 29 references, 7 figures, 3 tables.

Kobayashi, K.; Iwase, K.; Hashimoto, S.; Ueda, H.; Fujiwara, K.; Otsuki, A.; Keiichiro, F.; Haraguchi, H.

1987-03-01

86

Radioimmunoassay for prostatic acid phosphatase in human serum. Methodologic aspects  

International Nuclear Information System (INIS)

We propose a double antibody radioimmunoassay for human prostatic acid phosphatase (PAP) in serum for diagnosis and management of prostatic adenocarcinoma under treatment. The antigen is purified from human prostatic fluid by a gel-filtration on Sephadex G 100 followed by affinity chromatography on Con A Sepharose. A specific antibody is raised in rabbits and purified by immunoadsorption with a female serum. The described technique offers both radioisotopic sensibility and immunologic specificity. Physiological values determined in the serum of 125 healthy males are below 2 ng/ml. No significative differences are observed with age. The proposed technique also shows significant differences between values evaluated for benign prostatic hyperplasia and prostatic adenocarcinoma

87

Biochemical localization of alkaline phosphatase in the cell wall of a marine pseudomonad.  

Science.gov (United States)

The various layers of the cell envelope of marine pseudomonad B-16 (ATCC 19855) have been separated from the cells and assayed directly for alkaline phosphatase activity under conditions established previously to be optimum for maintenance of the activity of the enzyme. Under conditions known to lead to the release of the contents of the periplasmic space from the cells, over 90% of the alkaline phosphatase was released into the medium. Neither the loosely bound outer layer nor the outer double-track layer (cell wall membrane) showed significant activity. A small amount of the alkaline phosphatase activity of the cells remained associated with the mureinoplasts when the outer layers of the cell wall were removed. Upon treatment of the mureinoplasts with lysozyme, some alkaline phosphatase was released into the medium and some remained with the protoplasts formed. Cells washed and suspended in 0.5 M NaCl were lysed by treatment with 2% toluene, and 95% of the alkaline phosphatase in the cells was released into the medium. Cells washed and suspended in complete salts solution (0.3 M NaCl, 0.05 M MgSO(4), and 0.01 M KCl) or 0.05 M MgSO(4) appeared intact after treatment with toluene but lost 50 and 10%, respectively, of their alkaline phosphatase. The results suggest that the presence of Mg(2+) in the cell wall is necessary to prevent disruption of the cells by toluene and may also be required to prevent the release of alkaline phosphatase by toluene when disruption of the cells by toluene does not take place. PMID:4811547

Thompson, L M; MacLeod, R A

1974-02-01

88

The fate of purified radio-labelled alkaline phosphatase from the liver in the organism  

International Nuclear Information System (INIS)

Alkaline phosphatase (AP) from dog liver was enriched by a factor of 5.444 in various steps. Rabbit antiserum to the purified AP was produced; 125-I was used then to radiolabel the highly purified AP. Four dogs were cholecystectomized and subsequently received an extracorporal drainage of the bile ducts. Decrease rate of total radio-activity and of PBI in the serum was determined in one dog; likewise in three other dogs before and one week after occlusion of their main bile ducts. In addition, radioactivity above the organs was measured in some animals at short intervals. In the dogs with main bile duct drainage, bile was collected continuously for up to 70 h, samples were taken, and residual bile plus native dog bile were re-infused into the distal choledochus catheter. Total radioactivity, PBI and immunoprecipitability with antibodies were determined in the bile and serum samples. AP, GOT, CPT and bilirubin were determined in some serum samples. In addition, total radioactivity excreted by urine was established. Results show injected 125-I-AP to be rapidly stored in the liver and not to be excreted via bile to a decisive extent. The fact that 125-I-AP is not excreted via bile is further indicated by the identical decrease rate of injected 125-I-AP in the serum in dogs with and without main bile duct occlusion. Injected 125-I-AP appears to be metabolized very rapidly in the liver as is indicated by the rapid decrease of immuno precipitability of 125-I-AP in the serum. (orig./MG)

89

Control of placental alkaline phosphatase gene expression in HeLa cells: induction of synthesis by prednisolone and sodium butyrate  

International Nuclear Information System (INIS)

HeLa S3 cells produce an alkaline phosphatase indistinguishable from the enzyme from human term placenta. The phosphatase activity in these cells was induced by both prednisolone and sodium butyrate. Both agents stimulated de novo synthesis of the enzyme. The increase in phosphatase activity paralleled the increase in immunoactivity and biosynthesis of placental alkaline phosphatase. The fully processed phosphatase monomer in control, prednisolone-treated or butyrate-treated cells was a 64.5 K polypeptide, measured by both incorporation of L-[35S]methionine into enzyme protein and active-site labeling. The 64.5K polypeptide was formed by the incorporation of additional N-acetylneuraminic acid moieties to a precursor polypeptide of 61.5K. However, this biosynthetic pathway was identified only in butyrate-treated cells. In prednisolone-treated cells, the processing of 61.5K to 64.5K monomer was accelerated, and the presence of the 61.5 precursor could only be detected by either neuraminidase or monensin treatment. Phosphatase mRNA which comigrated with the term placental alkaline phosphatase mRNA of 2.7 kilobases was induced in the presence of either prednisolone or butyrate. Alkaline phosphatase mRNA is untreated HeLa S3 cells migrated slightly faster than the term placental alkaline phosphatase mRNA. Butyrate also induced a second still faster migrating alkaline phosphatase mRNA. Both prednisolone and butyrate increased the steady-statone and butyrate increased the steady-state levels of placental alkaline phosphatase mRNA. The data indicate that the increase in phosphatase mRNA by prednisolone and butyrate resulted in the induction of alkaline phosphatase activity and biosynthesis in HeLa S3 cells. Furthermore, both agents induced the expression of different alkaline phosphatase gene transcripts without altering its protein product

90

Control of placental alkaline phosphatase gene expression in HeLa cells: induction of synthesis by prednisolone and sodium butyrate  

Energy Technology Data Exchange (ETDEWEB)

HeLa S/sub 3/ cells produce an alkaline phosphatase indistinguishable from the enzyme from human term placenta. The phosphatase activity in these cells was induced by both prednisolone and sodium butyrate. Both agents stimulated de novo synthesis of the enzyme. The increase in phosphatase activity paralleled the increase in immunoactivity and biosynthesis of placental alkaline phosphatase. The fully processed phosphatase monomer in control, prednisolone-treated or butyrate-treated cells was a 64.5 K polypeptide, measured by both incorporation of L-(/sup 35/S)methionine into enzyme protein and active-site labeling. The 64.5K polypeptide was formed by the incorporation of additional N-acetylneuraminic acid moieties to a precursor polypeptide of 61.5K. However, this biosynthetic pathway was identified only in butyrate-treated cells. In prednisolone-treated cells, the processing of 61.5K to 64.5K monomer was accelerated, and the presence of the 61.5 precursor could only be detected by either neuraminidase or monensin treatment. Phosphatase mRNA which comigrated with the term placental alkaline phosphatase mRNA of 2.7 kilobases was induced in the presence of either prednisolone or butyrate. Alkaline phosphatase mRNA is untreated HeLa S/sub 3/ cells migrated slightly faster than the term placental alkaline phosphatase mRNA. Butyrate also induced a second still faster migrating alkaline phosphatase mRNA. Both prednisolone and butyrate increased the steady-state levels of placental alkaline phosphatase mRNA. The data indicate that the increase in phosphatase mRNA by prednisolone and butyrate resulted in the induction of alkaline phosphatase activity and biosynthesis in HeLa S/sub 3/ cells. Furthermore, both agents induced the expression of different alkaline phosphatase gene transcripts without altering its protein product.

Chou, J.Y.; Takahashi, S.

1987-06-16

91

Alkaline phosphatase activity of water column fractions and seagrass in a tropical carbonate estuary, Florida Bay  

Science.gov (United States)

Few phosphorus-depleted coastal ecosystems have been examined for their ability to hydrolyze phosphomonoesters. We examined seasonal (August 2006-April 2007) alkaline phosphatase activity in Florida Bay, a phosphorus-limited shallow estuary, using fluorescent substrate at low concentrations (?2.0 ?M). In situ dissolved inorganic and organic phosphorus levels and phosphomonoester concentrations were also determined. Water column alkaline phosphatase activity was partitioned into two particulate size fractions (>1.2 and 0.2-1.2 ?m) and freely dissolved enzymes (lysis. Pulses of inorganic phosphorus and labile organic phosphorus and nitrogen may stimulate autotrophs, particularly cyanobacteria, which in turn promote biological activity that increase alkaline phosphatase activity of both autotrophs and heterotrophs in the bay.

Koch, Marguerite S.; Kletou, Demetris C.; Tursi, Rosanna

2009-08-01

92

Intestinal disaccharidase and alkaline phosphatase activities in experimental rabbit mucoid enteropathy.  

Science.gov (United States)

Mucoid enteropathy was induced experimentally by ligation of the cecum, and the activities of mucosal disaccharidases and alkaline phosphatase were measured at different locations along the small intestine of the sick and control rabbits. In the duodenum of rabbits with mucoid enteropathy, the activity of acid beta-galactosidase II was elevated and hetero beta-galactosidase declined. In the jejunum, the activities of lactase, acid beta-galactosidase I and II, hetero beta-galactosidase, trehalase, sucrase and alkaline phosphatase were significantly lower in animals with mucoid enteropathy. In the ileum, acid beta-galactosidase II, hetero beta-galactosidase, maltase, trehalase, sucrase and alkaline phosphatase showed decreased activity in rabbits with mucoid enteropathy. PMID:4094400

Toofanian, F

1985-12-01

93

Activity of Follicular Fluid Phosphatases and Their Correlation with Levels of Serum Esteroidal Hormones and Gonadotropins  

Directory of Open Access Journals (Sweden)

Full Text Available Introduction: The aim of this study was to evaluate the correlation between serum levels of ovarian and gonadotropin hormones, age and number of follicles with follicular alkaline and acid phosphatase activity in infertile women under controlled ovarian hyperstimulation. Methods: After collection of follicular fluid and calculation of the number of follicles, the specific activity of alkaline (ALP and acid phosphatase (ACP was determined according to the total protein in 19 women at the time of puncture. Also at that time, the levels of progesterone, estradiol, and follicle stimulating hormone (FSH and leuteinizing hormone (LH of their sera were measured. The correlation of follicular ALP and ACP with each serum hormone levels, women age and number of follicles was calculated using non-parametric analysis. Results: The ALP has a correlation with progesterone (P=0.01 levels but doesn’t have any correlation with the other factors. However, the ACP activity has a correlation not only with follicular number but also with estradiol and progesterone levels (P=0.05. Conclusion: Thus ACP activity is more affected by ovarian hormone than ALP and it can affect the ovarian microenvironment and oocyte development.

Sh Byranvand

2006-10-01

94

Differentiating Intracellular from Extracellular Alkaline Phosphatase Activity in Soil by Sonication  

Digital Repository Infrastructure Vision for European Research (DRIVER)

Differentiating intracellular from extracellular enzyme activity is important in soil enzymology, but not easy. Here, we report on an adjusted sonication method for the separation of intracellular from extracellular phosphatase activity in soil. Under optimal sonication conditions [soil:water ratio ?=? 1/8 (w/v) and power density ?=? 15 watt ml-1], the activity of alkaline phosphomonoesterase (phosphatase) in a Haplic Cambisol soil increased with sonication time in two distinct steps....

Qin, S. P.; Hu, C. S.; Oenema, O.

2013-01-01

95

AVERRHOA CARAMBOLA (STAR FRUIT INDUCES HEPATIC ALKALINE PHOSPHATASE ACTIVITY IN RATS  

Directory of Open Access Journals (Sweden)

Full Text Available The objective of the study is to examine the in vivo effect of star fruit juice at different storage conditions on the activity of alkaline phosphates in female Sprague Dawley (SD rats. A total of 20 healthy female rats weighing 180-200g were used for the experiment. All animals were divided into four groups with five animals per group (n=5. First control was served as control group. Second, third and forth groups were orally treated with a single dose daily of freshly prepared star fruit juice, juice stored for 1-hour and juice stored for 3-hour, respectively for 14 days. All animals were sacrificed at day-15 and various organs such as liver, kidney and heart were removed. All organs were homogenised and centrifuged to prepare cytosolic fraction as a source of alkaline phosphatase enzyme. Protein content of each organ was determined.  p-nitrophenol phosphate was used as a substrate to determine the activity of alkaline phosphatase using spectrophotometer. All results were analysed using Dunnett’s test. Based on the results obtained, a significant increase (P<0.01 of the activity of alkaline phosphatase in rat liver was observed in all star fruit juice treatment groups when compared to control rats. In conclusion, star fruit juice at different storage times was selectively induced the activity of alkaline phosphatase in rat liver but not in the heart and kidney.

Shamala F.

2011-01-01

96

Differentiating intracellular from extracellular alkaline phosphatase activity in soil by sonication.  

Science.gov (United States)

Differentiating intracellular from extracellular enzyme activity is important in soil enzymology, but not easy. Here, we report on an adjusted sonication method for the separation of intracellular from extracellular phosphatase activity in soil. Under optimal sonication conditions [soil:water ratio ?=? 1/8 (w/v) and power density ?=? 15 watt ml(-1)], the activity of alkaline phosphomonoesterase (phosphatase) in a Haplic Cambisol soil increased with sonication time in two distinct steps. A first plateau of enzyme activity was reached between 60 and 100 s, and a second higher plateau after 300 s. We also found that sonication for 100 s under optimal conditions activated most (about 80%) of the alkaline phosphatase that was added to an autoclaved soil, while total bacteria number was not affected. Sonication for 300 s reduced the total bacteria number by three orders of magnitude but had no further effects on enzyme activity. Our results indicate that the first plateau of alkaline phosphatase activity was derived from extracellular enzymes attached to soil particles, and the second plateau to the combination of extracellular and intracellular enzymes after cell lysis. We conclude that our adjusted sonication method may be an alternative to the currently used physiological and chloroform-fumigation methods for differentiating intracellular from extracellular phosphatase activity in soil. Further testing is needed to find out whether this holds for other soil types. PMID:23527006

Qin, Shuping; Hu, Chunsheng; Oenema, Oene

2013-01-01

97

Dietary free fatty acids form alkaline phosphatase-enriched microdomains in the intestinal brush border membrane  

DEFF Research Database (Denmark)

Free fatty acids released during intralumenal digestion of dietary fat must pass through the enterocyte brush border membrane before triacylglycerol reassembly and subsequent chylomicron delivery to the lymph system. In the present work fluorescent BODIPY fatty acid analogs were used to study this membrane passage in organ cultured intestinal mucosal explants. We found that in addition to a rapid uptake into the cytoplasm, a fraction of the fatty acid analogs were inserted directly into the brush border membrane. Furthermore, a brief exposure of microvillar membrane vesicles to a fat mixture mimicking a physiological solution of dietary mixed micelles, rearranged the lipid raft microdomain organization of the membranes. Thus, the fat mixture generated a low-density subpopulation of microvillar detergent resistant membranes (DRMs) highly enriched in alkaline phosphatase (AP). Since this GPI-linked enzyme is the membrane protein in the brush border with the highest affinity for lipid rafts, this implies that free fatty acids selectively insert stably into these membrane microdomains. We have previously shown that absorption of dietary lipids transiently induce a selective endocytosis of AP from the brush border, and from work by others it is known that fat absorption is accompanied by a rise in serum AP and secretion of surfactant-like particles from enterocytes. We propose that these physiological processes may be triggered by the sequestering of dietary free fatty acids in lipid raft microdomains of the brush border.

Hansen, Gert H; Rasmussen, Karina

2011-01-01

98

EXPRESSION OF ALKALINE PHOSPHATASE DURING OSTEOGENIC DIFFERENTIATION OF RAT BONE MARROW STROMAL CELLS  

Directory of Open Access Journals (Sweden)

Full Text Available Introduction: Bone marrow contains a population of stem cells capable of differentiating to osteoblast and forming the bone nodule by dexamethasone. Material and Methods: The stromal cells of bone marrow obtained from 4 to 6 weeks old Spruge-Dawely male rats were grown in primary culture for 7 days and subcultured for 18 days. The cells were cultured in either DMEM medium containing 15% fetal calf serum and antibiotics as the controls or the above medium supplemented with osteogenic supplements (OS: include 10 mM Na-beta glycerophosphate (Na-betaGp, 10 nM dexamethasone (Dex and 50 g/ml ascordic acid (AsA as the examined cultures. After 6, 12 and 18 days of grow up in subculture, the cultures were examined for mineralization and alkaline phosphatase (Apase expression. Results: Mesenchymal stem cells (MSCs in examined cultures underwent a dramatic change in cellular morphology and a significat increase in Apase activity by day 12. The deposition of a calcified matrix on the surface of the culture flasks became evident between days 12 and 18. Conclusion: The addition of osteogenic supplements (OS to MSCs cultures induced Apase expression that contributes to cellular differentiation and mineralization of extracellular matrix.

AKBARI M

2001-01-01

99

Nature of immobilization surface affects antibody specificity to placental alkaline phosphatase.  

Science.gov (United States)

Retention of native conformation of immobilized protein is essential for various applications including selection and detection of specific recombinant antibodies (scFvs). Placental alkaline phosphatase (PAP), an onco-fetal antigen expressed on the surface of several tumors, was immobilized on supermagnetic particles for selection of recombinant antibodies from a human phage display antibody library. The isolated antibodies were found to be cross-reactive to either of the isozymes of alkaline phosphatase, i.e., bone alkaline phosphatase (BAP) or intestinal alkaline phosphatase (IAP) and could not be used for tumor targeting. A specific anti-PAP monoclonal antibody H17E2 was tested for retention of specificity under these conditions. Binding of the antibody to magnetic beads conjugated IAP and BAP along with PAP and the ability of the two isozymes to inhibit its binding to PAP depicted the loss of isozyme specificity of the antibody. However, the antibody retained its specificity to PAP immobilized on polyvinyl chloride (PVC) surface. Enzyme activity was observed on both surfaces. This demonstrates that nature of immobilization may affect antigen-antibody binding in subtle ways, resulting in alteration of conformation of the epitopes. This may have consequences for determining the specificity of antibody binding for proteins that share a high degree of homology. PMID:25321174

Kumar, Mukesh; Khan, Imran; Sinha, Subrata

2015-01-01

100

Radioprotective effect of MPG and WR-2721 against gamma-radiolysis of human placental alkaline phosphatase  

International Nuclear Information System (INIS)

Two-radioprotective drugs - MPG and WR-2721 have been found to protect human placental alkaline phosphatase against gamma radiolysis. Based on current literature and results obtained here free radical scavenging shielding of active sites and/or conformational change due to binding of the drug may be suggested as the possible mechanism of chemical radioprotection. (author)

 
 
 
 
101

Alkaline bone phosphatase activity as related to fluoride ingestion by dairy cattle  

Energy Technology Data Exchange (ETDEWEB)

Alkaline phosphatase prepared from bone and studied in vitro in relation to the effect of fluoride was relatively insensitive to fluoride. Concentrations of fluoride in excess of 10/sup -2/ M were required for inhibition. Experiments with dairy heifers and cows indicated a close correlation between fluoride ingested, fluoride content of the bone, osseous abnormalities, and alkaline bone phosphatase activity. Fluorosis occurred in heifers fed rations containing 66 and 68 ppm of fluoride in contaminated hay and NaF respectively, for a period of 20 months. Additions of about 69 ppm of CaF/sub 2/ to the rations for 20 months caused no detrimental effect to the dairy animals. In an experiment lasting seven years and 108 days, dairy cattle were fed rations containing 12, 27, 49, and 93 ppm of sodium fluoride. When the rations contained 49 or 93 ppm of NaF, osseous abnormalities, excessively high accumulation of fluoride in the bone, and significant increases in alkaline bone phosphatase activity occurred. Results indicated that alkaline bone phosphatase activity was related to abnormal formation.

Miller, G.W.; Shupe, J.L.

1962-01-01

102

Hydrolysis of membrane-bound liver alkaline phosphatase by GPI-PLD requires bile salts.  

Science.gov (United States)

Circulating liver plasma membrane fragments (LPMF) were purified from human serum by means of a monoclonal antileucine aminopeptidase antibody, AD-1. This was done by immunoaffinity chromatography or by incubating the sera with AD-1-coated nitrocellulose disks. Alkaline phosphatase (ALP, EC 3.1.3.1) is bound to these LPMF through a glycosylphosphatidylinositol (GPI) anchor and is referred to as membrane-bound liver ALP (Mem-LiALP). Low concentrations of Triton X-100 or high bile salt concentrations released GPI anchor-bearing LiALP (Anch-LiALP) from purified LPMF; once released, Anch-LiALP was slowly and progressively converted to hydrophilic dimeric LiALP [soluble LiALP (Sol-LiALP)], free from its GPI anchor. Low levels of GPI-specific phospholipase D (GPI-PLD) activity were measured in the pure LPMF. Apparently, this membrane-associated GPI-PLD was released by the action of detergents and contributed to the spontaneous conversion of Anch-LiALP to Sol-LiALP. In the absence of detergents, GPI-PLD had little effect on Mem-LiALP, both in purified form as well as in serum. In vitro, isolated Anch-LiALP was converted to Sol-LiALP by both GPI-specific phospholipase C and GPI-PLD. Sol-LiALP in serum, however, appeared to be the product of GPI-PLD activity only. Five- to tenfold higher concentrations of Triton X-100 were needed to release Anch-LiALP from LPMF in serum, compared with those required in a solution of purified LPMF. In serum, as well as in purified conditions, only a small range of detergent of bile salt concentrations permitted the conversion of Mem-LiALP to Sol-LiALP. A model is proposed for the release in the circulation of Mem-LiALP, Anch-LiALP, and Sol-LiALP, involving both LPMF-associated GPI-PLD and liver sinusoid bile salts. PMID:8897885

Deng, J T; Hoylaerts, M F; De Broe, M E; van Hoof, V O

1996-10-01

103

Membrane-bound alkaline phosphatase from ectopic mineralization and rat bone marrow cell culture.  

Science.gov (United States)

Cells from rat bone marrow exhibit the proliferation-differentiation sequence of osteoblasts, form mineralized extracellular matrix in vitro and release alkaline phosphatase into the medium. Membrane-bound alkaline phosphatase was obtained by method that is easy to reproduce, simpler and fast when compared with the method used to obtain the enzyme from rat osseous plate. The membrane-bound alkaline phosphatase from cultures of rat bone marrow cells has a MW(r) of about 120 kDa and specific PNPP activity of 1200 U/mg. The ecto-enzyme is anchored to the plasma membrane by the GPI anchor and can be released by PIPLC (selective treatment) or polidocanol (0.2 mg/mL protein and 1% (w/v) detergent). The apparent optimum pH for PNPP hydrolysis by the enzyme was pH 10. This fraction hydrolyzes ATP (240 U/mg), ADP (350 U/mg), glucose 1-phosphate (1100 U/mg), glucose 6-phosphate (340 U/mg), fructose 6-phosphate (460 U/mg), pyrophosphate (330 U/mg) and beta-glycerophosphate (600 U/mg). Cooperative effects were observed for the hydrolysis of PPi and beta-glycerophosphate. PNPPase activity was inhibited by 0.1 mM vanadate (46%), 0.1 mM ZnCl2 (68%), 1 mM levamisole (66%), 1 mM arsenate (44%), 10 mM phosphate (21%) and 1 mM theophylline (72%). We report the biochemical characterization of membrane-bound alkaline phosphatase obtained from rat bone marrow cells cultures, using a method that is simple, rapid and easy to reproduce. Its properties are compared with those of rat osseous plate enzyme and revealed that the alkaline phosphatase obtained has some kinetics and structural behaviors with higher levels of enzymatic activity, facilitating the comprehension of the mineralization process and its function. PMID:16798036

Simão, Ana Maria S; Beloti, Márcio M; Cezarino, Rodrigo M; Rosa, Adalberto Luiz; Pizauro, João M; Ciancaglini, Pietro

2007-04-01

104

Localization of alkaline phosphatase in three gram-negative rumen bacteria.  

Science.gov (United States)

Of the three species (Bacteroides ruminicola, B. succinogenes, and Megasphaera elsdenii) of anaerobic gram-negative rumen bacteria studied, only B. ruminicola produced significant amounts of alkaline phosphatase. This enzyme, which is constitutive, showed a greater affinity for p-nitrophenylphosphate than for sodium-beta-glycerophosphate and was shown to be located exclusively in the periplasmic space of log-phase cells. Small amounts of this enzyme were released from these cells in stationary-phase cultures, but washing in 0.01 M MgCl(2) and the production of spheroplasts by using lysozyme in 0.01 M MgCl(2) did not release significant amounts of the enzyme. Exposure to 0.2 M MgCl(2) did not release significant amounts of the periplasmic alkaline phosphatase of the cell, and when these cells were spheroplasted with lysozyme in 0.2 M MgCl(2) only 25% of the enzyme was released. Spheroplasts were formed spontaneously in aging cultures of B. ruminicola, but even these cells retained most of their periplasmic alkaline phosphatase. It was concluded that the alkaline phosphatase of B. ruminicola is firmly bound to a structural component within the periplasmic area of the cell wall and that the enzyme is released in large amounts only when the cells break down. The behavior of alkaline phosphatase in this bacterium contrasts with that of conventional periplasmic enzymes of aerobic bacteria, which are released upon conversion into spheroplasts by lysozyme and ethylenediaminetetraacetic acid and by other types of cell wall damage. All three species of bacteria studied here, as well as bacteria found in mixed populations in the rumen, have thick, complex layers external to the double-track layer of their cell walls. In addition, B. ruminicola produces a loose extracellular material. PMID:4147649

Cheng, K J; Costerton, J W

1973-10-01

105

Zinc and alkaline phosphatase in developing rat oral mucosa  

International Nuclear Information System (INIS)

Alkaline phosphates (AlkPase) of many different tissues and species have been shown to be zinc metalloenzymes. Specific regions of rat oral mucosa have a high activity of AlkPase. Combined autoradiography and enzyme histochemistry showed that they also retained injected radioactive zinc (65Zn). The AlkPase activity was inactivated by EDTA and reactivated with zinc. However, it could not be verified by polyacrylamide gel electrophoresis, combined with radioactivity measurements and enzyme analysis, that the 65Zn uptake of oral mucosa was incorporated in the AlkPase molecule

106

Detection of endogenous alkaline phosphatase activity in intact cells by flow cytometry using the fluorogenic ELF-97 phosphatase substrate  

Science.gov (United States)

BACKGROUND: The alkaline phosphatase (AP) substrate 2-(5'-chloro-2'-phosphoryloxyphenyl)-6-chloro-4-(3H)-quinazolinone (ELF((R))-97 for enzyme-labeled fluorescence) has been found useful for the histochemical detection of endogenous AP activity and AP-tagged proteins and oligonucleotide probes. In this study, we evaluated its effectiveness at detecting endogenous AP activity by flow cytometry. METHODS: The ELF-97 phosphatase substrate was used to detect endogenous AP activity in UMR-106 rat osteosarcoma cells and primary cultures of chick chondrocytes. Cells were labeled with the ELF-97 reagent and analyzed by flow cytometry using an argon ultraviolet (UV) laser. For comparison purposes, cells were also assayed for AP using a Fast Red Violet LB azo dye assay previously described for use in detecting AP activity by flow cytometry. RESULTS: The ELF-97 phosphatase substrate effectively detected endogenous AP activity in UMR-106 cells, with over 95% of the resulting fluorescent signal resulting from AP-specific activity (as determined by levamisole inhibition of AP activity). In contrast, less than 70% of the fluorescent signal from the Fast Red Violet LB (FRV) assay was AP-dependent, reflecting the high intrinsic fluorescence of the unreacted components. The ELF-97 phosphatase assay was also able to detect very low AP activity in chick chondrocytes that was undetectable by the azo dye method. CONCLUSIONS: The ELF-97 phosphatase assay was able to detect endogenous AP activity in fixed mammalian and avian cells by flow cytometry with superior sensitivity to previously described assays. This work also shows the applicability of ELF-97 to flow cytometry, supplementing its previously demonstrated histochemical applications. Copyright 1999 Wiley-Liss, Inc.

Telford, W. G.; Cox, W. G.; Stiner, D.; Singer, V. L.; Doty, S. B.

1999-01-01

107

Coupling immobilized alkaline phosphatase-based automated diagonal capillary electrophoresis to tandem mass spectrometry for phosphopeptide analysis.  

Science.gov (United States)

Automated diagonal capillary electrophoresis is a two-dimensional separation method that incorporates an immobilized enzyme reactor at the distal end of the first capillary and employs identical electrophoretic separation modes in both dimensions. Components undergo a preliminary separation in the first capillary. Fractions are parked in the reactor where some components undergo transformation. The fractions are then periodically transferred to the second capillary and replaced by the next components in the sample. Components that are not modified by the reactor will have identical mobility in both dimensions and fall on the diagonal of a reconstructed two-dimensional electropherogram, while analyte that undergoes modification will fall off the diagonal. In this study, alkaline phosphatase was immobilized in a monolithic reactor. An LTQ-Orbitrap Velos mass spectrometer was used to monitor analytes as they migrated from the second capillary. The system was used to characterize the phosphorylation status of a tryptic digest of ?-casein in a background prepared from a 22-fold excess of the tryptic digest of bovine serum albumin. 120 fractions underwent automated treatment in the alkaline phosphatase reactor and separation in the second dimension capillary for over 40 min; nine phosphorylated ?-casein peptides that produced 20 different phosphorylation states were detected with high confidence. PMID:24148505

Mou, Si; Sun, Liangliang; Wojcik, Roza; Dovichi, Norman J

2013-11-15

108

Kidney bone disease and mortality in CKD: revisiting the role of vitamin D, calcimimetics, alkaline phosphatase, and minerals.  

Science.gov (United States)

Recent evidence suggests that the traditional syndromes known as renal osteodystrophy, secondary hyperparathyroidism, and vitamin D deficiency are related to mortality in persons with moderate to advanced chronic kidney disease (CKD). The so-called 'kidney bone disease', also known as 'mineral and bone disorders', is defined to include bone disorders, mineral disarrays, and vascular calcification. We have identified 14 common and clinically relevant conditions of contemporary nature that are related to the kidney bone disease, including calcitriol (active vitamin D) deficiency, 25(OH)-vitamin D deficiency, biochemical hyperparathyroidism, relatively low parathyroid hormone (PTH) level, increased serum alkaline phosphatase (hyperphosphatasemia), elevated fibroblast growth factor (FGF)-23, high turnover bone disease, adynamic bone disease, uremic osteoporosis, vascular calcification, hyper- and hypophosphatemia, and hyper- and hypocalcemia. We present a critical review of these 14 conditions with emphasis on patient survival and other pertinent clinical outcomes. We also review unresolved controversies surrounding the management of these conditions by administration of nutritional vitamin D (ergocalciferol and cholecalciferol), vitamin D receptor activators (calcitriol, alphacalcidiol, doxercalciferol), D-mimetics (paricalcitol, maxacalcitol), calcimimetics (cinacalcet), recombinant PTH (teriparatide), and receptor activator of nuclear factor-kappaB ligand modulators (denosumab); compare mortality predictability of PTH and alkaline phosphatase; and examine potential risks of bone disorders and mineral disarrays in CKD patients. PMID:20671739

Kalantar-Zadeh, Kamyar; Shah, Anuja; Duong, Uyen; Hechter, Rulin C; Dukkipati, Ramanath; Kovesdy, Csaba P

2010-08-01

109

Effects of sodium nitroprusside activity of acid and alkaline invertases and alkaline phosphatase in lemongrass (Cymbopogon flexuosus Steud Wats  

Directory of Open Access Journals (Sweden)

Full Text Available Here we report the effects of SNP, a nitric oxide donor on sucrose metabolizing enzymes, acid and alkaline invertase (EC 3.2.1.26 and 3.2.1.153 and ubiquitous alkaline phosphatase (EC 3.1.3.1 in four lemongrass varieties viz., Krishna, Cauveri, Nima and Cheerharit. For the study, two 15 d lemongrass tillers were cut and immediately dipped into the test tubes containing SNP solution (5 mL of variable strength (1 to 5 mM and one without SNP (as control; kept for 4 h under mild sunlight. The results revealed that moderate SNP concentration (2 mM was most effective, caused drastic reduction (40% in protein content in var. Nima followed by Krishna (33%, Cauveri (17% and Cheerharit (12%. In contrast, SNP (1 mM has impressively enhanced protein content in all the lemongrass varieties. The SNP (2 mM markedly inhibited the activity of acid invertase by 38% in Cheerharit, 35% Nima and 28% Cauveri whereas and alkaline invertase by 21, 28 and 24% respectively in var. Cheerharit, Nima and Krishna. Similarly, SNP (5 mM severely inhibited (~ 63% the activity of the ALP in lemongrass var. Cauveri and Nima, 50% in Krishna and relatively less 23% in Cheerharit as compared to the control. However, in var. Nima, 50% loss in ALP activity had already been occurred after 2 mM SNP treatment. These results primarily suggests that NO interferes sucrose metabolism by anonymously hindering the activity of acid and alkaline invertase and ubiquitous alkaline phosphatase in lemongrasses.

Deepak Ganjewala

2010-01-01

110

Método fosfatasa alcalina anti-fosfatasa alcalina para el diagnóstico de inmunodeficiencias celulares Alkaline phosphatase-anti-alkaline phosphatase method for the diagnosis of cell immunodeficiencies  

Directory of Open Access Journals (Sweden)

Full Text Available Para el estudio de 25 pacientes con infecciones recurrentes y un grupo control de 25 individuos supuestamente sanos, se aplicó, en nuestro laboratorio, el método inmunocitoquímico de fosfatasa alcalina anti-fosfatasa alcalina, para la cuantificación de las principales subpoblaciones de linfocitos T identificados con los anticuerpos monoclonales: anti-CD3, anti-CD4 y anti-CD8. Se obtuvieron diferencias significativas para las subpoblaciones TCD3 y CD4 positivos (p For the study of 25 patients with recurrent infections and a control group of 25 supposedly healthy individuals, our Laboratory applied the immunocytochemical method of alkaline phosphatase - anti-alkaline phosphatase for the quantitation of the main T-lymphocyte subgroups identified with monoclonal antibodies:antiCD3, anti-CD4 and anti-CD8. There were significant differences in positive TCD3 and CD4 subsets (p<0,05. Because this is a low cost and quick method, it may be applied by other immunodiagnosis labs throughout the country

Berta B Socarrás Ferrer

2001-12-01

111

Development of conductometric biosensors based on alkaline phosphatases for the water quality control  

CERN Document Server

Researches are focused on the elaboration of enzymatic microconductometric device for heavy metal ions detection in water solutions. The manuscript includes a general introduction, the first chapter contains bibliographic review, the second chapter described the fundamentals of conductometric transducers, the third chapter examining the possibility to create and to optimize conductometric biosensor based on bovine alkaline phosphatase for heavy metals ions detection, the fourth chapter devoted to creation and optimization of conductometric biosensor based on alkaline phosphatase active microalgae and sol gel technology, the last chapter described application of the proposed algal biosensor for measurements of heavy metal ions toxicity of waste water, general conclusions stating the progresses achieved in the field of environmental monitoring

Berezhetskyy, A

2008-01-01

112

Significance of bone specific alkaline phosphatase as a tumor marker in malignant bone tumor  

International Nuclear Information System (INIS)

The relationship between total alkaline phosphatase activity and bone forming lesion is a well known fact. But alkaline phosphatase consist mainly of two portion (liver, bone). To clarify the exact activity of bone forming tissue, quantitative measurement of BALP is essential. Two finds of tests were performed for their feasibility as a laboratory test (wheat germ lectin vs electrophoresis). We analyzed 40 bony lesion and got 58 samples. Lectin method was simple, economic, with reliable resproducability. Owing to the small number of test sample, we could not identify the relationship between the disease activity and measured BALP level. Further collection of clinical sample and analysis the pattern of BALP on each clinical settings. (author). 8 refs

113

Export of Thermus thermophilus alkaline phosphatase via the twin-arginine translocation pathway in Escherichia coli.  

Science.gov (United States)

The bacterial twin-arginine translocation (Tat) pathway is distinct from the Sec system by its remarkable capacity to export folded enzymes. To address the question whether the two systems are capable of translocating homologous enzymes catalyzing the same reaction, we cloned the tap gene encoding Thermus thermophilus alkaline phosphatase (Tap) and expressed it in Escherichia coli. Unlike the alkaline phosphatase of E. coli, which is translocated through the Sec system and then activated in the periplasm, Tap was exported exclusively via the Tat pathway and active Tap precursor was observed in the cytoplasm. These results demonstrate that two sequence and functional related enzymes are exported by distinct protein transport systems, which may play an integral role in the bacterial adaptation to their environment during the evolution. PMID:11591380

Angelini, S; Moreno, R; Gouffi, K; Santini, C; Yamagishi, A; Berenguer, J; Wu, L

2001-10-01

114

Significance of bone specific alkaline phosphatase as a tumor marker in malignant bone tumor  

Energy Technology Data Exchange (ETDEWEB)

The relationship between total alkaline phosphatase activity and bone forming lesion is a well known fact. But alkaline phosphatase consist mainly of two portion (liver, bone). To clarify the exact activity of bone forming tissue, quantitative measurement of BALP is essential. Two finds of tests were performed for their feasibility as a laboratory test (wheat germ lectin vs electrophoresis). We analyzed 40 bony lesion and got 58 samples. Lectin method was simple, economic, with reliable resproducability. Owing to the small number of test sample, we could not identify the relationship between the disease activity and measured BALP level. Further collection of clinical sample and analysis the pattern of BALP on each clinical settings. (author). 8 refs.

Kim, Sug Jun; Jeon, Dae Geun; Huh, Kwang [Korea Cancer Center Hospital, Seoul (Korea, Republic of)

1998-01-01

115

Characterization of a highly thermostable alkaline phosphatase from the euryarchaeon Pyrococcus abyssi  

Digital Repository Infrastructure Vision for European Research (DRIVER)

This work reports the first isolation and characterization of an alkaline phosphatase (AP) from a hyperthermophilic archaeon. An AP gene from Pyrococcus abyssi, a euryarchaeon isolated from a deep-sea hydrothermal vent, was cloned and the enzyme expressed in Escherichia coli. Analysis of the sequence showed conservation of the active site and structural elements of the E. coli AP. The recombinant AP was purified and characterized. Monomeric and homodimeric active forms were detected, with a m...

Zappa, Se?bastien; Rolland, Jean-luc; Flament, Didier; Gueguen, Yannick; Boudrant, Joseph; Dietrich, Jacques

2001-01-01

116

Evaluation of Testosterone and Alkaline Phosphatase Activity Changes in Epidydimis of Toxoplasma gondii Infected Rats  

Digital Repository Infrastructure Vision for European Research (DRIVER)

Objective: Toxoplasma gondii is a widespread protozoan parasite that infects a broad range of warm blooded animals and humans. The present study was investigated to evaluate testosterone, alkaline phosphatase activity and malondialdehyde in male rats experimentally infected by Toxoplasma gondii, RH strain.Material & Methods: Male Wistar rats (n=20) were allocated into two groups, group one (n=10) that received 0.6 cc tachyzoites of T. gondii intraperitoneally (I.P.), and control group (n=10) ...

Fatemeh Afshari; Amir Mahdi Imani; Sasan Najjari Asl; Farhang, Hossein H.; Khazar Ghasempour; Ezzatzadeh; Nava Ainechi

2013-01-01

117

A Disposable Alkaline Phosphatase-Based Biosensor for Vanadium Chronoamperometric Determination  

Digital Repository Infrastructure Vision for European Research (DRIVER)

A chronoamperometric method for vanadium ion determination, based on the inhibition of the enzyme alkaline phosphatase, is reported. Screen-printed carbon electrodes modified with gold nanoparticles were used as transducers for the immobilization of the enzyme. The enzymatic activity over 4-nitrophenyl phosphate sodium salt is affected by vanadium ions, which results in a decrease in the chronoamperometric current registered. The developed method has a detection limit of 0.39 ± 0.06 ?M, a r...

Ana Lorena Alvarado-Gámez; María Asunción Alonso-Lomillo; Olga Domínguez-Renedo; María Julia Arcos-Martínez

2014-01-01

118

In situ hybridization of cytokine mRNA using alkaline phosphatase-labelled oligodeoxynucleotide probes  

DEFF Research Database (Denmark)

In situ hybridization is a powerful tool for visualizing cellular gene expression in morphologically preserved brain tissue giving precise information on the regional expression of specific mRNA sequences in cells of diverse phenotype. Here, we describe a sensitive, simple, and robust method using alkaline phosphatase (AP)-labelled oligodeoxynucleotide probes to detect cytokine mRNA in the acutely injured or inflamed mouse CNS.

Clausen, Bettina Hjelm; Fenger, Christina

2013-01-01

119

Alkaline Phosphatase and Other Hydrolyases Produced by Cenococcum graniforme, an Ectomycorrhizal Fungus  

Digital Repository Infrastructure Vision for European Research (DRIVER)

Cell extracts of Cenococcum graniforme have been found to contain the following hydrolytic enzymes: protease, esterase, ?-d-galactopyranosidase, ?-d-galactopyranosidase, ?-d-mannopyranosidase, ?-d-xylopyranosidase, ?-d-glucopyranosidase, ?-d-glucopyranosidase, and alkaline phosphatase. Sulfatase, inorganic pyrophosphatase, and ?-d-mannopyranosidase were not detected in the extracts. ?-d-Xylopyranosidase and ?-d-mannopyranosidase were most active in the neutral pH range, protease and ...

Bae, Kwang-sung; Barton, Larry L.

1989-01-01

120

Deletion map of the Escherichia coli structural gene for alkaline phosphatase, phoA.  

Digital Repository Infrastructure Vision for European Research (DRIVER)

Lambda transducing phages containing portions of the phoA gene have been isolated and used to construct a deletion map of the phoA gene. The isolation of a plaque-forming lambda transducing phage carrying the entire phoA gene is also described. Two new methods for screening or selection of mutants that have altered levels of alkaline phosphatase activity are reported.

Sarthy, A.; Michaelis, S.; Beckwith, J.

1981-01-01

 
 
 
 
121

A new signal sequence trap using alkaline phosphatase as a reporter.  

Digital Repository Infrastructure Vision for European Research (DRIVER)

Secreted and transmembrane proteins are critical to the cell-cell interactions governing normal development and carcinogenesis. To facilitate the identification of such molecules, we have developed a novel signal sequence trap that uses human placental alkaline phosphatase as a reporter. Libraries from mouse prostate and human prostatic carcinoma were constructed to test the PST (peptide signal trap) system, resulting in the identification of several secreted and transmembrane proteins.

Chen, H.; Leder, P.

1999-01-01

122

Use of leucocyte alkaline phosphatase (LAP) score in differentiating malignant from benign paraproteinaemias.  

Digital Repository Infrastructure Vision for European Research (DRIVER)

The leucocyte alkaline phosphatase (LAP) score of peripheral blood neutrophils was examined in 20 patients with multiple myeloma and compared with the score in 18 patients with monoclonal gammopathy of undetermined significance (MGUS). The mean (95% confidence limit) LAP score in those with multiple myeloma was 186 (169-218) compared with 92 (64-120) in the MGUS group. In the multiple myeloma group all but one patient had a high LAP score, irrespective of disease. No cause for raised LAP, suc...

Majumdar, G.; Hunt, M.; Singh, A. K.

1991-01-01

123

Distribution of alkaline and acid phosphatases in the duodenal wall of native sheep by using different fixatives  

Directory of Open Access Journals (Sweden)

Full Text Available Ten duodeni of adult ram were fixed in chilled acetone, 80% ethyl alcohol, formol- alcohol solution, alcoholic bouinssolution and neutral buffered formalin solution. The distribution of alkaline and acid phosphatases were similar in theirlocation but different in their intensity and distribution according to different fixative The distribution of alkaline phosphatasein absorptive columnar cell was more intense than in goblet cells, whereas the concentration of acid phosphatase was moreintense in goblet cells than in absorptive cells in the mucosa of sheep duodenum. The study revealed that the samples wasfixed with chilled acetone gave highest reaction for alkaline and acid phosphatases than other fixative samples. No reaction foralkaline phosphatase include the lower parts of intestinal glands, paneth cells and sub mucosal glands in different fixative,whereas, paneth cells and sub mucosal glands revealed wreaked reaction for acid phosphatase in samples fixed in 80% ethylalcohol and chilled acetone respectively in duodenum of native sheep.

N. S. Ahmed

2010-01-01

124

Effect of MPG on the radiation induced changes in the intestinal activity of alkaline phosphatase in mice  

International Nuclear Information System (INIS)

Adult male Swiss mice were exposed to 250, 500 and 1000 R of gamma rays with or without a prior intraperitoneal injection of 20 mg/kg MPG. Alkaline phosphatase activity in the ileum was estimated at different post irradiation intervals. The changes in the enzyme activity was dose dependent. MPG reduced the radiation induced increase in the alkaline phosphatase activity of the intestine. (author)

125

Lactoperoxidase-125I localization of salt-extractable alkaline phosphatase on the cytoplasmic membrane of Bacillus licheniformis.  

Digital Repository Infrastructure Vision for European Research (DRIVER)

Previous histochemical and biochemical localizations of alkaline phosphatase in Bacillus licheniformis MC14 have shown that the membrane-associated form of the enzyme is located on the inner surface of the cytoplasmic membrane, and soluble forms are located in the periplasmic space and in the growth medium. The distribution of salt-extractable alkaline phosphatase on the surfaces of the cytoplasmic membrane of B. licheniformis MC14 was determined by using lactoperoxidase-125I labeling techniq...

Spencer, D. B.; Hulett, F. M.

1981-01-01

126

Alterations in activities of acid phosphatase, alkaline phosphatase, ATPase and ATP content in response to seasonally varying Pi status in okra (Abelmoschus esculentus).  

Science.gov (United States)

Phosphorus (P) is the second most important macronutrient for plant growth. Plants exhibit numerous physiological and metabolic adaptations in response to seasonal variations in phosphorus content. Activities of acid and alkaline phosphatases, ATPase and ATP content were studied in summer, rainy and winter seasons at two different developmental stages (28 and 58 days after sowing) in Okra. Activities of both acid and alkaline phosphatases increased manifold in winter to cope up with low phosphorus content. ATP content and ATPase activity were high in summer signifying an active metabolic period. Phosphorus deficiency is characterized by low ATP content and ATPase activity (which are in turn partly responsible for a drastic reduction in growth and yield) and enhanced activities of acid and alkaline phosphatases which increase the availability of P in P-deficient seasons. PMID:15529876

Sen, Supatra; Mukherji, S

2004-04-01

127

Serum prostate-specific acid phosphatase: development and validation of a specific radioimmunoassay  

International Nuclear Information System (INIS)

We describe radioimmunoassay for human prostatic acid phosphatase [orthophosphoric-monoester phosphohydrolase (acid optimum), EC 3.1.3.2] in serum, with use of monospecific antisera raised in rabbits against highly purified acid phosphatase from human prostates. The antiserum did not cross react with partly purified acid phosphatases from human spleen, erythrocytes, or synovial tissues. 125I-labeled acid phosphatase was prepared by a Chloramine T method, and the bound and free antigen was separated in the assay by use of anti-rabbit gamma-globulin raised in sheep. Uniform low nonspecific binding of the [125I]acid phosphatase was achieved by using acid-phosphatase-free serum to prepare standard curves and diluted samples of serum with high acid phosphatase activities. Concentrations of immunoreactive acid phosphatase in the serum of healthy men ranged from <1 to 10 ?g/liter and for 12 patients with advanced prostatic carcinoma between 100 and 500 ?g/liter. The concentrations of the enzyme in sera of patients with benign prostatic hyperplasia were very similar to those in sera of the reference group

128

Epigenetic regulation of alkaline phosphatase in human cells of the osteoblastic lineage.  

Science.gov (United States)

Epigenetic mechanisms play an important role in the tissue-specific regulation of gene expression. This study analyzed the relationship between tissue non-specific alkaline phosphatase (ALPL) gene expression and the methylation of a CpG island located in its proximal region. Gene expression was analyzed by real time RT-qPCR in primary human osteoblasts (hOBs), the osteoblastic cell line MG-63, the mammary cell line MCF-7, and bone tissue. DNA methylation was analyzed by qMSP in those cells and also in lining osteoblasts and in osteocytes obtained from human bone samples by laser-assisted capture. hOBs expressed much more ALPL mRNA than MG-63 cells (7.3±3.2 vs. 0.2±0.1 arbitrary units, respectively). hOBs showed a very weak DNA methylation (MG-63 had a higher degree of methylation (58±6%). Likewise, MCF-7 cells, which scarcely expressed ALPL, had a hypermethylated CpG island. Thus, the degree of methylation in the CpG island was inversely associated with the transcriptional levels of ALPL in the studied cells. Furthermore, treatment with the DNA demethylating agent AzadC induced a 30-fold increase in ALPL expression, in MG-63 cells, accompanied by a parallel increase in alkaline phosphatase activity. However, AzadC did not affect ALPL levels in the already hypomethylated hOBs. In addition, in microdissected osteocytes, which do not express alkaline phosphatase, the CpG island was highly methylated (>90%), whereas lining osteoblasts showed an intermediate degree of methylation (58±13%). These results suggest an important role of DNA methylation in the regulation of ALPL expression through the osteoblast-osteocyte transition. PMID:21700004

Delgado-Calle, Jesús; Sañudo, Carolina; Sánchez-Verde, Lydia; García-Renedo, Raúl J; Arozamena, Jana; Riancho, José A

2011-10-01

129

Refined structure of alkaline phosphatase from Escherichia coli at 2.8 A resolution.  

Science.gov (United States)

The structure of alkaline phosphatase from Escherichia coli has been determined to 2.8 A resolution. The multiple isomorphous replacement electron density map of the dimer at 3.4 A was substantially improved by molecular symmetry averaging and solvent flattening. From these maps, polypeptide chains of the dimer were built using the published amino acid sequence. Stereochemically restrained least-squares refinement of this model against native data, starting with 3.4 A data and extending in steps to 2.8 A resolution, proceeded to a final overall crystallographic R factor of 0.256. Alkaline phosphatase-phosphomonoester hydrolase (EC 3.1.3.1) is a metalloenzyme that forms an isologous dimer with two reactive centers 32 A apart. The topology of the polypeptide fold of the subunit is of the alpha/beta class of proteins. Despite the similarities in the overall alpha/beta fold with other proteins, alkaline phosphatase does not have a characteristic binding cleft formed at the carboxyl end of the parallel sheet, but rather an active pocket that contains a cluster of three functional metal sites located off the plane of the central ten-stranded sheet. This active pocket is located near the carboxyl ends of four strands and the amino end of the antiparallel strand, between the plane of the sheet and two helices on the same side. Alkaline phosphatase is a non-specific phosphomonoesterase that hydrolyzes small phosphomonoesters as well as the phosphate termini of DNA. The accessibility calculations based on the refined co-ordinates of the enzyme show that the active pocket barely accommodates inorganic phosphate. Thus, the alcoholic or phenolic portion of the substrate would have to be exposed on the surface of the enzyme. Two metal sites, M1 and M2, 3.9 A apart, are occupied by zinc. The third site, M3, 5 A from site M2 and 7 A from site M1, is occupied by magnesium or, in the absence of magnesium, by zinc. As with other zinc-containing enzymes, histidine residues are ligands to zinc site M1 (three) and to zinc site M2 (one). Ligand assignment and metal preference indicate that the crystallographically found metal sites M1, M2 and M3 correspond to the spectroscopically deduced metal sites A, B and C, respectively. Arsenate, a product analog and enzyme inhibitor, binds between Ser102 and zinc sites M1 and M2. The position of the guanidinium group of Arg 166 is within hydrogen-bonding distance from the arsenate site.(ABSTRACT TRUNCATED AT 400 WORDS) PMID:3910843

Sowadski, J M; Handschumacher, M D; Murthy, H M; Foster, B A; Wyckoff, H W

1985-11-20

130

Effect of Diazinon on Acid and Alkaline Phosphatase Activities in Plasma and Organs of Clarias gariepinus  

Digital Repository Infrastructure Vision for European Research (DRIVER)

The aim of this study was to determine the effect of the pesticide, diazinon, on phosphatases in the plasma and organs on Clarias gariepinus. Adult Clarias gariepinus were exposed in four replicates to varying sublethal concentrations diazinon (ranging from 1.00 to 10.0 mg/L) in 30-day semi-static bioassays. Alkaline phoshatase (ALP) and acid phosphate (ACP) were determined in plasma and other organs (gastrointestinal tract - GIT, kidney, muscle, gill and liver) of the fish after the experime...

Inyang, I. R.; Ogamba, E. R. Daka And E. N.

2011-01-01

131

Biocompatibility and Alkaline Phosphatase Activity of Phosphorylated Chitooligosaccharides on the Osteosarcoma MG63 Cell Line  

Digital Repository Infrastructure Vision for European Research (DRIVER)

Phosphorylated chitooligosaccharides (P-COS) were prepared using a H3PO4, P2O5, Et3PO4 and hexanol solvent system. The P-COS were characterized by Fourier Transform Infrared Spectroscopy (FT-IR), Thermo gravimetric-Differential Thermal Analyzer (TG-DTA), 13C NMR, 31P NMR, X-ray diffraction analysis, solubility studies, biocompatibility and Alkaline Phosphatase Activity (ALP). The results reveal that phosphorylation occurred at the C3 and C6 position of OH groups and the C2 position of NH2 gro...

Jayachandran Venkatesan; Ratih Pangestuti; Zhong-Ji Qian; BoMi Ryu; Se-Kwon Kim

2010-01-01

132

Identification of enterotoxigenic Escherichia coli with synthetic alkaline phosphatase-conjugated oligonucleotide DNA probes.  

Digital Repository Infrastructure Vision for European Research (DRIVER)

Alkaline phosphatase-conjugated (AP) 26-base oligonucleotide DNA probes were compared with the same probes labeled with gamma-32P for the identification of heat-labile (LT) and heat-stable (ST) enterotoxigenic Escherichia coli (ETEC). The AP oligonucleotide probes were as sensitive as the radiolabeled (RL) probes in detecting LT and STA-2 target cell DNA, but the AP ST probe, which differed from STA-1 by two bases, was less sensitive than the RL probe in detecting STA-1 DNA (6.25 versus 0.78 ...

Seriwatana, J.; Echeverria, P.; Taylor, D. N.; Sakuldaipeara, T.; Changchawalit, S.; Chivoratanond, O.

1987-01-01

133

Comparative effectiveness of the cholera toxin B subunit and alkaline phosphatase as carriers for oral vaccines.  

Digital Repository Infrastructure Vision for European Research (DRIVER)

The purpose of this study was to determine whether the B subunit of cholera toxin (CtxB) has adjuvant activity over and above serving as a carrier protein for orally administered vaccines. An oligonucleotide that encodes an antigenic determinant (GtfB.1) from the glucosyltransferase B gene (gtfB) of Streptococcus mutans was genetically fused to the 5' terminus of either the CtxB gene (ctxB) or the Escherichia coli alkaline phosphatase gene (phoA). The resulting chimeric proteins were expresse...

Dertzbaugh, M. T.; Elson, C. O.

1993-01-01

134

Alternate modes of binding in two crystal structures of alkaline phosphatase-inhibitor complexes.  

Digital Repository Infrastructure Vision for European Research (DRIVER)

Two high resolution crystal structures of Escherichia coli alkaline phosphatase (AP) in the presence of phosphonate inhibitors are reported. The phosphonate compounds, phosphonoacetic acid (PAA) and mercaptomethylphosphonic acid (MMP), bind competitively to AP with dissociation constants of 5.5 and 0.6 mM, respectively. The structures of the complexes of AP with PAA and MMP were refined at high resolution to crystallographic R-values of 19.0 and 17.5%, respectively. Refinement of the AP-inhib...

Holtz, K. M.; Stec, B.; Myers, J. K.; Antonelli, S. M.; Widlanski, T. S.; Kantrowitz, E. R.

2000-01-01

135

Cloning and characterization of the Bacillus licheniformis gene coding for alkaline phosphatase.  

Digital Repository Infrastructure Vision for European Research (DRIVER)

The structural gene for alkaline phosphatase (orthophosphoric monoester phosphohydrolase; EC 3.1.3.1) of Bacillus licheniformis MC14 was cloned into the Pst1 site of pMK2004 from chromosomal DNA. The gene was cloned on an 8.5-kilobase DNA fragment. A restriction map was developed, and the gene was subcloned on a 4.2-kilobase DNA fragment. The minimum coding region of the gene was localized to a 1.3-kilobase region. Western blot analysis was used to show that the gene coded for a 60,000-molecu...

Hulett, F. M.

1984-01-01

136

Measurement of alkaline phosphatase of intestinal origin in plasma by p-bromotetramisole inhibition.  

Digital Repository Infrastructure Vision for European Research (DRIVER)

L-p-bromotetramisole was used to inhibit non-intestinal alkaline phosphatase (of liver or bone origin) (EC 3.1.3.1; ALP) in plasma, and intestinal ALP was measured from the uninhibited activity. The method of determination is convenient and correlated well with measurement by immunocapture assay. If carried out in parallel with wheat-germ lectin precipitation of bone ALP, subtraction of intestinal ALP activity from that of non-bone ALP in the supernatant can be used to measure the ALP that or...

Kuwana, T.; Rosalki, S. B.

1991-01-01

137

Influence of chronic Cd intoxication on the alkaline phosphatase activity of liver and kidney; biochemical, histochemical and histological investigations.  

Science.gov (United States)

To distinguish between: (a) the decrease of activity of alkaline phosphatase due to inhibition by cadmium; and (b) the decline of enzyme activity due to cell destruction, 20 adult female Wistar rats were treated 3 times/week with 0.5 mg/kg CdCl2 (by subcutaneous injection) during 28 weeks. Controls received the same volume of 0.9% NaCl solution. The animals were killed at different intervals. The liver and kidneys were investigated with biochemical, histochemical, light and electron microscopical techniques. The liver homogenates show an increase of alkaline phosphatase activity after about 12-13 weeks, whereas the activity of this enzyme in the kidney decreases. This activity could not be restored by the administration of Zn ions to the medium of the enzyme assay. However, in previous in vitro experiments the Cd inhibited alkaline phosphatase activity could be totally restored by such a Zn administration. By histochemical assay of alkaline phosphatase in the renal cortex a decrease of enzyme activity was demonstrated. By evaluation of the results obtained with light and electron microscopy, in combination with the biochemical results, the decrease of alkaline phosphatase activity in the kidney should be considered as a real decrease in the amount of this enzyme due to cell degeneration. PMID:231335

Peereboom-Stegeman, J H; Melet, J; Peereboom, J W; Hooghwinkel, G J

1979-09-01

138

A missense mutation in the human liver/bone/kidney alkaline phosphatase gene causing a lethal form of hypophosphatasia  

International Nuclear Information System (INIS)

Hypophosphatasia is an inherited disorder characterized by defective bone mineralization and a deficiency of serum and tissue liver/bone/kidney alkaline phosphatase (L/B/K ALP) activity. Clinical severity is variable, ranging from death in utero (due to severe rickets) to pathologic fractures first presenting in adult life. Affected siblings, however, are phenotypically similar. Severe forms of the disease are inherited in an autosomal recessive fashion; heterozygotes often show reduced serum ALP activity. The specific gene defects in hypophosphatasia are unknown but are thought to occur either at the L/B/K ALP locus or within another gene that regulates L/B/K ALP expression. The authors used the polymerase chain reaction to examine L/B/K ALP cDNA from a patient with a perinatal (lethal) form of the disease. They observed a guanine-to-adenine transition in nucleotide 711 of the cDNA that converts alanine-162 of the mature enzyme to threonine. The affected individual, whose parents are second cousins, is homozygous for the mutant allele. Introduction of this mutation into an otherwise normal cDNA by site-directed mutagenesis abolished the expression of active enzyme, demonstrating that a defect in the L/B/K ALP gene results in hypophosphatasia and that the enzyme is, therefore, essential for normal skeletal mineralization

139

Alkaline phosphatase in osteoblasts is down-regulated by pulsatile fluid flow  

Science.gov (United States)

It is our hypothesis that interstitial fluid flow plays a role in the bone remodeling response to mechanical loading. The fluid flow-induced expression of three proteins (collagen, osteopontin, and alkaline phosphatase) involved in bone remodeling was investigated. Rat calvarial osteoblasts subjected to pulsatile fluid flow at an average shear stress of 5 dyne/cm2 showed decreased alkaline phosphatase (AP) mRNA expression after only 1 hour of flow. After 3 hours of flow, AP mRNA levels had decreased to 30% of stationary control levels and remained at this level for an additional 5 hours of flow. Steady flow (4 dyne/cm2 fluid shear stress), in contrast, resulted in a delayed and less dramatic decrease in AP mRNA expression to 63% of control levels after 8 hours of flow. The reduced AP mRNA expression under pulsatile flow conditions was followed by reduced AP enzyme activity after 24 hours. No changes in collagen or osteopontin mRNA expression were detected over 8 hours of pulsatile flow. This is the first time fluid flow has been shown to affect gene expression in osteoblasts.

Hillsley, M. V.; Frangos, J. A.

1997-01-01

140

Biocompatibility and Alkaline Phosphatase Activity of Phosphorylated Chitooligosaccharides on the Osteosarcoma MG63 Cell Line  

Directory of Open Access Journals (Sweden)

Full Text Available Phosphorylated chitooligosaccharides (P-COS were prepared using a H3PO4, P2O5, Et3PO4 and hexanol solvent system. The P-COS were characterized by Fourier Transform Infrared Spectroscopy (FT-IR, Thermo gravimetric-Differential Thermal Analyzer (TG-DTA, 13C NMR, 31P NMR, X-ray diffraction analysis, solubility studies, biocompatibility and Alkaline Phosphatase Activity (ALP. The results reveal that phosphorylation occurred at the C3 and C6 position of OH groups and the C2 position of NH2 group. FT-IR confirmed no decomposition in pyranose ring in P-COS even with heating and treatment in acidic conditions. The amorphous nature of P-COS was confirmed by X-ray diffraction analysis. Further, the biocompatibility and alkaline phosphatase activity of P-COS were checked against the osteosarcoma MG63 cell line at different concentrations and no cytotoxicity was observed. After 12 h and 24 h of incubation, the ALP activity of P-COS was higher compared with the control group. These results suggest that P-COS is a biocompatible material and in future P-COS could open up a number of promising pharmaceutical and clinical applications to mankind.

Jayachandran Venkatesan

2010-10-01

 
 
 
 
141

Inhibitors of tissue-nonspecific alkaline phosphatase: design, synthesis, kinetics, biomineralization and cellular tests.  

Science.gov (United States)

Chronic kidney disease (CKD) is associated with numerous metabolic and endocrine disturbances, including abnormalities of calcium and phosphate metabolism and an inflammatory syndrome. The latter occurs early in the course of CKD and contributes to the development and progression of vascular calcification. A few therapeutic strategies are today contemplated to target vascular calcification in patients with CKD: vitamin K2, calcimimetics and phosphate binders. However, none has provided complete prevention of vascular calcification and there is an urgent need for alternate efficient treatments. Recent findings indicate that tissue-nonspecific alkaline phosphatase (TNAP) may represent a very promising drug target due to its participation in mineralization by vascular smooth muscle cells. We report the synthesis of four levamisole derivatives having better inhibition property on TNAP than levamisole. Their IC50, Ki and water solubility have been determined. We found that the four inhibitors bind to TNAP in an uncompetitive manner and are selective to TNAP. Indeed, they do not inhibit intestinal and placental alkaline phosphatases. Survival MTT tests on human MG-63 and Saos-2 osteoblast-like cells have been performed in the presence of inhibitors. All the inhibitors are not toxic at concentrations that block TNAP activity. Moreover, they are able to significantly reduce mineralization in MG63 and Saos-2 osteoblast-like cells, indicating that they are promising molecules to prevent vascular calcification. PMID:24183741

Debray, Julien; Chang, Lei; Marquès, Stéphanie; Pellet-Rostaing, Stéphane; Le Duy, Do; Mebarek, Saida; Buchet, René; Magne, David; Popowycz, Florence; Lemaire, Marc

2013-12-15

142

Expression of opioid receptors in osteoblast-like MG-63 cells, and effects of different opioid agonists on alkaline phosphatase and osteocalcin secretion by these cells.  

Science.gov (United States)

We have previously shown that several stressful situations associated with tissue injury determine a decrease in serum osteocalcin concentration. Since reduced osteocalcin production is a marker of decreased osteoblastic activity, this finding could be related to the pathogenesis of osteoporosis secondary to some diseases. Endogenous opioids are involved in stress response. Proenkephalin-derived peptides have been shown to inhibit alkaline phosphatase activity, another marker of bone formation, in the murine cell line ROS-17/2.8. On the other hand, serum osteocalcin has been reported as being low in heroin abusers. We have therefore thought it of interest to study the presence of opioid receptors in the human osteoblast-like cell line MG-63, and to evaluate the effects of different opioid agonists on the secretion of alkaline phosphatase and osteocalcin by these cells. The presence of opioid receptors was studied by means of RT-PCR and immunohistochemistry. RT-PCR studies suggested the presence of specific mRNA for the three types of receptors, and immunohistochemistry clearly showed their occurrence. Osteocalcin synthesis was significantly inhibited by high concentrations of the mu agonists morphine and (D-Ala(2), N-MePhe(4),Gly(5)-ol)-enkephalin although no changes were seen with the delta agonist (D-Ala(2),D-leu(5))-enkephalin. Morphine-induced osteocalcin inhibition was abolished when osteoblastic cells were incubated simultaneously with naloxone, whereas it was potentiated when cells were preincubated with naloxone. None of the opioid agonists modified the secretion of alkaline phosphatase. In conclusion, human osteoblast-like cells MG-63 express the three types of opioid receptors. Endogenous opioids may be involved in the reduction of osteocalcin observed in stressful situations associated with tissue injury. PMID:11025413

Pérez-Castrillón, J L; Olmos, J M; Gómez, J J; Barrallo, A; Riancho, J A; Perera, L; Valero, C; Amado, J A; González-Macías, J

2000-09-01

143

Endothelial alkaline phosphatase activity loss as an early stage in the development of radiation-induced heart disease in rats  

International Nuclear Information System (INIS)

Alkaline phosphatase activity of capillary endothelial cells in the heart of Wistar and Sprague-Dawley rats was studied sequentially after single doses of 10, 15, 20, or 25 Gy. Following irradiation capillary density and alkaline phosphatase activity were focally lost before myocardial degeneration or clinical symptoms of heart disease developed. Recovery from both changes took place after doses of 10 or 15 Gy. The decrease in capillary density and enzyme activity showed the same strain difference in latency times and in the extent of the lesions as previously described for pathological and clinical signs of heart disease

144

Cloning and Expression of the Alkaline Phosphatase Gene from the Persian Type Culture Collection Escherichia coli K-12  

Directory of Open Access Journals (Sweden)

Full Text Available The structural gene for alkaline phosphatase (phoA of E. coli K-12 strain obtained from the Persian type culture collection (PTCC 1268 was cloned into pTZ57R plasmid as cloning vector and pGEM-3Z plasmid as expression vector, respectively. The recombinant plasmids were confirmed by different restriction enzymes and determination of the nucleotide sequence. Protein expression was induced by isopropyl -D thiogalactopyranoside (IPTG and was analyzed using polyacrylamide gel electrophoresis (PAGE. The obtained results demonstrate a complete homology of the DNA sequence between the cloned alkaline phosphatase gene with the sequence present in the gene banks.

Hamid Mir Mohammad Sadeghi

2005-01-01

145

Purification and characterization of an alkaline phosphatase induced by phosphorus starvation in common bean (Phaseolus vulgaris L.) roots  

International Nuclear Information System (INIS)

Two phosphatase isoforms from roots of the common bean (Phaseolus vulgaris L.) showed an increase in activity in response to phosphate deficiency. One of them (APIII) was chosen for further purification through ionic exchange chromatography and preparative electrophoresis. The estimated molecular mass of APIII was 35 kDa by both SDS-Page and gel filtration analyses, suggesting a monomeric form of the active enzyme. The phosphatase was classified as an alkaline phosphatase based on the requirement of ph 8 for optimum catalysis. It not only exhibited broad substrate specificity, with the most activity against pyrophosphate, but also effectively catalyzed the hydrolysis of polyphosphate, glucose-1-phosphate and phospho enol-pyruvate. Activity was completely inhibited by molybdate, vanadate and phosphate but was only partially inhibited by fluoride. Although divalent cations were not essential for the pyro phosphatase activity of this enzyme, the hydrolysis of pyro phosphatase increased substantially in the presence of Mg2+.

146

Purification and characterization of an alkaline phosphatase induced by phosphorus starvation in common bean (Phaseolus vulgaris L.) roots  

Energy Technology Data Exchange (ETDEWEB)

Two phosphatase isoforms from roots of the common bean (Phaseolus vulgaris L.) showed an increase in activity in response to phosphate deficiency. One of them (APIII) was chosen for further purification through ionic exchange chromatography and preparative electrophoresis. The estimated molecular mass of APIII was 35 kDa by both SDS-Page and gel filtration analyses, suggesting a monomeric form of the active enzyme. The phosphatase was classified as an alkaline phosphatase based on the requirement of ph 8 for optimum catalysis. It not only exhibited broad substrate specificity, with the most activity against pyrophosphate, but also effectively catalyzed the hydrolysis of polyphosphate, glucose-1-phosphate and phospho enol-pyruvate. Activity was completely inhibited by molybdate, vanadate and phosphate but was only partially inhibited by fluoride. Although divalent cations were not essential for the pyro phosphatase activity of this enzyme, the hydrolysis of pyro phosphatase increased substantially in the presence of Mg{sup 2+}.

Morales, L.; Gutierrez, N.; Maya, V.; Parra, C.; Martinez B, E.; Coello, P., E-mail: pcoello@servidor.unam.mx [UNAM, Facultad de Quimica, Departamento de Bioquimica, Ciudad Universitaria, 04510 Mexico D. F. (Mexico)

2012-07-01

147

Improvement of Student Understanding of How Kinetic Data Facilitates the Determination of Amino Acid Catalytic Function through an Alkaline Phosphatase Structure/Mechanism Bioinformatics Exercise  

Science.gov (United States)

Laboratory exercises, which utilize alkaline phosphatase as a model enzyme, have been developed and used extensively in undergraduate biochemistry courses to illustrate enzyme steady-state kinetics. A bioinformatics laboratory exercise for the biochemistry laboratory, which complements the traditional alkaline phosphatase kinetics exercise, was…

Grunwald, Sandra K.; Krueger, Katherine J.

2008-01-01

148

Expression, secretion and surface display of a human alkaline phosphatase by the ciliate Tetrahymena thermophila  

Directory of Open Access Journals (Sweden)

Full Text Available Abstract Background Tetrahymena thermophila possesses many attributes that render it an attractive host for the expression of recombinant proteins. Surface proteins from the parasites Ichthyophthirius multifiliis and Plasmodium falciparum and avian influenza virus antigen H5N1 were displayed on the cell membrane of this ciliate. Furthermore, it has been demonstrated that T. thermophila is also able to produce a functional human DNase I. The present study investigates the heterologous expression of the functional human intestinal alkaline phosphatase (hiAP using T. thermophila and thereby presents a powerful tool for the optimization of the ciliate-based expression system. Results Functional and full length human intestinal alkaline phosphatase was expressed by T. thermophila using a codon-adapted gene containing the native signal-peptide and GPI (Glycosylphosphatidylinositol anchor attachment signal. HiAP activity in the cell extract of transformants suggested that the hiAP gene was successfully expressed. Furthermore, it was demonstrated that the enzyme was modified with N-glycosylation and localized on the surface membrane by the C-terminal GPI anchor. A C-terminally truncated version of hiAP lacking the GPI anchor signal peptide was secreted into the medium as an active enzyme. In a first approach to establish a high level expression system up to 14,000 U/liter were produced in a time frame of two days, which exceeds the production rate of other published expression systems for this enzyme. Conclusions With the expression of hiAP, not only a protein of commercial interest could be produced, but also a reporter enzyme that offers the possibility to analyze T. thermophila genes that play a role in the regulation of protein secretion. Additionally, the fact that ciliates do not secrete an endogenous alkaline phosphatase provides the possibility to use the truncated hiAP as a reporter enzyme, allowing the quantification of measures that will be necessary for further optimization of the host strains and the fermentation processes.

Herrmann Lutz

2011-01-01

149

[Cytotoxicity test based on luminescent assay of alkaline phosphatase released from target cells].  

Science.gov (United States)

Assay of 51Cr release from target cells has been commonly used in various methods of examining the cytotoxic properties of lymphocytes. In this paper a non-isotopic assay of cytotoxicity based on the leak of endogenous alkaline phosphatase (AIP) in target cells, is described. Enzyme activities were assayed by the luminescence on hydrolysis of the lumigen-PPD substrate. P3-X63-Ag8-U1 (P3U-1) cells were demonstrated to contain AIP and proved sensitive to the IL-2-induced killer lymphocytes, while no AIP activity was detected in human effector lymphocytes. Comparative studies of the test with 51Cr- and AIP-release in P3U-1 target cells were carried out, and the results obtained suggested that the AIP release test is useful as a new, simple lymphocyte cytotoxicity test. PMID:7996714

Kasatori, N; Urayama, T; Mori, T; Ishikawa, F

1994-10-01

150

Alkaline Phosphatase and CD34 Reaction of Deciduous Teeth Pulp Stem Cells  

Directory of Open Access Journals (Sweden)

Full Text Available Endothelial progenitor cells from the pulp of milk teeth were isolated for use in clinical applications and tissue engineering. Normal deciduous teeth from children of 7 to 8 years of age, which more than half the tooth root was extracted, were selected from the dental centre. Cells from enzyme treated pulps were cultured and cells resulting from the fifth and eight subculture were combined for cell surface marker determination experiments. Cells were positive for CD34 marker with a total of 99/45%, determined by flowcytometry. Cells also demonstrated alkaline phosphatase (ALP activity. From the developmental point of view, stem cells from the dental pulp seem to have derived from the neural crest, which our findings technically support this theory. In essence mobile progenitor cells from bone marrow of endothelial origin could also play a significant role in the derivation of dental pulp stem cells.

Fatemeh Abedini

2007-01-01

151

Ethnic differences in pre-adipocyte intracellular lipid accumulation and alkaline phosphatase activity.  

Science.gov (United States)

Alkaline phosphatase (ALP) increases lipid accumulation in human pre-adipocytes. This study was performed to assess whether ethnic differences in the prevalence of obesity in African and European females are related to differences in pre-adipocyte lipid accretion and ALP activity. Pre-adipocytes were isolated from 13 black and 14 white females. Adipogenesis was quantified using the lipid dye, Oil red O, whilst ALP activity was assayed in cell extracts on day zero and 12days after initiating adipogenesis. Lipid levels (OD units/mg protein) were lower in pre-adipocytes from white than black females on day 0 (0.36±0.05 versus 0.44±0.03, respectively; pblack compared to white females which parallels the obesity prevalence rates in these population groups. The reason for higher fat accumulation in pre-adipocytes isolated from black females may be related to higher ALP activity. PMID:25281857

Ali, Aus T; Chirambo, George; Penny, Clement; Paiker, Janice E; Ikram, Faisel; Psaras, George; Crowther, Nigel J

2015-01-01

152

Near-infrared fluorescence probe for the determination of alkaline phosphatase.  

Science.gov (United States)

Water-soluble CuInS2 quantum dots (QDs) were directly synthesized in an aqueous solution with mercaptopropionic acid (MPA) as stabilizers, and were functionalized using tryptophan molecules to form tryptophan-functionalized CuInS2 QDs (W-CuInS2 QDs) that had a strong fluorescence emission around 689 nm. The fluorescence of W-CuInS2 QDs could be quenched by Cu(2+) and then the addition of pyrophosphate (PPi) could effectively turn on the quenched fluorescence due to the strong interaction between Cu(2+) and PPi. The alkaline phosphatase (ALP) could catalyze the hydrolysis of PPi that would disassemble the complex of PPi-Cu(2+)-PPi. Therefore, the recovered fluorescence could be quenched again by the addition of ALP. In this paper, we developed a novel near-infrared fluorescence probe for the simple and convenient assay of ALP. PMID:24388906

Liu, Siyu; Pang, Shu; Na, Weidan; Su, Xingguang

2014-05-15

153

[Alkaline phosphatase isoenzymes. Selection of the seperation methods and their diagnostic value].  

Science.gov (United States)

Polyacrylamide gel electrophoresis (PAGE) of alkaline phosphatase (AP) was performed in 116 patients with elevated AP and in 36 controls. Results were compared with clinical data and with the heat inactivation techniques for AP isoenzyme separation. This latter method is simple but approximate; it is without value in the case of coexistent bone and liver pathology. PAGE gave a clear separation of liver, bile, bone, and intestinal AP isoenzymes. Disagreement with clinical data was rare (6 out of 86) and occurred in cases of associated bone and liver pathology (with 1 exception only), where one of these two abnormalities was not detected. Heat inactivation is useful as a screening test. If the cause of the elevated AP remains unknown after a few simple examinations, PAGE should then be utilized. Although more time-consuming, it is certainly more effective in the separation of AP isoenzymes. PMID:1153982

Balant, L; Fabre, J; Jung, A; Rosenbusch, C A

1975-05-10

154

Ratiometric fluorescent probe for alkaline phosphatase based on betaine-modified polyethylenimine via excimer/monomer conversion.  

Science.gov (United States)

Alkaline phosphatase (ALP) is an important diagnostic indicator for a number of human diseases since abnormal level of ALP is closely related to a variety of pathological processes; hence, the development of convenient and reliable assay methods for monitoring ALP is of great significance for medical sciences as well as biological diagnostics. Herein, we report the first ratiometric fluorescent sensing system for ALP. This sensing system consists of two components: the betaine-modified and positively charged polyethylenimine (PEI) and the negatively charged pyrene derivative containing one ALP-responsive phosphate group (Py-P, an aliphatic phosphate ester). In the absence of ALP, the two-component sensing system shows the excimer's emission of Py-P, since Py-P molecules complex with the positively charged polyelectrolyte via electrostatic interactions, leading to the formation of pyrene excimers. While in the presence of ALP, the phosphate moieties are cleaved from Py-P molecules due to the enzymatic reaction, thereby destroying the electrostatic interactions; as a result, the system displays the monomer emission of Py-P. This assay system is operable in aqueous media with a very low detection limit of 0.1 U/mL. The system is capable of detecting ALP in such biological fluid as serum, and this strategy may provide a new and effective approach for designing ratiometric sensing systems for detecting other biomolecules. PMID:25211600

Zheng, Fangyuan; Guo, Sihua; Zeng, Fang; Li, Jun; Wu, Shuizhu

2014-10-01

155

Fluorescent light-up probe with aggregation-induced emission characteristics for alkaline phosphatase sensing and activity study.  

Science.gov (United States)

Fluorogens with aggregation-induced emission (AIE) characteristics have attracted intensified research interest in biosensing applications, and those with specific targeting ability are especially desirable. In this work, we designed and synthesized an AIE fluorescent probe by functionalizing a tetraphenylethylene (TPE) fluorogen with two phosphate groups (TPE-phos) for the detection of alkaline phosphatase (ALP) and its enzymatic activity based on the specific interaction between the probe and ALP. The probe is virtually nonfluorescent in aqueous media due to good water solubility. In the presence of ALP, the phosphate groups are cleaved through enzymatic hydrolysis, yielding a highly fluorescent product as a result of activated AIE process. This light-up probe shows excellent selectivity toward ALP among a group of proteins. The detection limit is found to be 11.4 pM or 0.2 U L(-1) in Tris buffer solution with a linear quantification range of 3-526 U L(-1). The assay is also successfully performed in diluted serum with a linear range up to 175 U L(-1), demonstrating its potential application in clinical analysis of ALP levels in real samples. Furthermore, by conducting kinetic analysis of the enzyme using TPE-phos as the substrate, the kinetic parameter kcat/KM is determined to be 5.1×10(5) M(-1) s(-1), indicating a high efficiency of the substrate. PMID:23957823

Liang, Jing; Kwok, Ryan Tsz Kin; Shi, Haibin; Tang, Ben Zhong; Liu, Bin

2013-09-11

156

Reduced activity of alkaline phosphatase due to host-guest interactions with humic superstructures.  

Science.gov (United States)

Nuclear Magnetic Resonance (NMR) spectroscopy was applied to directly study the interactions between the alkaline phosphatase enzyme (AP) and two different humic acids from a volcanic soil (HA-V) and a Lignite deposit (HA-L). Addition of humic matter to enzyme solutions caused signals broadening in (1)H-NMR spectra, and progressive decrease and increase of enzyme relaxation (T1 and T2) and correlation (?C) times, respectively. Spectroscopic changes were explained with formation of ever larger weakly-bound humic-enzyme complexes, whose translational and rotational motion was increasingly restricted. NMR diffusion experiments also showed that the AP diffusive properties were progressively reduced with formation of large humic-enzyme complexes. The more hydrophobic HA-L affected spectral changes more than the more hydrophilic HA-V. (1)H-NMR spectra also showed the effect of progressively greater humic-enzyme complexes on the hydrolysis of an enzyme substrate, the 4-nitrophenyl phosphate disodium salt hexahydrate (p-NPP). While AP catalysis concomitantly decreased NMR signals of p-NPP and increased those of nitrophenol, addition of humic matter progressively and significantly slowed down the rate of change for these signals. In agreement with the observed spectral changes, the AP catalytic activity was more largely inhibited by HA-L than by HA-V. Contrary to previous studies, in which humic-enzyme interactions were only indirectly assumed from changes in spectrophotometric behavior of enzyme substrates, the direct measurements of AP behavior by NMR spectroscopy indicated that humic materials formed weakly-bound host-guest complexes with alkaline phosphatase, and the enzyme catalytic activity was thereby significantly inhibited. These results suggest that the role of extracellular enzymes in soils may be considerably reduced when they come in contact with organic matter dissolved in the soil solution. PMID:23953249

Mazzei, Pierluigi; Oschkinat, Hartmut; Piccolo, Alessandro

2013-11-01

157

Alkaline phosphatase activity in the western English Channel: Elevations induced by high summertime rainfall  

Science.gov (United States)

Alkaline phosphatase activity (APA) was determined in bulk particulate material and in a single-cell (ELF) assay at station L4 in the western English Channel during the summer of 2007. Throughout this period, the UK experienced its heaviest summertime rainfall since records began in 1914; with the result that riverine run-off into coastal waters was also elevated relative to long-term averages. Between May and August 2007, three distinct periods of elevated river run-off were observed which resulted in salinity minima at L4 on days 141, 190 and 232. An extended period of high river run-off between days 170 and 210 was responsible for decreases in near-surface salinity at L4 from 35.2068 to a minimum on day 190 of 34.7422. This contributed to the development of haline stratification which supported the development of an intense bloom of the centric diatom Chaetoceros debelis, with maximum observed chlorophyll a concentration of 8.69 ?g l -1. Minima in salinity, and maxima in chlorophyll concentration on day 190 were coincident with a peak in river-derived dissolved inorganic nitrogen (DIN) of 1.9 ?mol l -1 which was >5 times greater than the summertime mean and 24 times the concentrations experienced at L4 on weeks immediately before and after. There was no accompanying increase in dissolved inorganic phosphorus (DIP), and the DIN:DIP ratio increased to 49. With the inherent phosphorus stress that this caused, rates of APA increased from <4 to 42.4 nmolP l -1 h -1. ELF analysis on day 197 identified two taxa actively expressing alkaline phosphatase: the dinoflagellate Prorocentrum micans and ciliate Tiarana sp.

Rees, Andrew P.; Hope, Sam B.; Widdicombe, Claire E.; Dixon, Joanna L.; Woodward, E. Malcolm S.; Fitzsimons, Mark F.

2009-03-01

158

Properties of Na+/K+ ATPase and alkaline phosphatase alter during spontaneous and radiation-induced leukemogenesis in mice  

International Nuclear Information System (INIS)

Properties are characterized of Na+/K+ ATPase and alkaline phosphatase in thymocytes or thymoblasts from mice of two strains: AKR in which thymoma developed spontaneously, and C57Bl in which the development was induced by X-irradiation (total dose: 5.4 Gy in 3 fractions). It was found that before thymoma could be discerned morphologically the properties of the two enzymes changed. There was a decrease in 86Rb uptake and in the rate of ATP hydrolysis per cell (both strains) as well as an increase in alkaline phosphatase activity per cell (C57Bl mice). In both spontaneous and radiation-induced thymomas 86Rb uptake, ATP hydrolysis and 3H-ouabain binding per cell were higher than in normal thymuses. Likewise, alkaline phosphatase activity per cell was higher in the thymomas than in the thymuses; this increase was accompanied by the appearance of additional isoenzyme(s) (1 in AKR, 2 in C57Bl). These changes were compared with cAMP content and 3H-thymidine incorporation, taken as indicators of the proliferative activity, and their high correlation in both AKR and C57Bl mice allowed to distinguish a pre-leukemic period. In that period thymoblasts clearly differed from the normal ones in Na+/K+ ATPase and alkaline phosphatase properties as well as proliferation, although the morphology of the thymus was still unchanged. (author)

159

The effect of 50 kV X-ray irradiation on the alkaline phosphatase activity of growing rat bone  

International Nuclear Information System (INIS)

Alkaline phosphatase activity was decreased in tibial metaphysis of growing rats on the first day after 50 kV x-irradiation with 0.5-8.0 Gy. There were no differences in enzyme activity between the control and the irradiated metaphysis 30 days after irradiation. (author)

160

Pst I restriction fragment length polymorphism of the human placental alkaline phosphatase gene in normal placentae and tumors  

International Nuclear Information System (INIS)

The structure of the human placental alkaline phosphatase gene from normal term placentae was studied by restriction enzyme digestion and Southern blot analysis using a cDNA probe to the gene for the placental enzyme. The DNA digests fall into three distinct patterns based on the presence and intensity of an extra 1.1-kilobase Pst I Band. The extra 1.1-kilobase band is present in 9 of 27 placenta samples, and in 1 of these samples the extra band is present at double intensity. No polymorphism was revealed by digestion with restriction enzymes EcoRI, Sma I, BamHI, or Sac I. The extra Pst I-digestion site may lie in a noncoding region of the gene because no correlation was observed between the restriction fragment length polymorphism and the common placental alkaline phosphatase alleles identified by starch gel electrophoresis. In addition, because placental alkaline phosphatase is frequently re-expressed in neoplasms, the authors examined tissue from ovarian, testicular, and endometrial tumors and from BeWo choriocarcinoma cells in culture. The Pst I-DNA digestion patterns from these cells and tissues were identical to those seen in the normal ovary and term placentae. The consistent reproducible digestion patterns seen in DNA from normal and tumor tissue indicate that a major gene rearrangement is not the basis for the ectopic expression of placental alkaline phosphatase in neoplasia

 
 
 
 
161

Changes in Bone Alkaline Phosphatase and Procollagen Type-1 C-Peptide after Static and Dynamic Exercises  

Science.gov (United States)

We investigated the effects of two types of nonweight-bearing exercise on changes in bone-specific alkaline phosphatase (BAP) and pro-collagen type 1 C-peptide (P1P). BAP is a specific marker of bone synthesis, whereas P1P reflects synthesis of type 1 collagen in other organs as well as bone. Eight participants performed static and dynamic…

Kubo, Keitaro; Yuki, Kazuhito; Ikebukuro, Toshihiro

2012-01-01

162

Leucine Aminopeptidase, ?-Glucosidase and Alkaline Phosphatase Activity Rates and Their Significance in Nutrient Cycles in Some Coastal Mediterranean Sites  

Digital Repository Infrastructure Vision for European Research (DRIVER)

In aquatic microbial ecology, knowledge of the processes involved in the turnover of organic matter is of utmost importance to understand ecosystem functioning. Microorganisms are major players in the cycling of nutrients (nitrogen, phosphorus) and carbon, thanks to their enzymatic activities (leucine aminopeptidase, LAP, alkaline phosphatase, AP, and ?-glucosidase, ?-GLU) on organic polymers (proteins, organic phosphates and polysaccharides, respectively). Estimates of the decomposition ra...

Gabriella Caruso

2010-01-01

163

Evaluation of non-specific tissue alkaline phosphatase on bone samples from traditional and piezoelectric osteotomy.  

Science.gov (United States)

The aim of this study is to test the response of bone during cutting actions in dental procedures by sampling alkaline phosphatase (ALP) as a biological reference marker. ALP is found abundantly in bone tissue. In the first series of experiments a temporal-minimum quantity of ALP enzyme response was recorded, the observed period was 40 minutes. The ALP samples treated with piezoelectric surgery showed a rapid increase, with peak at 30 min, and then declined rapidly within the next 10 minutes. A second experiment was performed to evaluate 4 cutting instruments: drill bits high speed turbine (T1); drill bits contra-angle (T2) Piezoelectric insertions (T3), and manual instruments (controls). This second experiment was to evaluate the ALP activity at 30 min. The T1 samples produced the highest results (3,66499 +- 0,51394); control groups had a lower response (0,72793 +- 0,22353), while the T2 group produced statistically significant higher results (2,77793 +- 0,40553) than T3 (1,16608 +- 0,32676). The different values obtained for ALP in these two experiments for a short period of time (30 min) cannot be interpreted as a response of bone tissue regeneration subjected to surgical trauma. The MINIMUM trauma caused by the surgical piezoelectric instruments, in respect to conventional surgical instruments is clearly evident from the phosphatase inflammatory activity. PMID:18187021

Perfetti, G; Calderini, M; Berardi, D; Leoni, S; Ferrante, M; Spoto, G

2006-01-01

164

Streptolysin-O induces release of glycosylphosphatidylinositol-anchored alkaline phosphatase from ROS cells by vesiculation independently of phospholipase action.  

Science.gov (United States)

Streptolysin-O (SLO), a cholesterol-binding agent, was used for studies on the release of glycosylphosphatidylinositol (GPI)-anchored alkaline phosphatase (AP) from ROS cells. Treatment of cells with SLO resulted in a time- and concentration-dependent release of AP into the extracellular medium. This release was potentiated by Ca2+ and bovine serum, but not by GPI-specific phospholipase D (GPI-PLD) purified from bovine serum. The released AP distributed to the detergent phase after Triton X-114 phase separation. This result suggested that the released AP contained an intact GPI anchor, and thus both proteolysis and anchor degradation by anchor-specific hydrolases, including GPI-PLD, as the potential mechanisms for SLO-mediated AP release were ruled out. The released AP sedimented at 100,000 g. A substantial amount of lipids was detected in the 100,000 g pellet. Cholesterol and sphingomyelin were enriched in SLO-released material, compared with intact cells. These results were consistent with vesiculation as the mechanism for SLO induction of AP release. Two other cholesterol-binding agents, saponin and digitonin, were also able to release AP, possibly by a similar vesiculation mechanism, whereas others, including nystatin, filipin and beta-escin, failed to elicit any AP release. Eight GPI-anchored proteins were identified in ROS cells, and all were substantially enriched in the vesicles released by SLO. Taken together, these results do not provide any support for the hypothesis that the clustering of GPI-anchored proteins in the plasma membrane is responsible for their resistance to GPI-PLD cleavage. Images Figure 5 Figure 6 PMID:7832771

Xie, M; Low, M G

1995-01-01

165

Biological Apatite Formed from Polyphosphate and Alkaline Phosphatase May Exchange Oxygen Isotopes from Water through Carbonate  

Science.gov (United States)

The oxygen isotopic composition in bone mineral phosphate is known to reflect the local water composition, environmental humidity, and diet1. Once ingested, biochemical processes presumably equilibrate PO43- with "body water" by the many biochemical reactions involving PO43- 2. Blake et al. demonstrated that enzymatic release of PO43- from organophosphorus compounds, and microbial metabolism of dissolved orthophosphate, significantly exchange the oxygen in precipitated apatite within environmental water3,4, which otherwise does not exchange with water at low temperatures. One of the enzymes that can cleave phosphates from organic substrates is alkaline phosphastase5, the enzyme also associated with bone mineralization. The literature often states that the mineral in bone in hydroxylapatite, however the mineral in bone is carbonated apatite that also contains some fluoride6. Deprotonation of HPO32- occurs at pH 12, which is impossibly high for biological system, and the predominate carbonate species in solution at neutral pH is HCO3-. To produce an apatite mineral without a significant hydroxyl content, it is possible that apatite biomineralization occurs through a polyphosphate pathway, where the oxygen atom required to transform polyphosphate into individual phosphate ions is from carbonate: [PO3-]n + CO32- -> [PO3-]n-1 + PO43- + CO2. Alkaline phosphatase can depolymerise polyphosphate into orthophosphate5. If alkaline phosphatase cleaves an oxygen atom from a calcium-carbonate complex, then there is no requirement for removing a hydrogen atom from the HCO3- or HPO43- ions of body water to form bioapatite. A mix of 1 mL of 1 M calcium polyphosphate hydogel, or nano-particles of calcium polyphosphate, and amorphous calcium carbonate were reacted with alkaline phosphatase, and maintained at neutral to basic pH. After two weeks, carbonated apatite and other calcium phosphate minerals were identified by powder x-ray diffraction. Orthophosphate and unreacted polyphosphate species were detected by Raman and IR spectroscopy. The oxygen isotope data of the reactants and products will also be presented. The possibility that carbonate acts as an intermediate reagent, transferring the oxygen from water to phosphate in biological apatite mineral formation may explain why biological apatite exhibits a significant carbonate content, and how this mineral is formed with an insignificant hydroxyl content. 1 Kohn, M.J., and Cerling, T.E. Rev Mineral Geochem 2002 (48) 455 2 Kolodny, Y., Luz, B., Navon, O. Earth Planet Sci Lett 1983 (64) 398 3 Blake, R.E., O'Neil, J.R., Garcia, G.A. Geochim et Cosmochim Acta 1997 (61) 4411 4 Blake, R.E., Alt, J.C., and Martini, A.M. PNAS 2001 (98) 2148-2153 5 Liang, Y., and Blake, R.E. Geochim Cosmochim Acta 2009 (73) 3782) 6 Pasteris, J.D. et al. Biomaterials 2004 (35) 229 7 Omelon et al., PLoS ONE 2009 4(5), e5634

Omelon, S. J.; Stanley, S. Y.; Gorelikov, I.; Matsuura, N.

2011-12-01

166

Extracellular alkaline phosphatase is a sensitive marker for cellular stimulation and exocytosis in heterotroph cell cultures of Chenopodium rubrum.  

Science.gov (United States)

We investigated the response of extracellular phosphatase to heat shock in heterotrophic Chenopodium rubrum L. cell cultures. Surprisingly, in contrast to the generally used acid phosphatase, an extracellular alkaline phosphatase showed the most sensitive response. This phosphatase was characterized as a marker for cellular stimulation by its high correlations with induced changes of extracellular pH: 10microM nigericin (correlation coefficient r=0.91), 100microM salicylic acid (r=0.84), heat shock 5min 37 degrees C (r=0.79), and heat shock after pre-treatment with 5microM fusicoccin (r=0.92) or 0.5% ethanol (r=0.90). Cellular stimulation was estimated with concentrations of acids and bases, yielding similar levels of pH change (0.5 pH) in cell-free supernatant: salicylic acid (200microM), benzoic acid (600microM), HCl (140microM), NaOH (100microM), and KOH (100microM). The Golgi apparatus inhibitor Brefeldin A (200microM) reduced the heat-shock-induced phosphatase (-33%). The pH optimum of heat-shock-induced phosphatase was 3; however, there the proportion of constitutive phosphatase was higher than at pH 8-9.5, indicating different pH dependence of constitutive and induced activity. Thus, heat-shock-induced phosphatase was characterized by alkaline activity with inhibitors (10microM molybdate: -52%, 2.5mM phosphate: -64%, 10microM ZnCl(2): -82%), substrates (2.5mM, tyrosine phosphate: 255pkat g(-1), p-nitrophenyl phosphate: 92pkat g(-1), serine phosphate: 0, threonine phosphate: 0), Hill coefficient (nH=1.4) indicating two binding sites, and the extent of heat-shock stimulation (p-nitrophenyl phosphate: +190%, tyrosine phosphate: +180%). SDS-PAGE showed a correlation of alkaline phosphatase with the heat-shock-induced release of highly N-glycosylated 53kDa protein, detected by peroxidase-labeled concanavalin A affinoblotting after endoglycosidase H treatment. The 53kDa protein showed no in-gel phosphatase activity after SDS-PAGE and regeneration treatment, in contrast to a putative dimer (105kDa). PMID:18433930

Chaidee, Anchalee; Wongchai, Chatchawal; Pfeiffer, Wolfgang

2008-11-01

167

Improving Escherichia coli alkaline phosphatase efficacy by additional mutations inside and outside the catalytic pocket.  

Science.gov (United States)

We describe a strategy that allowed us to confer on a bacterial (E. coli) alkaline phosphatase (AP) the high catalytic activity of the mammalian enzyme while maintaining its high thermostability. First, we identified mutations, at positions other than those occupied by essential catalytic residues, which inactivate the bacterial enzyme without destroying its overall conformation. We transferred concomitantly into the bacterial enzyme four residues of the mammalian enzyme, two being in the catalytic pocket and two being outside. Second, the gene encoding the inactive mutant was submitted to random mutagenesis. Enzyme activity was restored upon the single mutation D330N, at a position that is 12 A away from the center of the catalytic pocket. Third, this mutation was combined with other mutations previously reported to increase AP activity slightly in the presence of magnesium. As a result, at pH 10.0 the phosphatase activity of both mutants D330N/D153H and D330N/D153G was 17-fold higher than that of the wild-type AP. Strikingly, although the two individual mutations D153H and D153G destabilize the enzyme, the double mutant D330N/D153G remained highly stable (T(m)=87 degrees C). Moreover, when combining the phosphatase and transferase activities, the catalytic activity of the mutant D330N/D153G increased 40-fold (k(cat)=3200 s-1) relative to that of the wild-type enzyme (k(cat)=80 s-1). Due to the simultaneous increase in K(m), the resulting k(cat)/K(m) value was only increased by a factor of two. Therefore, a single mutation occurring outside a catalytic pocket can dramatically control not only the activity of an enzyme, but also its thermostability. Preliminary crystallographic data of a covalent D330N/D153G enzyme-phosphate complex show that the phosphate group has significantly moved away from the catalytic pocket, relative to its position in the structure of another mutant previously reported. PMID:11828484

Muller, B H; Lamoure, C; Le Du, M H; Cattolico, L; Lajeunesse, E; Lemaître, F; Pearson, A; Ducancel, F; Ménez, A; Boulain, J C

2001-08-01

168

Structure and expression of rat osteosarcoma (ROS 17/2.8) alkaline phosphatase: product of a single copy gene.  

Digital Repository Infrastructure Vision for European Research (DRIVER)

Alkaline phosphatase [ALP; orthophosphoric-monoester phosphohydrolase (alkaline optimum), EC 3.1.3.1] is a ubiquitous enzyme of unknown function expressed at high levels in cells of mineralizing tissues. To study the structure, function, and expression of ALP, a full-length cDNA of rat ALP (2415 bases) was isolated from a ROS 17/2.8 osteosarcoma cell lambda gt10 cDNA library. The predicted amino acid sequence spans 524 residues and includes an N-terminal signal peptide of 17 amino acids, the ...

Thiede, M. A.; Yoon, K.; Golub, E. E.; Noda, M.; Rodan, G. A.

1988-01-01

169

Titanium dioxide nanotube films: Preparation, characterization and electrochemical biosensitivity towards alkaline phosphatase.  

Science.gov (United States)

Titania nanotubes (TNTs) were prepared by anodization on different substrates (titanium, Ti6Al4V and Ti6Al7Nb alloys) in ethylene glycol and glycerol. The influence of the applied potential and processing time on the nanotube diameter and length is analyzed. The as-formed nanotube layers are amorphous but they become crystalline when subjected to subsequent thermal treatment in air at 550°C; TNT layers grown on titanium and Ti6Al4V alloy substrates consist of anatase and rutile, while those grown on Ti6Al7Nb alloy consist only of anatase. The nanotube layers grown on Ti6Al7Nb alloy are less homogeneous, with supplementary islands of smaller diameter nanotubes, spread across the surface. Better adhesion and proliferation of osteoblasts was found for the nanotubes grown on all three substrates by comparison to an unprocessed titanium plate. The sensitivity towards bovine alkaline phosphatase was investigated mainly by electrochemical impedance spectroscopy in relation to the crystallinity, the diameter and the nature of the anodization electrolyte of the TNT/Ti samples. The measuring capacity of the annealed nanotubes of 50nm diameter grown in glycerol was demonstrated and the corresponding calibration curve was built for the concentration range of 0.005-0.1mg/mL. PMID:24582263

Roman, Ioan; Trusca, Roxana Doina; Soare, Maria-Laura; Fratila, Corneliu; Krasicka-Cydzik, Elzbieta; Stan, Miruna-Silvia; Dinischiotu, Anca

2014-04-01

170

Schistosoma mansoni: molecular characterization of Alkaline Phosphatase and expression patterns across life cycle stages.  

Science.gov (United States)

Here we describe the cloning and characterization of the Schistosoma mansoni Alkaline Phosphatase (SmAP), previously identified in the tegument of adult worms. SmAP encodes a complete sequence composed of 536 amino acids containing an N-terminal signal peptide, five N-glycosylation sites, and a GPI anchor signal, similar to that described for mammalian orthologs. Real-time RT-PCR and Western blot experiments suggest a rapid translation as soon as cercariae are transformed into schistosomula. Immunolocalization analysis shows that the protein is widely distributed in the worm tissues, with increased concentration in the vitelline glands of female parasites. Furthermore, the surface localization of this enzyme was quantitatively supported by its enzymatic activity in live ex vivo or cultured parasites throughout the life cycle stages. The fact that cercariae accumulate large amounts of SmAP mRNA, which rapidly translates into protein upon schistosomula transformation, indicates it may have an important role in host invasion. PMID:21784070

Araujo-Montoya, B O; Rofatto, H K; Tararam, C A; Farias, L P; Oliveira, K C; Verjovski-Almeida, S; Wilson, R A; Leite, L C C

2011-11-01

171

Growth and alkaline phosphatase activity of Chattonella marina and Heterosigma akashiwo in response to phosphorus limitation.  

Science.gov (United States)

The growth and alkaline phosphatase activity (APA) of two raphidophyceae species Chattonella marina and Heterosigma akashiwo were investigated in response to P-limitation and subsequent addition of dissolved inorganic phosphorus (DIP, NaH2PO4) and two dissolved organic phosphorus (DOP) compounds: guanosine 5-monophosphate (GMP) and triethyl phosphate (TEP). APA levels increased greatly after P-starvation as the decrease of the cellular phosphorus quotes (Qp). C. marina responded to P-limitation quickly and strongly, with 10-fold increase in APA within 24hr after P-starvation. The larger difference between maximal and minimal QP values in C. marina indicated its high capacity in P storage. APA of H. akashiwo was maximally enlarged about 2.5 times at 48hr of P-starvation. After the addition of nutrients, cell numbers of C. marina increased in all treatments including the P-free culture, demonstrating the higher endurance of C. marina to P-limitation. However, those of H. akashiwo increased only in DIP and GMP cultures. APA increased only after the addition of the monophosphate ester GMP. The results suggest that quick responses of C. marina to P-limitation, high capacity in P storage as well as endurance for P-depletion provide this species an ecological advantage in phytoplankton community competition under DIP-limited conditions. PMID:25662231

Wang, Zhao-Hui; Liang, Yu

2015-02-01

172

Alkaline phosphatase-positive cells isolated from human hearts have mesenchymal stem cell characteristics  

Directory of Open Access Journals (Sweden)

Full Text Available Tissue-specific resident cells have been identified as a promising population of progenitor cells for cell-based therapies. We describe here the isolation from adult human hearts of tissue nonspecific alkaline phosphatase-positive cells (ALPL+ cells with mesenchymal stem cell (MSC characteristics. Samples from 24 adult cadaveric donors were obtained from a valve bank. Mean total ischemia time was 21.5 ± 9.1 hours. The success rate for the isolation of human heart-derived cells by the explant culture technique was 70% for the right auricle (14 of 20 trials and 33% for the right ventricle (7 of 21 trials. The total auricle-derived cell population (TAD was used for the purification of ALPL+ cells. TAD and ALPL+ cells expressed markers for MSC and pericytes. TAD cells and ALPL+ cells differentiated into adipocytes, osteoblasts and chondroblasts, and ALPL+ cells expressed markers of these three lineages more strongly than TAD cells, as shown by RT-PCR. This population therefore has a greater potential for differentiation into mesechymal lineages than TAD cells. Both cell populations express some markers of cardiac progenitors, such as GATA4, CD117 and VEGF. ALPL+ cells expressed troponin T and ABCG2, which are also markers of the cardiac lineage. Heart samples from tissue banks could be considered as sources of MSC with putative commitment towards cardiac lineages, even after prolonged ischemia times.

Alessandra Melo de Aguiar

2011-10-01

173

Curcumin and Chronic Kidney Disease (CKD: Major Mode of Action through Stimulating Endogenous Intestinal Alkaline Phosphatase  

Directory of Open Access Journals (Sweden)

Full Text Available Curcumin, an active ingredient in the traditional herbal remedy and dietary spice turmeric (Curcuma longa, has significant anti-inflammatory properties. Chronic kidney disease (CKD, an inflammatory disease, can lead to end stage renal disease resulting in dialysis and transplant. Furthermore, it is frequently associated with other inflammatory disease such as diabetes and cardiovascular disorders. This review will focus on the clinically relevant inflammatory molecules that play a role in CKD and associated diseases. Various enzymes, transcription factors, growth factors modulate production and action of inflammatory molecules; curcumin can blunt the generation and action of these inflammatory molecules and ameliorate CKD as well as associated inflammatory disorders. Recent studies have shown that increased intestinal permeability results in the leakage of pro-inflammatory molecules (cytokines and lipopolysaccharides from gut into the circulation in diseases such as CKD, diabetes and atherosclerosis. This change in intestinal permeability is due to decreased expression of tight junction proteins and intestinal alkaline phosphatase (IAP. Curcumin increases the expression of IAP and tight junction proteins and corrects gut permeability. This action reduces the levels of circulatory inflammatory biomolecules. This effect of curcumin on intestine can explain why, despite poor bioavailability, curcumin has potential anti-inflammatory effects in vivo and beneficial effects on CKD.

Siddhartha S. Ghosh

2014-12-01

174

Rapid and highly sensitive electrochemical determination of alkaline phosphatase using a composite tyrosinase biosensor.  

Science.gov (United States)

The use of an amperometric graphite-Teflon composite tyrosinase biosensor for the rapid monitoring of alkaline phosphatase (ALP), with no need of an incubation step and using phenyl phosphate as the substrate, is reported. Phenol generated by the action of ALP is monitored at the tyrosinase composite electrode through the electrochemical reduction of the o-quinone produced to catechol, which produces a cycle between the tyrosinase substrate and the electroactive product, giving rise to the amplification of the biosensor response and to the sensitive detection of ALP. The current was measured at -0.10 V 5 min after the addition of ALP. As a compromise between high ALP activity and high sensitivity for the detection of phenol, a pH of 8.5 was chosen. The substrate concentration was also optimized. A linear calibration plot was obtained for ALP between 2.0 x 10(-13) and 2.5 x 10(-11), with a detection limit of 6.7 x 10(-14) M. Different types of milk were analyzed with good results, using an extremely simple and rapid procedure. PMID:15620894

Serra, B; Morales, M D; Reviejo, A J; Hall, E H; Pingarrón, J M

2005-01-15

175

Alkaline phosphatase variation during carfilzomib treatment is associated with best response in multiple myeloma patients.  

Science.gov (United States)

The ubiquitin-proteasome pathway regulates bone formation through osteoblast differentiation. We analyzed variation alkaline phosphatase (ALP) during carfilzomib treatment. Data from 38 patients enrolled in the PX-171-003 and 29 patients in PX-171-004 studies, for patients with relapsed/refractory myeloma, were analyzed. All patients received 20 mg/m(2) of carfilzomib on Days 1, 2, 8, 9, 15, and 16 of a 28-day cycle. Sixty-seven patients from ALP data were evaluable. In PX-171-003, the ORR (>PR) was 18% and the clinical benefit response (CBR; >MR) was 26%, while in PX-171-004, the ORR was 35.5% overall and 57% in bortezomib-naive patients. ALP increment from baseline was statistically different in patients who achieved ? VGPR compared with all others on Days 1 (P = 0.0049) and 8 (P = 0.006) of Cycle 2. In patients achieving a VGPR or better, ALP increased more than 15 units per liter at Cycle 2 Day 1 over baseline. An ALP increase over the same period of time was seen in 26%, 13% and 11% of patients achieving PR, MR, and SD, respectively. This retrospective analysis of patients with relapsed or refractory myeloma treated with single-agent carfilzomib indicates that early elevation in ALP is associated with subsequent myeloma response. PMID:21477075

Zangari, Maurizio; Aujay, Monette; Zhan, Fenghuang; Hetherington, Kristina L; Berno, Tamara; Vij, Ravi; Jagannath, Sundar; Siegel, David; Keith Stewart, A; Wang, Luhua; Orlowski, Robert Z; Belch, Andrew; Jakubowiak, Andrzej; Somlo, George; Trudel, Suzanne; Bahlis, Nizar; Lonial, Sagar; Singhal, Seema; Kukreti, Vishal; Tricot, Guido

2011-06-01

176

Enhanced catalysis by active-site mutagenesis at aspartic acid 153 in Escherichia coli alkaline phosphatase.  

Science.gov (United States)

Bacterial alkaline phosphatase catalyzes the hydrolysis and transphosphorylation of phosphate monoesters. Site-directed mutagenesis was used to change the active-site residue Asp-153 to Ala and Asn. In the wild-type enzyme Asp-153 forms a second-sphere complex with Mg2+. The activity of mutant enzymes D153N and D153A is dependent on the inclusion of Mg2+ in the assay buffer. The steady-state kinetic parameters of the D153N mutant display small enhancements, relative to wild type, in buffers containing 10 mM Mg2+. In contrast, the D153A mutation gives rise to a 6.3-fold increase in kcat, a 13.7-fold increase in kcat/Km (50 mM Tris, pH 8), and a 159-fold increase in Ki for Pi (1 M Tris, pH 8). In addition, the activity of D153A increases 25-fold as the pH is increased from 7 to 9. D153A hydrolyzes substrates with widely differing pKa's of their phenolic leaving groups (PNPP and DNPP), at similar rates. As with wild type, the rate-determining step takes place after the initial nucleophilic displacement (k2). The increase in kcat for the D153A mutant indicates that the rate of release of phosphate from the enzyme product complex (k4) has been enhanced. PMID:1525159

Matlin, A R; Kendall, D A; Carano, K S; Banzon, J A; Klecka, S B; Solomon, N M

1992-09-01

177

Effect of gallic acid on alkaline phosphatase and peptidase activities in rat intestine.  

Science.gov (United States)

Gallic acid is a normal constituent of many edible foods, thus directly interacts with epithelial tissue in intestine. In the present study, the effect of gallic acid on intestinal alkaline phosphatase (IAP) and peptidase activities in rat intestine was evaluated. Gallic acid (0.27-0.5 mM) inhibited activities of leucine aminopeptidase (LAP) and y-glutamyl transpeptidase (y-GTP) by over 90%, compared to controls in rat intestine. In contrast, 0.1-0.6 mM gallic acid either had no effect or stimulated the activity of IAP in rat intestine. The observed inhibition of peptidases by gallic acid was reversible in nature. Kinetic analysis revealed no change in Vmx of LAP (0.42-0.44 units/mg protein) and gamma-GTP (0.22-0.24 units/mg protein), while the values of apparent Km were increased 6-7 fold, exhibiting competitive-type of enzyme inhibition by gallic acid. The values of Ki for LAP and gamma-GTP were 0.037 mM and 0.017 mM, respectively. These observations indicate that gallic acid is a potent inhibitor of brush border peptidases, and thus may interfere in the digestion and absorption of proteins in the intestine. PMID:20027867

Mahajan, Nidhi; Mahmood, Akhtar

2009-10-01

178

Conversion of glycosylphosphatidylinositol (GPI)-anchored alkaline phosphatase by GPI-PLD.  

Science.gov (United States)

Enzymatic conversion of brain glycosylphosphatidylinositol-linked alkaline phosphatase (GPI-AP), amphiphilic, was examined. When GPI-AP was incubated with glycosylphosphatidylinositol-specific phospholipase D (GPI-PLD), a negligible conversion of GPI-AP to hydrophilic form was observed. The inclusion of monoacylglycerols enhanced the enzymatic conversion, although the action of monoacylglycerols differed greatly according to the size of acyl group; the enzymatic conversion was enhanced considerably in the presence of monoacylglycerols possessing acyl group of longer chain length (C10-C18), while monoacylglycerols with acyl moiety of shorter length (C4-C8) did fail to augment the enzymatic conversion. Noteworthy, monooleoylglycerol was much more effective than the other monoacylglycerols in promoting the enzymatic conversion, indicating a beneficial role of the unsaturation in acyl chain. Meanwhile, ionic amphiphiles such as monohexadecyllysophosphatidylcholine and palmitoyl-carnitine decreased the enzymatic conversion of GPI-AP in a concentration-dependent manner, with monohexadecyllysophosphatidylcholine being more inhibitory than palmitoylcarnitine. Separately, when GPI-AP was exposed to various oxidants prior to the incubation with GPI-PLD, a remarkable decrease of the enzymatic conversion was observed with hypochlorite and peroxynitrite generators, but not H2O2. In further study, hypochlorite was found to inactivate GPI-PLD at low concentrations (3 to approximately 100 microM). From these results, it is suggested that the enzymatic conversion of GPI-AP by GPI-PLD may be regulated in vivo system. PMID:10403126

Moon, Y G; Lee, H J; Kim, M R; Myung, P K; Park, S Y; Sok, D E

1999-06-01

179

Use of leucocyte alkaline phosphatase (LAP) score in differentiating malignant from benign paraproteinaemias.  

Science.gov (United States)

The leucocyte alkaline phosphatase (LAP) score of peripheral blood neutrophils was examined in 20 patients with multiple myeloma and compared with the score in 18 patients with monoclonal gammopathy of undetermined significance (MGUS). The mean (95% confidence limit) LAP score in those with multiple myeloma was 186 (169-218) compared with 92 (64-120) in the MGUS group. In the multiple myeloma group all but one patient had a high LAP score, irrespective of disease. No cause for raised LAP, such as infection, was present in any of the patients with multiple myeloma. In the MGUS group six patients had a raised LAP score; in two of them another cause for such a rise was present (autoimmune haemolytic anaemia and primary thrombocythaemia). In neither group did the LAP score correlate with duration of the disease, bone marrow plasma cell count, paraprotein concentration, haemoglobin, total white cell or neutrophil count. It is concluded that a normal LAP count in patients with paraproteinaemia suggests a benign condition, but a raised count does not indicate a malignant condition. PMID:1713223

Majumdar, G; Hunt, M; Singh, A K

1991-07-01

180

Identification of enterotoxigenic Escherichia coli with synthetic alkaline phosphatase-conjugated oligonucleotide DNA probes  

International Nuclear Information System (INIS)

Alkaline phosphatase-conjugated (AP) 26-base oligonucleotide DNA probes were compared with the same probes labeled with gamma-32P for the identification of heat-labile (LT) and heat-stable (ST) enterotoxigenic Escherichia coli (ETEC). The AP oligonucleotide probes were as sensitive as the radiolabeled (RL) probes in detecting LT and STA-2 target cell DNA, but the AP ST probe, which differed from STA-1 by two bases, was less sensitive than the RL probe in detecting STA-1 DNA (6.25 versus 0.78 ng). Of 94 ETEC that were identified with the RL probe, the AP probes detected 93% (28 of 30) of ST, 73% (25 of 34) of LT, and 67% (20 of 30) of LTST ETEC. When colony lysates of these ETEC were examined, the AP probes identified all 94 ETEC. In examinations of stool blots, the RL and AP probes were shown to have sensitivities of 71 and 59%, specificities of 91 and 86%, positive predictive values of 87 and 73%, and negative predictive values of 86 and 74%, respectively. AP oligonucleotide probes to detect ETEC were less sensitive in detecting ETEC by colony or stool blot hybridization than the RL probes but could be used by laboratories without access to radioisotopes to examine colony lysates

 
 
 
 
181

Reduced Levels of Membrane-Bound Alkaline Phosphatase Are Common to Lepidopteran Strains Resistant to Cry Toxins from Bacillus thuringiensis  

Digital Repository Infrastructure Vision for European Research (DRIVER)

Development of insect resistance is one of the main concerns with the use of transgenic crops expressing Cry toxins from the bacterium Bacillus thuringiensis. Identification of biomarkers would assist in the development of sensitive DNA-based methods to monitor evolution of resistance to Bt toxins in natural populations. We report on the proteomic and genomic detection of reduced levels of midgut membrane-bound alkaline phosphatase (mALP) as a common feature in strains of Cry-resistant Heliot...

Jurat-fuentes, Juan Luis; Karumbaiah, Lohitash; Jakka, Siva Rama Krishna; Ning, Changming; Liu, Chenxi; Wu, Kongming; Jackson, Jerreme; Gould, Fred; Blanco, Carlos; Portilla, Maribel; Perera, Omaththage; Adang, Michael

2011-01-01

182

Combined influence of temperature and metal ions on the level of activity of alkaline phosphatase of intestinal mucosa of Acipenseridae  

Digital Repository Infrastructure Vision for European Research (DRIVER)

The combined influence of divalent metal ions (Mn, Fe, Co, Ni, Cu and Zn) and temperature on the level of alkaline phosphatase activity of the mucous membrane of the Acipenseridae is shown. The dependence of the response of the enzyme to the action of metal ions according to their position in the periodic table of chemical elements is presented. This dependence is kept during the change of incubation temperature as well, but only at low temperatures the activating effect of metals is maximum ...

Bednyakov Dmitriy Andreevich; Nevalennyy Alexander Nickolaevich

2012-01-01

183

Modulation of placental alkaline phosphatase activity and cytokeratins in human HN-1 cells by butyrate, retinoic acid, catecholamines and histamine  

Digital Repository Infrastructure Vision for European Research (DRIVER)

The effects of butyrate and retinoic acid in combination with catecholamines or histamine on the HN-1 human head and neck squamous carcinoma cell line were investigated analysing cell proliferation, placental alkaline phosphatase (PLAP) activity, and relative cytokeratin content. Butyrate inhibited cell proliferation in agar, whereas retinoic acid induced a small inhibitory effect. Butyrate enhanced PLAP activity in a time related manner in contrast to retinoic acid, which had no significant ...

Bijman, J. T.; Wagener, D. J.; Rennes, H.; Wessels, J. M.; Ramaekers, F. C.; Den Broek, P.

1987-01-01

184

Effects of Colchicine on Localization of Alkaline Phosphatase in McA-RH 7777 Rat Hepatoma Cells  

Digital Repository Infrastructure Vision for European Research (DRIVER)

We investigated the changes caused by microtubule disruption in cell contact-induced translocation of alkaline phosphatase (ALP) from the Golgi area to the plasma membrane in McA-RH 7777 cells. When the cells were treated with colchicine, the tubular structure of microtubules in the cytoplasm was lost. Colchicine treatment also resulted in the appearance of numerous dots containing mannosidase II (man II) throughout the cytoplasm. Moreover, ALP was distributed in small dots throughout the cyt...

Taguchi, Meiko; Chida, Kohsuke

2008-01-01

185

Change in Localization of Alkaline Phosphatase and Mannosidase II by Colchicine Treatment of Primary Cultures of Fetal Rat Hepatocytes  

Digital Repository Infrastructure Vision for European Research (DRIVER)

We examined the changes in localization of alkaline phosphatase (ALP) and mannosidase II (man II), a Golgi marker, after colchicine treatment of primary cultures of fetal rat hepatocytes, using double immunofluorescence staining and confocal laser microscopy. In hepatocytes cultured in basal medium, ALP was localized in the perinuclear cytoplasm, and man II was observed in the Golgi region of the cytoplasm. When hepatocytes were cultured in dexamethasone-supplemented medium, ALP was also loca...

Chida, Kohsuke; Taguchi, Meiko

2008-01-01

186

Localization of Alkaline Phosphatase and Cathepsin D during Cell Restoration after Colchicine Treatment in Primary Cultures of Fetal Rat Hepatocytes  

Digital Repository Infrastructure Vision for European Research (DRIVER)

Localization of alkaline phosphatase (ALP) and cathepsin D (CAPD) in primary cultures of fetal rat hepatocytes was examined using double immunofluorescent staining in order to investigate the relationship between lysosome movement and the fate of ALP during cell restoration after microtubule disruption by colchicine. At 3 hr and 24 hr after colchicine treatment, numerous coarse dots containing ALP were observed throughout the cytoplasm, and some of these showed colocalization with CAPD. At 48...

Chida, Kohsuke; Taguchi, Meiko

2011-01-01

187

Crevicular Alkaline Phosphatase Activity and Rate of Tooth Movement of Female Orthodontic Subjects under Different Continuous Force Applications  

Digital Repository Infrastructure Vision for European Research (DRIVER)

Purpose. This study is aimed to compare the effects of two different orthodontic forces on crevicular alkaline phosphatase activity, rate of tooth movement, and root resorption. Materials and Methods. Twelve female subjects of class II division 1 malocclusion participated. Maxillary canines with bonded fixed appliances acted as the tested teeth, while their antagonists with no appliances acted as the controls. Canine retraction was performed using nickel titanium coil spring that delivered fo...

Rohaya Megat Abdul Wahab; Maryati Md Dasor; Sahidan Senafi; Asma Alhusna Abang Abdullah; Zulham Yamamoto; Abdul Aziz Jemain; Shahrul Hisham Zainal Ariffin

2013-01-01

188

Activity of alkaline and acidic phosphatase in glandular cells of uterine endometrium of puerperal ewes after exposure to polychlorinated biphenyls  

Digital Repository Infrastructure Vision for European Research (DRIVER)

The study is focused on the observation of alkaline and acidic phosphatase activity in the glandular cells of uterine endometrium in puerperal ewes after exposure to polychlorinated biphenyls. Ewes of Slovak merino breed (n=25) divided into 2 groups were included in the experiment. The animals in the experimental group (n=14) and control group (n=11) were euthanised on day 17, 25 and 34 postpartum. The ewes in the experimental group were given per os capsules of the chemical preparation Delor...

Valocky I.; Krajni?akova Maria; Legath J.; Lenhardt L.; Ostro A.; Danko J.; Udmila, Tkac?ikova L.; Mojžišova Jana; Fialkovi?ova Maria; Mardzinova Silvia

2005-01-01

189

HISTOENZYMOLOGICAL DEMONSTRATION OF ALKALINE PHOSPHATASE ACTIVITY IN THE ANTERIOR INTESTINE OF HgCl - TREATED HERBIVOROUS 2 FISH, LABEO ROHITA.  

Directory of Open Access Journals (Sweden)

Full Text Available Present studies incorporate enterotoxic effects of HgCl on the relative distribution of alkaline 2 phosphatase (AKP activity in the anterior intestine of fish, Labeo rohita. It was observed that there appears to be slightly decreased AKP activity accompanied by histolytic changes in the epithelial cells lining of intestinal villi and intestinal glands of the anterior intestine of treated fresh water fish. However, in control, these changes were invisible.

Dhirender

2014-07-01

190

HISTOENZYMOLOGICAL DEMONSTRATION OF ALKALINE PHOSPHATASE ACTIVITY IN THE ANTERIOR INTESTINE OF HgCl - TREATED HERBIVOROUS 2 FISH, LABEO ROHITA.  

Digital Repository Infrastructure Vision for European Research (DRIVER)

Present studies incorporate enterotoxic effects of HgCl on the relative distribution of alkaline 2 phosphatase (AKP) activity in the anterior intestine of fish, Labeo rohita. It was observed that there appears to be slightly decreased AKP activity accompanied by histolytic changes in the epithelial cells lining of intestinal villi and intestinal glands of the anterior intestine of treated fresh water fish. However, in control, these changes were invisible.

Dhirender; Jagdish Prasad

2014-01-01

191

Partial purification, characterisation and histochemical localisation of alkaline phosphatase from ascocarps of the edible desert truffle Terfezia claveryi Chatin.  

Science.gov (United States)

In the present paper, we confirmed that alkaline phosphatase (ALP) is the main phosphatase present in ascocarps of the edible mycorrhizal fungus Terfezia claveryi. The enzyme was partially purified by precipitation with polyethylene glycol. The purification achieved from a crude extract was fivefold, with 53% of the activity recovered, and acid phosphatase, most of the lipids and phenolic compounds were eliminated. Alkaline phosphatase was kinetically characterised at pH 10.0, the optimum for this enzyme, using p-nitrophenyl phosphate as substrate. The V(max) and K(m) values were 0.3 micromol.min(-1).mg(-1) protein and 9.0 mM, respectively. Orthovanadate was a competitive inhibitor of ALP, with a K(i) of 42.5 microM. The enzyme was histochemically localised in the peridium, the hypothecium and in the ascogenic hyphae of the gleba using both colour and fluorescent reactions. The results presented suggest that the ascocarp of T. claveryi, at some stages of its development, may become nutritionally autonomous and independent of the host plant. PMID:19689775

Navarro-Ródenas, A; Morte, A; Pérez-Gilabert, M

2009-09-01

192

[Studies into carcino-embryonic antigen (CEA) and enzyme activities of alkaline and acid phosphatase in patients with fibrocystic breast disease (author's transl)].  

Science.gov (United States)

CEA as well as alkaline and acid phosphatase were measured in patients with fibrocystic breast disease. The values recorded from the punctured fluids were compared to those in peripheral blood. CEA concentrations in cyst fluid were elevated in patients with proliferative breast disease, whereas no correlations could be established between alkaline and acid phosphatase, on the one hand, and various histological forms of breast disease, on the other. PMID:7269869

Opri, F; Kirchner, H

1981-01-01

193

The toxicity of four native Indian plants: effect on AChE and acid/alkaline phosphatase level in fish Channa marulius.  

Science.gov (United States)

The latex of four plants viz. Euphorbia royleana, Jatropha gossypifolia (Euphorbiaceae), Nerium indicum and Thevetia peruviana (Apocynaceae) caused significant reduction in acid/alkaline phosphatase activity and anti-acetylcholinesterase activity in nervous tissue of freshwater air breathing fish Channa marulius. The reduction in the activity of both phosphatases and AChE were time as well as dose dependent. PMID:15910912

Singh, Digvijay; Singh, Ajay

2005-06-01

194

Activity of alkaline and acidic phosphatase in glandular cells of uterine endometrium of puerperal ewes after exposure to polychlorinated biphenyls  

Directory of Open Access Journals (Sweden)

Full Text Available The study is focused on the observation of alkaline and acidic phosphatase activity in the glandular cells of uterine endometrium in puerperal ewes after exposure to polychlorinated biphenyls. Ewes of Slovak merino breed (n=25 divided into 2 groups were included in the experiment. The animals in the experimental group (n=14 and control group (n=11 were euthanised on day 17, 25 and 34 postpartum. The ewes in the experimental group were given per os capsules of the chemical preparation Delor 105 of domestic proveniance containing polychlorinated biphenyls (PCB for a period of 30 days. This preparation is equivalent to the foreign preparation Aroclor 1254. A dose of 100 µg/kg of Delor 105 was given to the animals of the experimental group. These animals were euthanised on day 17 postpartum (n=4 i. e. 5 days from the end of a 30-day period of application; on day 25 postpartum (n=5 i.e. 17 days from the last application of PCB; on day 34 postpartum (n=5, which was equivalent to day 28 from the last application. The ewes from the control group were euthanised on day 17 (n=3, day 25 (n=4 and on day 34 (n-4 postpartum. When evaluating alkaline phosphatase (ALP activity in the glandular cells of the endometrium in the control group, a statistically significant increase (P<0.01 was observed on day 25 and on day 34 (P<0.001 compared to day 17 postpartum. No statistically significant differences in alkaline phosphatase (ALP activity were observed (P>0.05 in the experimental group. The mean values of its activity in the observed period were below the level of values of day 17 in the control group. Acidic phosphatase activity in the glandular cells of the ewes' endometrium showed a statistically conclusive increase between day 17 and day 25 as well as day 34 postpartum (P<0.001. Acidic phosphatase density in the experimental group of ewes showed no statistically marked change (P>0.05 at the observed intervals postpartum. The discussion is focused on PCB effect on the activity of alkaline and acidic phosphatase in the glandular cells of the endometrium of ewes in the puerperal period.

Valocky I.

2005-01-01

195

TNF-? stimulates alkaline phosphatase and mineralization through PPAR? inhibition in human osteoblasts.  

Science.gov (United States)

The aims of the present study were to determine whether prostaglandins (PGs) and PPAR? are involved in the stimulation of tissue-non-specific alkaline phosphatase (TNAP) activity and mineralization by TNF-? in human osteoblasts. We used osteoblasts differentiated from MSCs from three different donors and MG-63 osteoblast-like cells. Inhibition of prostaglandin synthesis with the cyclooxygenase (COX) inhibitor indomethacin or the specific COX-2 blocker NS-398 abolished mineralization in the absence and presence of 1 ng/ml of TNF-?, suggesting that PGs were involved. The TNAP inhibitor levamisole abolished TNF-? effects on mineralization, suggesting that PGs were involved in TNAP expression and mineralization. TNF-? stimulated expression of COX-2 and PG E synthase before that of TNAP, but expression of PG D synthase later suggesting that PGE? and PGF?? but not 15d-PGJ? were involved in TNF-? effects. However, both PGE? and PGF?? dose-dependently inhibited mineralization indicating that endogenous PG are required for mineralization but that TNF-? does not increase mineralization by increasing PG synthesis. Interestingly, TNF-? inhibited PPAR? expression and binding activity to PPRE consensus sequences independently of 15d-PGJ?. Inhibition of PPAR? activity with GW-9662 mimicked TNF-? effects in MG-63 cells, indicating that TNF-? stimulates mineralization by inhibiting PPAR? in osteoblasts. In MSC-derived osteoblast cultures, inhibition of PPAR? dropped TNAP expression and mineralization. Treatment of MG-63 cells with conditioned media from MSC-derived osteoblasts or MSC-derived adipocytes treated or not with GW-9662 revealed that TNF-? inhibition of PPAR? in undifferentiated MSCs and/or adipocytes was responsible for the decreased expression of TNAP in osteoblasts. In conclusion, TNF-? increases TNAP expression and stimulates mineralization by inhibiting PPAR? in osteoblasts, but PPAR? in adipocytes or undifferentiated MSCs controls the secretion of a factor leading to TNAP stimulation in osteoblasts. PMID:20832511

Lencel, P; Delplace, S; Hardouin, P; Magne, D

2011-02-01

196

Reversible inactivation of alkaline phosphatase from Atlantic cod (Gadus morhua) in urea.  

Science.gov (United States)

Alkaline phosphatase (AP) from Atlantic cod (Gadus morhua) is a zinc and magnesium containing homodimer that requires the oligomeric state for activity. Its kinetic properties are indicative of cold-adaptation. Here, the effect of urea on the structural stability was studied in order to correlate the activity with metal content, the microenvironment around tryptophan residues, and events at the subunit interface. At the lowest concentrations of urea, the first detected alteration in properties was an increase in the activity of the enzyme. This was followed by inactivation, and the release of half of the zinc content when the amount of urea reached levels of 2 M. Intrinsic tryptophan fluorescence and circular dichroism ellipticity changed in the range 2.5 to 8 M urea, signaling dimer dissociation, followed by one major monomer unfolding transition at 6-8 M urea as indicated by ANS fluorescence and KI fluorescence quenching. Gibbs free energy was estimated by the linear extrapolation method using a three-state model as 8.6 kcal/mol for dimer stability and 11.6 kcal/mol for monomer unfolding giving a total of 31.8 kcal/mol. Dimer association had a very small ionic contribution. Dimers were stable in relatively high concentration of urea, whereas the immediate vicinity around the active site was vulnerable to low concentrations of urea. Thus, inactivation did not coincide with dimer dissociation, suggesting that the active site is the most dynamic part of the molecule and closest related to cold-adaptation of its enzymatic activity. PMID:16443405

Asgeirsson, Bjarni; Guojónsdóttir, Katrín

2006-02-01

197

Characterization of rat heart alkaline phosphatase isoenzymes and modulation of activity  

Scientific Electronic Library Online (English)

Full Text Available SciELO Brazil | Language: English Abstract in english Alkaline phosphatase (ALP) is important in calcification and its expression seems to be associated with the inflammatory process. We investigated the in vitro acute effects of compounds used for the prevention or treatment of cardiovascular diseases on total ALP activity from male Wistar rat heart h [...] omogenate. ALP activity was determined by quantifying, at 410 nm, the p-nitrophenol released from p-nitrophenylphosphate (substrate in Tris buffer, pH 10.4). Using specific inhibitors of ALP activity and the reverse transcription-polymerase chain reaction, we showed that the rat heart had high ALP activity (31.73 ± 3.43 nmol p-nitrophenol·mg protein-1·min-1): mainly tissue-nonspecific ALP but also tissue-specific intestinal ALP type II. Both ALP isoenzymes presented myocardial localization (striated pattern) by immunofluorescence. ALP was inhibited a) strongly by 0.5 mM levamisole, 2 mM theophylline and 2 mM aspirin (91, 77 and 84%, respectively) and b) less strongly by 2 mM L-phenylalanine, 100 mL polyphenol-rich beverages and 0.5 mM progesterone (24, 21 to 29 and 11%, respectively). ?-estradiol and caffeine (0.5 and 2 mM) had no effect; 0.5 mM simvastatin and 2 mM atenolol activated ALP (32 and 36%, respectively). Propranolol (2 mM) tended to activate ALP activity and corticosterone activated (18%) and inhibited (13%) (0.5 and 2 mM, respectively). We report, for the first time, that the rat heart expresses intestinal ALP type II and has high total ALP activity. ALP activity was inhibited by compounds used in the prevention of cardiovascular pathology. ALP manipulation in vivo may constitute an additional target for intervention in cardiovascular diseases.

A., Mota; P., Silva; D., Neves; C., Lemos; C., Calhau; D., Torres; F., Martel; H., Fraga; L., Ribeiro; M.N.M.P., Alçada; M.J., Pinho; M.R., Negrão; R., Pedrosa; S., Guerreiro; J.T., Guimarães; I., Azevedo; M.J., Martins.

2008-07-01

198

Structural characteristics of alkaline phosphatase from the moderately halophilic bacterium Halomonas sp. 593.  

Science.gov (United States)

Alkaline phosphatase (AP) from the moderate halophilic bacterium Halomonas sp. 593 (HaAP) catalyzes the hydrolysis of phosphomonoesters over a wide salt-concentration range (1-4?M NaCl). In order to clarify the structural basis of its halophilic characteristics and its wide-range adaptation to salt concentration, the tertiary structure of HaAP was determined by X-ray crystallography to 2.1?Å resolution. The unit cell of HaAP contained one dimer unit corresponding to the biological unit. The monomer structure of HaAP contains a domain comprised of an 11-stranded ?-sheet core with 19 surrounding ?-helices similar to those of APs from other species, and a unique `crown' domain containing an extended `arm' structure that participates in formation of a hydrophobic cluster at the entrance to the substrate-binding site. The HaAP structure also displays a unique distribution of negatively charged residues and hydrophobic residues in comparison to other known AP structures. AP from Vibrio sp. G15-21 (VAP; a slight halophile) has the highest similarity in sequence (70.0% identity) and structure (C(?) r.m.s.d. of 0.82?Å for the monomer) to HaAP. The surface of the HaAP dimer is substantially more acidic than that of the VAP dimer (144 exposed Asp/Glu residues versus 114, respectively), and thus may enable the solubility of HaAP under high-salt conditions. Conversely, the monomer unit of HaAP formed a substantially larger hydrophobic interior comprising 329 C atoms from completely buried residues, whereas that of VAP comprised 264 C atoms, which may maintain the stability of HaAP under low-salt conditions. These characteristics of HaAP may be responsible for its unique functional adaptation permitting activity over a wide range of salt concentrations. PMID:24598750

Arai, Shigeki; Yonezawa, Yasushi; Ishibashi, Matsujiro; Matsumoto, Fumiko; Adachi, Motoyasu; Tamada, Taro; Tokunaga, Hiroko; Blaber, Michael; Tokunaga, Masao; Kuroki, Ryota

2014-03-01

199

Properties of a constitutive alkaline phosphatase from strain 74A of the mold Neurospora crassa  

Directory of Open Access Journals (Sweden)

Full Text Available A constitutive alkaline phosphatase was purified to apparent homogeneity as determined by polyacrylamide gel electrophoresis from mycelia of the wild strain 74A of the mold Neurospora crassa, after growth on acetate and in the presence of saturating amounts of inorganic phosphate (Pi for 72 h at 30ºC. The molecular mass was 58 kDa and 56 kDa as determined by exclusion chromatography and SDS-PAGE, respectively. This monomeric enzyme shows an apparent optimum pH ranging from 9.5 to 10.5 and Michaelis kinetics for the hydrolysis of p-nitrophenyl phosphate (the Km and Hill coefficient values were 0.35 mM and 1.01, respectively, alpha-naphthyl phosphate (the Km and Hill coefficient values were 0.44 mM and 0.97, respectively, ß-glycerol phosphate (the Km and Hill coefficient values were 2.46 mM and 1.01, respectively and L-histidinol phosphate (the Km and Hill coefficient values were 0.47 mM and 0.94, respectively at pH 8.9. The purified enzyme is activated by Mg2+, Zn2+ and Tris-HCl buffer, and is inhibited by Be2+, histidine and EDTA. Also, 0.3 M Tris-HCl buffer protected the purified enzyme against heat inactivation at 70ºC(half-life of 19.0 min, k = 0.036 min-1 as compared to 0.3 M CHES (half-life of 2.3 min, k = 0.392 min-1 in the same experiment.

A.C. Morales

2000-08-01

200

Contribution of matrix vesicles and alkaline phosphatase to ectopic bone formation  

Scientific Electronic Library Online (English)

Full Text Available SciELO Brazil | Language: English Abstract in english Endochondral calcification involves the participation of matrix vesicles (MVs), but it remains unclear whether calcification ectopically induced by implants of demineralized bone matrix also proceeds via MVs. Ectopic bone formation was induced by implanting rat demineralized diaphyseal bone matrix i [...] nto the dorsal subcutaneous tissue of Wistar rats and was examined histologically and biochemically. Budding of MVs from chondrocytes was observed to serve as nucleation sites for mineralization during induced ectopic osteogenesis, presenting a diameter with Gaussian distribution with a median of 306 ± 103 nm. While the role of tissue-nonspecific alkaline phosphatase (TNAP) during mineralization involves hydrolysis of inorganic pyrophosphate (PPi), it is unclear how the microenvironment of MV may affect the ability of TNAP to hydrolyze the variety of substrates present at sites of mineralization. We show that the implants contain high levels of TNAP capable of hydrolyzing p-nitrophenylphosphate (pNPP), ATP and PPi. The catalytic properties of glycosyl phosphatidylinositol-anchored, polidocanol-solubilized and phosphatidylinositol-specific phospholipase C-released TNAP were compared using pNPP, ATP and PPi as substrates. While the enzymatic efficiency (k cat/Km) remained comparable between polidocanol-solubilized and membrane-bound TNAP for all three substrates, the k cat/Km for the phosphatidylinositol-specific phospholipase C-solubilized enzyme increased approximately 108-, 56-, and 556-fold for pNPP, ATP and PPi, respectively, compared to the membrane-bound enzyme. Our data are consistent with the involvement of MVs during ectopic calcification and also suggest that the location of TNAP on the membrane of MVs may play a role in determining substrate selectivity in this micro-compartment.

P., Ciancaglini; A.M.S., Simão; F.L., Camolezi; J.L., Millán; J.M., Pizauro.

2006-05-01

 
 
 
 
201

Properties of a constitutive alkaline phosphatase from strain 74A of the mold Neurospora crassa  

Scientific Electronic Library Online (English)

Full Text Available SciELO Brazil | Language: English Abstract in english A constitutive alkaline phosphatase was purified to apparent homogeneity as determined by polyacrylamide gel electrophoresis from mycelia of the wild strain 74A of the mold Neurospora crassa, after growth on acetate and in the presence of saturating amounts of inorganic phosphate (Pi) for 72 h at 30 [...] ºC. The molecular mass was 58 kDa and 56 kDa as determined by exclusion chromatography and SDS-PAGE, respectively. This monomeric enzyme shows an apparent optimum pH ranging from 9.5 to 10.5 and Michaelis kinetics for the hydrolysis of p-nitrophenyl phosphate (the Km and Hill coefficient values were 0.35 mM and 1.01, respectively), alpha-naphthyl phosphate (the Km and Hill coefficient values were 0.44 mM and 0.97, respectively), ß-glycerol phosphate (the Km and Hill coefficient values were 2.46 mM and 1.01, respectively) and L-histidinol phosphate (the Km and Hill coefficient values were 0.47 mM and 0.94, respectively) at pH 8.9. The purified enzyme is activated by Mg2+, Zn2+ and Tris-HCl buffer, and is inhibited by Be2+, histidine and EDTA. Also, 0.3 M Tris-HCl buffer protected the purified enzyme against heat inactivation at 70ºC(half-life of 19.0 min, k = 0.036 min-1) as compared to 0.3 M CHES (half-life of 2.3 min, k = 0.392 min-1) in the same experiment.

A.C., Morales; S.R., Nozawa; G., Thedei Jr.; W., Maccheroni Jr.; A., Rossi.

2000-08-01

202

Intestinal Alkaline Phosphatase Has Beneficial Effects in Mouse Models of Chronic Colitis  

Science.gov (United States)

Background The brush border enzyme intestinal alkaline phosphatase (IAP) functions as a gut mucosal defense factor and is protective against dextran sulfate sodium (DSS)-induced acute injury in rats. The present study evaluated the potential therapeutic role for orally administered calf IAP (cIAP) in two independent mouse models of chronic colitis: (1) DSS-induced chronic colitis, and (2) chronic spontaneous colitis in Wiskott-Aldrich Syndrome protein (WASP) deficient (knockout) mice that is accelerated by irradiation. Methods The wild-type (WT) and IAP knockout (IAP-KO) mice received 4 cycles of 2% DSS ad libitum for 7 days. Each cycle was followed by a 7-day DSS-free interval during which mice received either cIAP or vehicle in the drinking water. The WASP-KO mice received either vehicle or cIAP for 6 weeks beginning on the day of irradiation. Results Microscopic colitis scores of DSS-treated IAP-KO mice were higher than DSS-treated WT mice (52 ± 3.8 vs. 28.8 ± 6.6, respectively, P < 0.0001). cIAP treatment attenuated the disease in both groups (KO = 30.7 ± 6.01, WT = 18.7 ± 5.0, P < 0.05). In irradiated WASP-KO mice cIAP also attenuated colitis compared to control groups (3.3 ± 0.52 vs. 6.2 ± 0.34, respectively, P < 0.001). Tissue myeloperoxidase activity and pro-inflammatory cytokines were significantly decreased by cIAP treatment. Conclusions Endogenous IAP appears to play a role in protecting the host against chronic colitis. Orally administered cIAP exerts a protective effect in two independent mouse models of chronic colitis and may represent a novel therapy for human IBD. PMID:20645323

Ramasamy, Sundaram; Nguyen, Deanna D.; Eston, Michelle; Alam, Sayeda Nasrin; Moss, Angela K.; Ebrahimi, Farzad; Biswas, Brishti; Mostafa, Golam; Chen, Kathryn T.; Kaliannan, Kanakaraju; Yammine, Halim; Narisawa, Sonoko; Millán, José Luis; Warren, H. Shaw; Hohmann, Elizabeth L.; Mizoguchi, Emiko; Reinecker, Hans-Christian; Bhan, Atul K.; Snapper, Scott B.; Malo, Madhu S.; Hodin, Richard A.

2011-01-01

203

Colocation and role of polyphosphates and alkaline phosphatase in apatite biomineralization of elasmobranch tesserae.  

Science.gov (United States)

Elasmobranchs (e.g. sharks and rays), like all fishes, grow continuously throughout life. Unlike other vertebrates, their skeletons are primarily cartilaginous, comprising a hyaline cartilage-like core, stiffened by a thin outer array of mineralized, abutting and interconnected tiles called tesserae. Tesserae bear active mineralization fronts at all margins and the tesseral layer is thin enough to section without decalcifying, making this a tractable but largely unexamined system for investigating controlled apatite mineralization, while also offering a potential analog for endochondral ossification. The chemical mechanism for tesserae mineralization has not been described, but has been previously attributed to spherical precursors, and alkaline phosphatase (ALP) activity. Here, we use a variety of techniques to elucidate the involvement of phosphorus-containing precursors in the formation of tesserae at their mineralization fronts. Using Raman spectroscopy, fluorescence microscopy and histological methods, we demonstrate that ALP activity is located with inorganic phosphate polymers (polyP) at the tessera-uncalcified cartilage interface, suggesting a potential mechanism for regulated mineralization: inorganic phosphate (Pi) can be cleaved from polyP by ALP, thus making Pi locally available for apatite biomineralization. The application of exogenous ALP to tissue cross-sections resulted in the disappearance of polyP and the appearance of Pi in uncalcified cartilage adjacent to mineralization fronts. We propose that elasmobranch skeletal cells control apatite biomineralization by biochemically controlling polyP and ALP production, placement and activity. Previous identification of polyP and ALP shown previously in mammalian calcifying cartilage supports the hypothesis that this mechanism may be a general regulating feature in the mineralization of vertebrate skeletons. PMID:24948547

Omelon, Sidney; Georgiou, John; Variola, Fabio; Dean, Mason N

2014-09-01

204

Evaluation of spectrophotometric and fluorometric methods for alkaline phosphatase activity determination in ewe's milk.  

Science.gov (United States)

The alkaline phosphatase (ALP) activity test has been used since 1935 to assess the effectiveness of pasteurization. Different analytical methods exist for detecting ALP in milk. Unfortunately, there is little information about ALP activity in ewe's milk. The aim of this study was to assess and compare the official European method (spectrophotometric method) and the Fluorophos method (fluorometric method) regarding their use in ewe's milk. Bulk ewe's milk samples were taken from a flock and from three different dairies. A portion of the original sample was pasteurized at 63 degrees C for 30 min in a circulating bath; another portion was heated to and kept at 95 degrees C for about 2 min, and 0.1% (vol/vol) of raw milk was added. The samples obtained were analyzed in duplicate using the spectrophotometric and fluorometric methods. The relation between ALP activity determined by the two methods was characterized by the following equation: Y = 1.34 + 0.0039X (where Y = ALP in microg of phenol per ml of milk and X = ALP in mU/liter; R2 = 91.5%). Precision parameters (repeatability [r], standard deviation of repeatability [s(r)], and relative standard deviation of repeatability [RSDr]) for both methods were calculated. The values of RSDr for the Fluorophos method were 4.30 for pasteurized milk and 2.96 for 0.1% raw milk, close to the value indicated by Rocco in whole cow's milk (RSDr = 4.4). The repeatability for the official method (r = 2.16) was close to that indicated for whole cow's milk (r = 2). PMID:10983802

Scintu, M F; Daga, E; Ledda, A

2000-09-01

205

Evaluation of alkaline phosphatase levels and salivary hydroxyapatite ion activity in 6?12-year-old children with different caries severity  

Directory of Open Access Journals (Sweden)

Full Text Available Introduction: Salivary calcium and phosphate levels might have a relationship with dental caries severity. Evaluation of different indexes such as alkaline phosphatase, which increases salivary phosphate levels, and hydroxyapatite ion activity, which reflects salivary calcium and phosphate levels, might help determine the risk of dental caries. The aim of the present study was to compare salivary alkaline phosphatase activity and hydroxyapatite ion activity index in 6?12-year-old children with different rates of dental caries. Materials and methods: In this descriptive-analytical study the teeth were examined in 152 children, aged 6?12, by a dentist and were placed into three experimental groups according to the dental caries severity as follows: group 1: severe dental caries (DMFT ? 6; group 2: moderate dental caries rate (1 < DMFT < 6; and group 3: mild dental caries rate (DMFT ? 1. After collection of saliva, salivary alkaline phosphatase levels and hydroxyapatite ion activity were measured for each sample. Data was analyzed using one-way ANOVA and a post hoc Tukey test (? = 0.05. Results: The means and standard deviations of alkaline phosphatase levels in groups 1 to 3 were 5.39 ± 2.96, 5.71 ± 3.68 and 5.83 ± 4.53 unit/dL, respectively. Hydroxyapatite ion activity rates were 25.80 ± 0.70, 28.28 ± 0.76 and 28.50 ± 0.56 in groups 1 to 3, respectively. The results of one-way ANOVA showed no significant differences between the experimental groups in alkaline phosphatase levels (p value = 0.830 and hydroxyapatite ion activity (p value = 0.065.Conclusion: According to the finding of the present study, alkaline phosphatase level and hydroxyapatite ion activity have no effect on tooth caries severity in 6?12-year-old children. Future studies are recommended. Key words: Alkaline phosphatase, Dental caries, Saliva

Loghman Rezaei- Soufi

2013-01-01

206

Fluorometric detection of active alkaline phosphatase and gamma-glutamyl transferase in fluid dairy products from multiple species.  

Science.gov (United States)

Over the past 80 years, a variety of methods have been developed to detect underpasteurized or improperly pasteurized milks used in dairy products. Existing methods are hampered by duration of analysis, poor reproducibility, and in some cases the use of hazardous chemicals. To overcome these issues, two new methods have been developed using fluorogenic substrates for two marker enzymes, alkaline phosphatase and gamma-glutamyl transferase. In 30 min, up to 18 samples can be analyzed in triplicate by both methods on two separate 96-well plates. Sample preparation is not necessary for liquid milks when using these methods. The relative standard deviation for each assay is less than 9%, and the correlation coefficient for results of the two methods is greater than 0.98. Using the new methods, milks from four species and nine commercially available liquid milk products were tested. The new methods were also tested directly against an existing phosphatase method (Fluorophos) in spiked whole milk samples. PMID:23643136

Ziobro, George C; McElroy, Kevin M

2013-05-01

207

The effect of the alkaline tide on serum-ionized calcium concentration in man.  

Science.gov (United States)

The effect of the gastric alkaline tide on serum-ionized calcium levels was determined in human subjects. Gastric acid seretion was stimulated by a standard steak meal, human synthetic gastrin, and betazole hydrochloride. Ionized calcium levels fell to a similar extent after each stimulus. The mean decrease in calcium ion concentration in all experiments was 5.4% of the basal concentration. The fall in serum calcium ion concentration was highly correlated with the rise in serum pH. We speculate that increased formation of calcium bicarbonate complex in the serum as well as increased binding of ionized calcium by serum protein accounts for the surprisingly large effect of the alkaline tide on serumionized calcium levels. PMID:14862

Hughes, W; Cohen, S; Arvan, D; Seamonds, B

1977-01-01

208

Combined influence of temperature and metal ions on the level of activity of alkaline phosphatase of intestinal mucosa of Acipenseridae  

Directory of Open Access Journals (Sweden)

Full Text Available The combined influence of divalent metal ions (Mn, Fe, Co, Ni, Cu and Zn and temperature on the level of alkaline phosphatase activity of the mucous membrane of the Acipenseridae is shown. The dependence of the response of the enzyme to the action of metal ions according to their position in the periodic table of chemical elements is presented. This dependence is kept during the change of incubation temperature as well, but only at low temperatures the activating effect of metals is maximum in the early period at high temperatures the inhibitory effects of metals is maximum at the end of the period.

Bednyakov Dmitriy Andreevich

2012-11-01

209

Cloning of and complementation tests with alkaline phosphatase regulatory genes (phoS and phoT) of Escherichia coli.  

Digital Repository Infrastructure Vision for European Research (DRIVER)

The regulatory genes of alkaline phosphatase, phoS and phoT, of Escherichia coli were cloned on pBR322, initially as an 11.8-kilobase EcoRI fragment. A restriction map of the hybrid plasmid was established. Deletion plasmids of various sizes were constructed in vitro, and the presence of phoS and phoT genes on the cloned DNA fragments was tested by introducing the plasmids into phoS64 and phoT9 strains for complementation tests. One set complemented only phoS64 but not phoT9; the other set co...

Amemura, M.; Shinagawa, H.; Makino, K.; Otsuji, N.; Nakata, A.

1982-01-01

210

A Phos-Tag-Based Fluorescence Quenching System for Activity Assay and Inhibitor Screening for Alkaline Phosphatase  

Digital Repository Infrastructure Vision for European Research (DRIVER)

Fluorescence resonance energy transfer (FRET) is a distance-dependent interaction between the electronic excited states of two dye molecules. Here we introduce a novel FRET-based fluorescence quenching system for assaying the activity of alkaline phosphatase (AP) by using a phos-phate-binding tag molecule, Phos-tag {1,3-bis[bis(pyridine-2-ylmethyl)amino]propan-2-olato dizinc(II) complex}, attached to a nonfluorescent 4-{[4-(dimethylamino)phenyl]diazenyl}benzoyl (Dabcyl: ?max

Emiko Kinoshita-Kikuta; Hiromasa Kurosaki; Natsumi Kunisada; Eiji Kinoshita; Tohru Koike

2014-01-01

211

Recombinant antibody-alkaline phosphatase conjugates for diagnosis of human IgGs: application to anti-HBsAg detection.  

Science.gov (United States)

We have designed an expression vector permitting the production in the periplasm of Escherichia coli of a fusion protein comprising a dimer of bacterial alkaline phosphatase (PhoA) and two Fab or scFv fragments of a monoclonal antibody directed against human IgG. Each hybrid protein expressed both high specificity for the antigen and full PhoA activity. We show that crude periplasmic extracts containing these conjugates can be used as such in enzyme immunoassays for the detection of human IgG, as exemplified in the case of anti-hepatitis B immunoglobulin. PMID:7745247

Carrier, A; Ducancel, F; Settiawan, N B; Cattolico, L; Maillère, B; Léonetti, M; Drevet, P; Ménez, A; Boulain, J C

1995-04-26

212

Use of an alkaline phosphatase-labeled synthetic oligonucleotide probe for detection of Campylobacter jejuni and Campylobacter coli.  

Digital Repository Infrastructure Vision for European Research (DRIVER)

A commercially available synthetic nucleic acid probe (SNAP) conjugated to alkaline phosphatase was compared with standard culture techniques for detecting Campylobacter species. The SNAP was able to detect either 5 ng of C. jejuni DNA or 10(5) CFU of bacteria. The SNAP could also detect DNA extracted from 10(5) CFU in mock-infected stool samples. The SNAP detected C. jejuni and C. coli but showed no reactivity with C. laridis, C. fetus subsp. fetus, C. fetus subsp. venerealis, C. fennelliae,...

Olive, D. M.; Johny, M.; Sethi, S. K.

1990-01-01

213

Activity of the enzymes of pentose phosphate way and of alkaline phosphatase in rats under incorporation of nitrates and radionuclides  

International Nuclear Information System (INIS)

The pentose-phosphate way major enzymes (transketolase, glucose-6-phosphate dehydrogenase) and the alkaline phosphatase activity in thymus, spleen and mesenteric lymph nodes were studied after introducing of nitrates (5% NaNO3) with drinking water and radionuclides (cesium 137 and strontium 90) with grain during 45-50 days to intact rats and to those suffering B1 hypovitaminosis by injection of oxythiamine. It was revealed changes in the lymphoid tissue cells protein synthesis connected with impairments in the immune mechanisms. The oxythiamine hypovitaminosis caused more marked changes in the enzyme activity in the lymphoid organs

214

Effect of vanadate, a potent alkaline phosphatase inhibitor, on 45Ca and 32P/sub i/ uptake by matrix vesicle-enriched fractions from chicken epiphyseal cartilage  

International Nuclear Information System (INIS)

Vanadate was tested in a hydroxyapatite-seeded ion uptake system to determine possible direct effects on mineral formation. The effect of vanadate on vesicle mineral ion uptake was complex; low dosages of vanadate (2-20 ?M) were stimulatory to Ca2+ uptake, but were inhibitory to P/sub i/. Higher dosages (>67 ?M) were inhibitory to both ions. The effect of vanadate on ion uptake was strongly influenced by the stage of vesicle loading; major effects were seen during the lag and early uptake phases, and minimal effects were seen in the terminal stages. Concentrations of vanadate highly inhibitory to vesicle ion uptake had minimal effects on ion accretion by a hydroxyapatite-seeded system. Inhibition of alkaline phosphatase activity by vanadate broadly paralleled inhibition of P/sub i/ and Ca2+ uptake; however, at low vanadate concentrations, inhibition of P/sub i/ uptake closely paralleled that of alkaline phosphatase. The data indicate that vanadate binds with high affinity to P/sub i/-loading sites, blocking initial P/sub i/ uptake. The close parallelism between inhibition of early P/sub i/ uptake and of alkaline phosphatase activity supports the concept that alkaline phosphatase is involved in P/sub i/ transport during the early stages of matrix vesicle ion loading. However, the fact that only about half of the P/sub i/ uptake was affected by vanadate, despite the progressive inhibition of alkaline phosphatase activity, indicates that alkaline atase activity, indicates that alkaline phosphatase is not solely responsible for P/sub i/ uptake by the matrix vesicle-enriched fraction

215

Perinatal undernutrition alters intestinal alkaline phosphatase and its main transcription factors KLF4 and Cdx1 in adult offspring fed a high-fat diet.  

Science.gov (United States)

Nutrient restriction during gestation and/or suckling is associated with an increased risk of developing inflammation, obesity and metabolic diseases in adulthood. However, the underlying mechanisms, including the role of the small intestine, are unclear. We hypothesized that intestinal adaptation to the diet in adulthood is modulated by perinatal nutrition. This hypothesis was tested using a split-plot design experiment with 20 controls and 20 intrauterine growth-retarded (IUGR) rats aged 240 days and randomly assigned to be fed a standard chow or a high-fat (HF) diet for 10 days. Jejunal tissue was collected at necropsy and analyzed for anatomy, digestive enzymes, goblet cells and mRNA levels. Cecal contents and blood serum were analyzed for alkaline phosphatase (AP). IUGR rats failed to adapt to HF by increasing AP activity in jejunal tissue and cecal content as observed in controls. mRNA levels of transcription factors KLF4 and Cdx1 were blunted in jejunal epithelial cell of IUGR rats fed HF. mRNA levels of TNF-? were lower in IUGR rats. They also displayed exacerbated aminopeptidase N response and reduced jejunal goblet cell density. Villus and crypt architecture and epithelial cell proliferation increased with HF in both control and IUGR rats. Serum AP tended to be lower, and serum levamisole inhibition-resistant AP fraction was lower, in IUGR than controls with HF. Serum fatty acids and triglycerides were higher in IUGR rats and higher with HF. In conclusion, the adult intestine adapts to an HF diet differentially depending on early nutrition, jejunal AP and transcription factors being blunted in IUGR individuals fed HF. PMID:22405696

Lallès, Jean-Paul; Orozco-Solís, Ricardo; Bolaños-Jiménez, Francisco; de Coppet, Pierre; Le Dréan, Gwénola; Segain, Jean-Pierre

2012-11-01

216

Variations of alkaline phosphatase activity and P fractions in sediments of a shallow Chinese eutrophic lake (Lake Taihu)  

International Nuclear Information System (INIS)

The distribution of alkaline phosphatase activity (APA) and P fractions in sediment cores and the relationship between them were studied in a shallow Chinese freshwater lake (Lake Taihu). Sediment cores were collected from four sites, characterized by different degrees of eutrophication in June 2004. Sediment P was fractionated into Fe/Al-P, Ca-P, organic P (OP), inorganic P (IP) and total P (TP). The former two species made the largest contribution to the sediment P pool. Results show that trophic status and hydrological conditions have great impact on the APA of the sediments. The order of the APA in sediments was conjectured to be: macrophyte dominated lake > transitional lake > algal dominated lake. APA profiles follow a similar downcore decreasing trend. There was a positive relationship between the APA and the TP, IP. The multiple linear regression equation of the APA and P fractions is: APA = -97 + 0.768TP - 0.985Fe/Al-P. - Characteristics of the alkaline phosphatase activity and P fractions in sediments of different trophic status lake were studied in Lake Taihu

217

Multiple forms of human intestinal alkaline phosphatase: chemical and enzymatic properties, and circulating clearances of the fast- and slow-moving enzymes  

International Nuclear Information System (INIS)

Two forms of alkaline phosphatase orthophosphoric monoester phosphohydrolase (alkaline optimum, EC 3.1.3.1) have been purified from human small intestine by column chromatography on DEAE-cellulose and tyraminyl derivative affinity gel, and by preparative disc gel electrophoresis. Intestinal phosphatases were electrophoretically separated into two components, fast- and slow-moving enzymes, with apparent molecular weights of 140000 and 168000 and with subunit weights of 68000 and 80000, respectively. Organ distribution of injected 125I-labelled enzymes indicates that the desialylated hepatic enzyme was selectively distributed in liver, while the degalactosylated intestinal enzyme was incorporated into liver, lymph fluid, and small intestine. These results suggest that the pathway of circulating clearance of alkaline phosphatase has several routes. (Auth.)

218

Effect of gingival application of melatonin on alkaline and acid phosphatase, osteopontin and osteocalcin in patients with diabetes and periodontal disease  

Digital Repository Infrastructure Vision for European Research (DRIVER)

Objectives: To assess the effect of topical application of melatonin to the gingiva on salivary fluid concentrations of acid phosphatase, alkaline phosphatase, osteopontin, and osteocalcin. Study Design: Cross-sectional study of 30 patients with diabetes and periodontal disease and 30 healthy subjects. Diabetic patients were treated with topical application of melatonin (1% orabase cream formula) once daily for 20 days and controls with a placebo formulation. Results: Before treatment w...

Cutando, Antonio; Lo?pez-valverde, Antonio; Go?mez-de-diego, Rafel; Arias-santiago, Salvador; Vicente-jime?nez, Joaqui?n

2013-01-01

219

Ultrasensitive detection of cancer cells and glycan expression profiling based on a multivalent recognition and alkaline phosphatase-responsive electrogenerated chemiluminescence biosensor  

Science.gov (United States)

A multivalent recognition and alkaline phosphatase (ALP)-responsive electrogenerated chemiluminescence (ECL) biosensor for cancer cell detection and in situ evaluation of cell surface glycan expression was developed on a poly(amidoamine) (PAMAM) dendrimer-conjugated, chemically reduced graphene oxide (rGO) electrode interface. In this strategy, the multivalency and high affinity of the cell-targeted aptamers on rGO provided a highly efficient cell recognition platform on the electrode. The ALP and concanavalin A (Con A) coated gold nanoparticles (Au NPs) nanoprobes allowed the ALP enzyme-catalyzed production of phenols that inhibited the ECL reaction of Ru(bpy)32+ on the rGO electrode interface, affording fast and highly sensitive ECL cytosensing and cell surface glycan evaluation. Combining the multivalent aptamer interface and ALP nanoprobes, the ECL cytosensor showed a detection limit of 38 CCRF-CEM cells per mL in human serum samples, broad dynamic range and excellent selectivity. In addition, the proposed biosensor provided a valuable insight into dynamic profiling of the expression of different glycans on cell surfaces, based on the carbohydrates recognized by lectins applied to the nanoprobes. This biosensor exhibits great promise in clinical diagnosis and drug screening.A multivalent recognition and alkaline phosphatase (ALP)-responsive electrogenerated chemiluminescence (ECL) biosensor for cancer cell detection and in situ evaluation of cell surface glycan expression was developed on a poly(amidoamine) (PAMAM) dendrimer-conjugated, chemically reduced graphene oxide (rGO) electrode interface. In this strategy, the multivalency and high affinity of the cell-targeted aptamers on rGO provided a highly efficient cell recognition platform on the electrode. The ALP and concanavalin A (Con A) coated gold nanoparticles (Au NPs) nanoprobes allowed the ALP enzyme-catalyzed production of phenols that inhibited the ECL reaction of Ru(bpy)32+ on the rGO electrode interface, affording fast and highly sensitive ECL cytosensing and cell surface glycan evaluation. Combining the multivalent aptamer interface and ALP nanoprobes, the ECL cytosensor showed a detection limit of 38 CCRF-CEM cells per mL in human serum samples, broad dynamic range and excellent selectivity. In addition, the proposed biosensor provided a valuable insight into dynamic profiling of the expression of different glycans on cell surfaces, based on the carbohydrates recognized by lectins applied to the nanoprobes. This biosensor exhibits great promise in clinical diagnosis and drug screening. Electronic supplementary information (ESI) available: CV and EIS during the electrode assembly, activity of the nanoprobes and the glycan-binding specificities of the lectins. See DOI: 10.1039/c4nr03053b

Chen, Xiaojiao; He, Yao; Zhang, Youyu; Liu, Meiling; Liu, Yang; Li, Jinghong

2014-09-01

220

Probing the origins of catalytic discrimination between phosphate and sulfate monoester hydrolysis: comparative analysis of alkaline phosphatase and protein tyrosine phosphatases.  

Science.gov (United States)

Catalytic promiscuity, the ability of enzymes to catalyze multiple reactions, provides an opportunity to gain a deeper understanding of the origins of catalysis and substrate specificity. Alkaline phosphatase (AP) catalyzes both phosphate and sulfate monoester hydrolysis reactions with a ?10(10)-fold preference for phosphate monoester hydrolysis, despite the similarity between these reactions. The preponderance of formal positive charge in the AP active site, particularly from three divalent metal ions, was proposed to be responsible for this preference by providing stronger electrostatic interactions with the more negatively charged phosphoryl group versus the sulfuryl group. To test whether positively charged metal ions are required to achieve a high preference for the phosphate monoester hydrolysis reaction, the catalytic preference of three protein tyrosine phosphatases (PTPs), which do not contain metal ions, were measured. Their preferences ranged from 5 × 10(6) to 7 × 10(7), lower than that for AP but still substantial, indicating that metal ions and a high preponderance of formal positive charge within the active site are not required to achieve a strong catalytic preference for phosphate monoester over sulfate monoester hydrolysis. The observed ionic strength dependences of kcat/KM values for phosphate and sulfate monoester hydrolysis are steeper for the more highly charged phosphate ester with both AP and the PTP Stp1, following the dependence expected based on the charge difference of these two substrates. However, the dependences for AP were not greater than those of Stp1 and were rather shallow for both enzymes. These results suggest that overall electrostatics from formal positive charge within the active site is not the major driving force in distinguishing between these reactions and that substantial discrimination can be attained without metal ions. Thus, local properties of the active site, presumably including multiple positioned dipolar hydrogen bond donors within the active site, dominate in defining this reaction specificity. PMID:25299936

Andrews, Logan D; Zalatan, Jesse G; Herschlag, Daniel

2014-11-01

 
 
 
 
221

Ratio of serum tartrate-inhibitable acid phosphatase to total serum protein in benign prostatic hypertrophy and prostatic carcinoma.  

Science.gov (United States)

The activity concentration and the specific activity (the ratio of enzyme activity to total serum protein) of the tartrate-inhibitable fraction of acid phosphatase [orthophosphoric monoester phosphohydrolase (acid optimum), EC 3.1.3.2; TIAP] were related to benign prostatic hypertrophy and to prostatic carcinoma. As expected, the TIAP activity concentrations assayed in the sera of patients with benign prostatic hypertrophy were within the range of those assayed in normal human sera. In contrast, the specific activities of TIAP determined in the sera of patients with benign prostatic hypertrophy were significantly higher than those determined in the control group. In the sera of prostatic carcinoma patients, both the TIAP activity concentrations and the TIAP specific activities differed significantly (F = 730) from the normal values. PMID:1371723

Petrovich, G; Ursich-Jankovich, J; Prijovich, Z

1992-02-01

222

Formation of a vitamin B-12-serum complex on heating at alkaline pH  

International Nuclear Information System (INIS)

The binding of vitamin B-12 to serum proteins during heating at alkaline pH was investigated by gel filtration of serum supplemented with cyano[57Co]-cobalamin. Heating for 5 min at 1000C destroyed most of the vitamin B-12 binding activity of serum but, with further heating, the vitamin B-12 became incorporated into a complex that did not correspond in molecular size to the original vitamin B-12 binding proteins. Radioassay of vitamin B-12 in heated serum showed correspondingly first an increase then a progressive decrease in the apparent vitamin B-12 level suggesting that, on heating, vitamin B-12 was initially released then subsequently complexed by the serum. The formation of complexed vitamin B-12 was abolished by the presence of the reducing agent dithiothreitol during the heating step. (Auth.)

223

Treatment with bortezomib in multiple myeloma is associated with only a transient and brief increase of bone specific alkaline phosphatase  

DEFF Research Database (Denmark)

There are indications of a bone anabolic effect associated with bortezomib treatment. We present a study with long follow up, measuring bone specific alkaline phosphatase (bALP) for a year during and after treatment in an unselected cohort of myeloma patients treated with bortezomib, and assess factors of potential influence on the increase of bALP. Our main findings are that bALP increase is of short duration and declines significantly even during continued treatment with bortezomib. Only myeloma response was associated with a significant increase of bALP; whereas previous treatment with bortezomib, previous or concomitant treatment with zoledronic acid i.v., dose of bortezomib, line of treatment, or combination with other chemotherapy was not.

Haidl, Felix; Plesner, Torben

2012-01-01

224

Radionuclide bone scan, radiographic bone survey, and alkaline phosphatase: studies of limited value in asymptomatic patients with ovarian carcinoma  

International Nuclear Information System (INIS)

Bone scans or skeletal surveys were obtained in 104 patients with ovarian carcinoma. No metastases were identified at staging in the 43 patients with Stage I or II disease. Four patients in the entire series had osseous metastases. Three of the 40 patients with Stage III epithelian ovarian carcinoma has osseous metastases at the time of staging. All of these were Grade III lesions. One Stage I, Grade III patient demonstrated osseous metastases two years after initial diagnosis. None of the four patients with osseous metastases had an elevated alkaline phosphatase; three of the four had bone pain. Based on these results, it is suggested that radiographic bone survey and radionuclide bone scans are not indicated as screening procedures in asymptomatic patients with ovarian carcinoma

225

Surface modification of Ti-6Al-4 V alloy for biomineralization and specific biological response: part II, alkaline phosphatase grafting.  

Science.gov (United States)

Titanium and its alloys are the most widespread materials for the realization of orthopaedic and dental implants due to their good mechanical properties and biocompatibility. Surface functionalization of biomaterials aimed to improve and quicken implant integration and tissue regeneration is an active research field. The opportunity to confer biological activity (ability to directly stimulate cells with proper biological signals) to the Ti6Al4 V alloy, previously modified to be bioactive from the inorganic point of view (apatite precipitation), was explored in this research work. The alkaline phosphatase (ALP) enzyme was grafted to metal surface via tresyl chloride activation, maintaining its activity. A synergistic effect between biological functionalization and inorganic bioactivity was observed. PMID:21660585

Ferraris, S; Spriano, S; Bianchi, C L; Cassinelli, C; Vernè, E

2011-08-01

226

Localization of Alkaline Phosphatase and Cathepsin D during Cell Restoration after Colchicine Treatment in Primary Cultures of Fetal Rat Hepatocytes  

Science.gov (United States)

Localization of alkaline phosphatase (ALP) and cathepsin D (CAPD) in primary cultures of fetal rat hepatocytes was examined using double immunofluorescent staining in order to investigate the relationship between lysosome movement and the fate of ALP during cell restoration after microtubule disruption by colchicine. At 3 hr and 24 hr after colchicine treatment, numerous coarse dots containing ALP were observed throughout the cytoplasm, and some of these showed colocalization with CAPD. At 48 hr and 72 hr after colchicine treatment, although most of the dots containing ALP in the cytoplasm disappeared, dots containing CAPD remained. The present results suggest that the denatured ALP proteins remaining in the cytoplasm of hepatocytes during cell restoration after colchicine treatment are digested by lysosomes. PMID:21753861

Chida, Kohsuke; Taguchi, Meiko

2011-01-01

227

Active Site Detection by Spatial Conformity and Electrostatic Analysis—Unravelling a Proteolytic Function in Shrimp Alkaline Phosphatase  

Science.gov (United States)

Computational methods are increasingly gaining importance as an aid in identifying active sites. Mostly these methods tend to have structural information that supplement sequence conservation based analyses. Development of tools that compute electrostatic potentials has further improved our ability to better characterize the active site residues in proteins. We have described a computational methodology for detecting active sites based on structural and electrostatic conformity - CataLytic Active Site Prediction (CLASP). In our pipelined model, physical 3D signature of any particular enzymatic function as defined by its active sites is used to obtain spatially congruent matches. While previous work has revealed that catalytic residues have large pKa deviations from standard values, we show that for a given enzymatic activity, electrostatic potential difference (PD) between analogous residue pairs in an active site taken from different proteins of the same family are similar. False positives in spatially congruent matches are further pruned by PD analysis where cognate pairs with large deviations are rejected. We first present the results of active site prediction by CLASP for two enzymatic activities - ?-lactamases and serine proteases, two of the most extensively investigated enzymes. The results of CLASP analysis on motifs extracted from Catalytic Site Atlas (CSA) are also presented in order to demonstrate its ability to accurately classify any protein, putative or otherwise, with known structure. The source code and database is made available at www.sanchak.com/clasp/. Subsequently, we probed alkaline phosphatases (AP), one of the well known promiscuous enzymes, for additional activities. Such a search has led us to predict a hitherto unknown function of shrimp alkaline phosphatase (SAP), where the protein acts as a protease. Finally, we present experimental evidence of the prediction by CLASP by showing that SAP indeed has protease activity in vitro. PMID:22174814

Chakraborty, Sandeep; Minda, Renu; Salaye, Lipika; Bhattacharjee, Swapan K.; Rao, Basuthkar J.

2011-01-01

228

Nicotine inhibits collagen synthesis and alkaline phosphatase activity, but stimulates DNA synthesis in osteoblast-like cells  

International Nuclear Information System (INIS)

Use of smokeless tobacco is associated with various oral lesions including periodontal damage and alveolar bone loss. This study was performed to test the effects of nicotine on bone-forming cells at concentrations that occur in the saliva of smokeless tobacco users. Confluent cultures of osteoblast-like cells isolated from chick embryo calvariae were incubated for 2 days with nicotine added to the culture medium (25-600 micrograms/ml). Nicotine inhibited alkaline phosphatase in the cell layer and released to the medium, whereas glycolysis (as indexed by lactate production) was unaffected or slightly elevated. The effects on medium and cell layer alkaline phosphatase were concentration dependent with maximal inhibition occurring at 600 micrograms nicotine/ml. Nicotine essentially did not affect the noncollagenous protein content of the cell layer, but did inhibit collagen synthesis (hydroxylation of [3H]proline and collagenase-digestible protein) at 100, 300, and 600 micrograms/ml. Release of [3H]hydroxyproline to the medium was also decreased in a dose-dependent manner, as was the collagenase-digestible protein for both the medium and cell layer. In contrast, DNA synthesis (incorporation of [3H]thymidine) was more than doubled by the alkaloid, whereas total DNA content was slightly inhibited at 600 micrograms/ml, suggesting stimulated cell turnover. Morphologic changes occurred in nicotine-treated cells including rounding up, detachmeated cells including rounding up, detachment, and the occurrence of numerous large vacuoles. These results suggest that steps to reduce the salivary concentration of nicotine in smokeless tobacco users might diminish damaging effects of this product on alveolar bone

229

The effect of 1,25-dihydroxycholecalciferol on the multiple forms of alkaline phosphatase and the sialic acid incorporation into microsomes of chick duodenum  

International Nuclear Information System (INIS)

Polyacrylamide disc gel electrophoresis of n-butanol solubilized alkaline phosphatase from chick duodenum revealed that the change of alkaline phosphatase induced by 1,25-(OH)2D3 involved the transformation of desialoenzyme to sialoenzyme. The initial stimulation by 1,25-(OH)2D3 of the incorporation of sialic acid into duodenal microsomes corresponded with the initial increase in calcium absorption. After this initial stimulation, there was a rapid decline in sialic acid incorporation into microsomes decreasing below control levels at 24 hr. Calcium concentration in the microsomes followed a pattern similar to the incorporation of sialic acid into microsomes. The depressed sialic acid incorporation was reversed by the addition of calcium in vitro. These results suggest that the initial action of 1,25-(OH)2D3 is to change the membrane permeability to calcium and to change the subcellular distribution of calcium in the small intestine. The accumulated calcium in the microsomes then stimulates the sialic acid incorporation into desialoenzyme. This results in the changes of isozyme pattern of alkaline phosphatase, viz, the transformation of desialoenzyme to sialoenzyme. The transformed alkaline phosphatase might be one of the factors involved more directly in the regulation of calcium transport in intestine. (auth.)

230

CONTROL OF ALKALINE PHOSPHATASE ACTIVITY IN C3H10T1/2 CELLS: ROLE OF RETINOIC ACID AND CELL DENSITY  

Science.gov (United States)

The enzyme alkaline phosphatase (AP) has been shown to be lost or inappropriately expressed during carcinogenesis in some tissues. ecause retinoic acid (RA) appears to play a role in the normal regulation of the enzyme (RA up-regulates AP in a variety of cell types) we have sugge...

231

Evidence of associations between feto-maternal vitamin D status, cord parathyroid hormone and bone-specific alkaline phosphatase, and newborn whole body bone mineral content  

Science.gov (United States)

In spite of a high prevalence of vitamin D inadequacy in pregnant women and neonates, relationships among vitamin D status [25(OH)D], parathyroid hormone (PTH), bone specific alkaline phosphatase (BALP), and whole body bone mineral content (WBBMC) in the newborn are poorly characterized. The purpose...

232

Effect of glyphosate toxicity on growth, pigment and alkaline phosphatase activity in cyanobacterium Anabaena doliolum: a role of inorganic phosphate in glyphosate tolerance.  

Science.gov (United States)

Response of glyphosate toxicity on photoautotrophic cyanobacterium A. doliolum and its mutant strain was investigated. Chlorophyll a content of both the wild type and mutant strain in the presence of glyphosate (N-phosphonomethyl glycine) initially showed an increasing trend when supplemented with Pi and a declining tendency under the Pi-starved condition. The results suggested that both the wild type and mutant strains were more sensitive to glyphosate in the absence of phosphate. Alkaline phosphatase activity of wild type strain in the presence of Pi, enhanced in response to addition of glyphosate (40 microg/ml), but the activity remained unaltered by addition of glyphosate in the Pi-starved cells, whereas the alkaline phosphatase activity in the mutant strain under both Pi-starved as well as unstarved conditions was stimulated (approximately 5.4 and 3.1-fold, respectively) by addition of glyphosate. The results on alkaline phosphatase activity indicated a glyphosate-induced depletion in the phosphate content of the cells, particularly in the mutant strain, as evident from the stimulated activity of alkaline phosphatase. It is suggested that enzyme activity in the Pi-starved wild type cells may not be influenced any further by glyphosate, as cellular phosphate reserve might not be available for further depletion. PMID:15282956

Shikha; Singh, D P; Darmwal, N S

2004-02-01

233

ALKALINE PHOSPHATASE REACTIVITY IN THE VAGINA AND UTEROVAGINAL JUNCTION SPERM-STORAGE TUBULES OF TURKEYS IN EGG PRODUCTION: IMPLICATIONS FOR SPERM STORAGE  

Science.gov (United States)

Currently there remains contradictory information on the localization and possible role of alkaline phosphatase (AP) in the chicken and Japanese quail oviducts. Using turkeys with a hard-shelled egg in their uteri, vaginal and uterovaginal junction mucosae were stretched and fixed as whole mounts ...

234

Residual alkaline phosphatase activity in pasteurized milk heated at various temperatures--measurement with the fluorophos and Scharer rapid phosphatase tests.  

Science.gov (United States)

Milk containing three levels of milkfat (skim [0.5%], lowfat [2.0%], and whole [3.25%]) were heat treated at five different temperatures (59, 61, 63, 65, and 67 degrees C) using a laboratory scale, batch pasteurization method. Heated milk samples were removed at 5-min intervals, immediately cooled, and then assayed using the quantitative fluorometric method and the qualitative Scharer rapid test. Mean alkaline phosphatase (ALP) activity values as measured with the Fluorophos method decreased in all milk preparations as the time of sampling and temperature of heating increased. Samples representing the three fat levels and heat treated at 63 degrees C for 30 min, the minimum time/temperature allowed by the 1995 pasteurized milk ordinance (PMO), had ALP activity values <100 mU/liter. All values were below the 350 mU/liter standard for fluid milk products established by the Food and Drug Administration and cited in the 1995 PMO. Evaluation of the milks for adequacy of pasteurization with the Scharer rapid method indicated that those same milks were adequately pasteurized. PMID:9921835

Angelino, P D; Christen, G L; Penfield, M P; Beattie, S

1999-01-01

235

Purification and Characterization of an Alkaline Phosphatase Induced by Phosphorus Starvation in Common Bean (Phaseolus vulgaris L.) Roots  

Scientific Electronic Library Online (English)

Full Text Available SciELO Mexico | Language: English Abstract in spanish Dos isoformas de fosfatasas obtenidas de raíz de frijol (Phaseolus vulgaris L.) mostraron un incremento en la actividad en respuesta a la deficiencia de fosfato. Una de ellas (APIII) se purificó a través de una cromatografía de intercambio iónico y una electroforesis preparativa. La masa molecular e [...] stimada para APIII fue de 35 kDa tanto por SDS-PAGE como por filtración molecular, sugiriendo que la enzima activa es monomérica. APIII se clasificó como una fosfatasa alcalina basada en sus requerimientos de pH 8 para catálisis. Esta enzima es activa sobre un amplio espectro de sustratos como polifosfato, glucose 1-fosfato y fosfoenolpiruvato, aunque muestra preferencia por pirofosfato. Su actividad se inhibe completamente por molibdato, vanadato y fosfato, aunque es inhibida parcialmente por fluoruro. Aún cuando los cationes divalentes no fueron escenciales para su actividad, la hidrólisis de pirofosfato se incrementó notablemente en presencia de Mg2+. Abstract in english Two phosphatase isoforms from roots of the common bean (Phaseolus vulgaris L.) showed an increase in activity in response to phosphate deficiency. One of them (APIII) was chosen for further purification through ionic exchange chromatography and preparative electrophoresis. The estimated molecular ma [...] ss of APIII was 35 kDa by both SDS-PAGE and gel filtration analyses, suggesting a monomeric form of the active enzyme. The phosphatase was classified as an alkaline phosphatase based on the requirement of pH 8 for optimum catalysis. It not only exhibited broad substrate specificity, with the most activity against pyrophosphate, but also effectively catalyzed the hydrolysis of polyphosphate, glucose-1-phosphate and phosphoenol-pyruvate. Activity was completely inhibited by molybdate, vanadate and phosphate but was only partially inhibited by fluoride. Although divalent cations were not essential for the pyrophosphatase activity of this enzyme, the hydrolysis of pyrophosphate increased substantially in the presence of Mg2+.

Lorena, Morales; Natalia, Gutiérrez; Vanessa, Maya; Carmen, Parra; Eleazar, Martínez-Barajas; Patricia, Coello.

2012-03-01

236

Effect of x-irradiation on the acid and alkaline phosphatase activities in the south Indian scorpion Heterometrus fulvipes (C.L. Koch)  

International Nuclear Information System (INIS)

Variations in the activity levels of acid phosphatase and alkaline phosphatase in the tissues of scorpion Heterometrus fulvipes after whole-body irradiation with X-rays are studied. The animals were exposed to sublethal dose (1/3 of LD50) of radiation for different periods of time ranging from 1/2h to 4h. Tissue homogenates were also irradiated and studied. The activity of phosphatases was found to decrease with increase in exposure time both at in vivo and in vitro, the per cent decrease being higher at in vitro. The activity levels of phosphatases in different tissues were in the order of hepato pancreas > heart pedipalpal > muscle > nervous tissue. (M.G.B.)

237

Aplicación del método inmunocitoquímico de la fosfatasa alcalina anti-fosfatasa alcalina para la clasificación inmunológica de las leucemias mieloides agudas Application of the alkaline phosphatase anti-alkaline phosphatase immunocytochemical method for the immunological classification of acute myeloid leukemias  

Directory of Open Access Journals (Sweden)

Full Text Available Se realizó el inmunofenotipaje celular de 30 pacientes con el diagnóstico de leucemia mieloide aguda por el método inmunocitoquímico fosfatasa alcalina anti-fosfatasa alcalina (APAAP introducido en nuestro laboratorio. Los marcadores estudiados fueron: CD3, CD13, CD15, CD19, CD33 y CD41. Para el estudio se utilizaron extendidos de médula ósea o sangre periférica fijados en acetona pura e incubados con el respectivo anticuerpo monoclonal. Posteriormente se añadió la inmunoglobulina anti ratón obtenida en conejo ( Linking y por último, el complejo APAAP. Los períodos de incubación fueron de 30 minutos y se realizaron lavados con solución amortiguadora entre cada uno de los pasos. La lectura de las láminas se realizó en microscopio óptico y se consideró positivo cuando el número de células marcadas era mayor o igual a 20 %. De los pacientes estudiados, el 93,3 % y el 90 %, respectivamente, expresaron antígenos pan mieloides CD13 y CD33; 16 de ellos expresaron el CD15 (53,3 %; 3 el CD19 (10 % y 2 el CD41 (6,6 %. Se concluyó que el método APAAP es rápido y de bajo costo y puede ser aplicado con confiabilidad en la clasificación inmunológica de las leucemias mieloides agudasThe cellular immunophenotyping of 30 patients with the diagnosis of acute myeloid leukemia was conducted by the alkaline phosphatase anti-alkaline phosphatase immunocytochemical method (APAAP introduced in our laboratory. The markers studied were: CD3, CD13, CD15, CD19, CD33 y CD41. Specimens of bone marrow or peripheral blood fixed in pure acetone and incubated with the respective monoclonal antibody were used for the study. Later on, the anti-mouse immunoglobulin obtained in rabbit (Linking was added and, finally, the APAAP complex. The incubation periods were of 30 minutes and lavages with buffer solution were carried out between one step and the other. The reading of the slides was performed on the optical microscope and it was considered positive when the number of marked cells was higher than or equal to 20 %. Of the studied patients, 93.3 % and 90 %, respectively, expressed panmyeloid antigens CD13 and CD33; 16 of them expressed the CD15 (53.3 %; 3 the CD19 (10 %; and 2 the CD41 (6.6 %. It was concluded that the APAAP method is rapid and inexpensive and that it may be reliably applied in the immunological classification of the acute myeloid leukemias

Bertha B Socarrás Ferrer

2006-04-01

238

Aplicación del método inmunocitoquímico de la fosfatasa alcalina anti-fosfatasa alcalina para la clasificación inmunológica de las leucemias mieloides agudas / Application of the alkaline phosphatase anti-alkaline phosphatase immunocytochemical method for the immunological classification of acute myeloid leukemias  

Scientific Electronic Library Online (English)

Full Text Available SciELO Cuba | Language: Spanish Abstract in spanish Se realizó el inmunofenotipaje celular de 30 pacientes con el diagnóstico de leucemia mieloide aguda por el método inmunocitoquímico fosfatasa alcalina anti-fosfatasa alcalina (APAAP) introducido en nuestro laboratorio. Los marcadores estudiados fueron: CD3, CD13, CD15, CD19, CD33 y CD41. Para el es [...] tudio se utilizaron extendidos de médula ósea o sangre periférica fijados en acetona pura e incubados con el respectivo anticuerpo monoclonal. Posteriormente se añadió la inmunoglobulina anti ratón obtenida en conejo ( Linking ) y por último, el complejo APAAP. Los períodos de incubación fueron de 30 minutos y se realizaron lavados con solución amortiguadora entre cada uno de los pasos. La lectura de las láminas se realizó en microscopio óptico y se consideró positivo cuando el número de células marcadas era mayor o igual a 20 %. De los pacientes estudiados, el 93,3 % y el 90 %, respectivamente, expresaron antígenos pan mieloides CD13 y CD33; 16 de ellos expresaron el CD15 (53,3 %); 3 el CD19 (10 %) y 2 el CD41 (6,6 %). Se concluyó que el método APAAP es rápido y de bajo costo y puede ser aplicado con confiabilidad en la clasificación inmunológica de las leucemias mieloides agudas Abstract in english The cellular immunophenotyping of 30 patients with the diagnosis of acute myeloid leukemia was conducted by the alkaline phosphatase anti-alkaline phosphatase immunocytochemical method (APAAP) introduced in our laboratory. The markers studied were: CD3, CD13, CD15, CD19, CD33 y CD41. Specimens of bo [...] ne marrow or peripheral blood fixed in pure acetone and incubated with the respective monoclonal antibody were used for the study. Later on, the anti-mouse immunoglobulin obtained in rabbit (Linking) was added and, finally, the APAAP complex. The incubation periods were of 30 minutes and lavages with buffer solution were carried out between one step and the other. The reading of the slides was performed on the optical microscope and it was considered positive when the number of marked cells was higher than or equal to 20 %. Of the studied patients, 93.3 % and 90 %, respectively, expressed panmyeloid antigens CD13 and CD33; 16 of them expressed the CD15 (53.3 %); 3 the CD19 (10 %); and 2 the CD41 (6.6 %). It was concluded that the APAAP method is rapid and inexpensive and that it may be reliably applied in the immunological classification of the acute myeloid leukemias

Bertha B, Socarrás Ferrer; Lázaro O, del Valle Pérez; Vianed, Súarez; Julio C, Merlín Linares; Consuelo, Macías Abraham.

2006-04-01

239

Comparison of Salivary Ion Activity Product for Hydroxyapatite (IPHA, Alkaline Phosphatase and Buffering Capacity of Adults According to Age and Caries Severity  

Directory of Open Access Journals (Sweden)

Full Text Available Statement of Problem: Tooth caries is influenced by different biochemical characteristics of saliva. As hydroxyapatite is the main component of enamel, salivary ion activity product for hydroxyapatite (IPHA as well as alkaline phosphatase may be attributed to dental caries. Purpose: The aim of the present study was to compare salivary buffering capacity, alkaline phosphatase and IPHA of adults according to the dental caries and age. Materials and Method: One hundred and twenty 19 to 44 years old male individuals were divided into four groups according to the dental caries rate and age: group 1: 19-35 years old low dental caries (DMFT <5; group 2: 19-35 years old high dental caries (DMFT 5<; group 3: 35-44 years old low dental caries (DMFT <11 and 35-44 years old high dental caries (DMFT 11<. Five millilitre of unstimulated saliva was collected, and then buffering capacity, the level of alkaline phosphatase activity and IPHA was determined for each sample. Data was analyzed by soft ware SPSS using two-way ANOVA, Friedman and Mann-Whitney tests.Results: Mean and standard deviation of buffering capacity of group 1 to 4 was 2.66±0.54, 2.64±0.56, 2.70±0.70 and 2.26±0.82, respectively. The difference was not significance (p= 0. 305. Mean and standard deviation of alkaline phosphatase activity of group 1 to 4 was 5.82±2.91, 5.30±1.52, 4.77±1.82 and 4.55±1.61, respectively. There was no significant difference (p= 0.692. Mean and standard deviation of IPHA of group 1 to 4 was 29.39±0.61, 29.51±0.76, 29.14±0.56 and 29.75±0.75, respectively. The difference was significant (p= 0.049.Conclusion: Based on the results of the present study, buffering capacity and the level of alkaline phosphatase couldn’t affect dental caries, independently. However, the higher value of IPHA may be attributed to the higher dental caries rate. Ageing decreases alkaline phosphatase activity.

Vahedi M.

2012-12-01

240

Alkaline phosphatase activity at the southwest coast of India: A comparison of locations differently affected by upwelling  

Science.gov (United States)

The realization of the potential importance of phosphorus (P) as a limiting nutrient in marine ecosystem is increasing globally. Hence, the contribution of biotic variables in mobilizing this nutrient would be relevant especially in productive coastal waters. As alkaline phosphatase activity (APA) indicates the status of P for primary production in aquatic environments, we asked the following question: is the level of APA indicative of P sufficiency or deficiency in coastal waters, especially, where upwelling is a regular phenomenon? Therefore, we have examined the total APA, chlorophyll a along with phosphatase producing bacteria (PPB) and related environmental parameters from nearshore to offshore in coastal waters off Trivandrum and Kochi regions differently affected by upwelling during the onset of monsoon. Off Trivandrum, APA in the offshore waters of 5-m layer at 2.23 ?M P h- 1 was > 4 times higher than nearshore. Thus, low APA could be indicative of P sufficiency in coastal waters and higher activity suggestive of deficiency in offshore waters off Trivandrum. In contrast, there was less difference in APA between near and offshore surface waters off Kochi. Our results show that the regions differently affected by upwelling respond differently according to ambient P concentration, distance from shore or depth of water. These observations could apparently be applicable to other coastal systems as well, where gradients in upwelling and phosphate runoff have been noticed. Further studies on other transects would throw more light on the extent and direction of the relationship between APA and ambient P concentration. Such studies would help in understanding the level of control of this nutrient on the productivity of coastal waters.

Mamatha, S. S.; Malik, Ashish; Varik, Sandesh; Parvathi, V.; Jineesh, V. K.; Gauns, Mangesh U.; LokaBharathi, P. A.

2015-01-01

 
 
 
 
241

Assessment of the colorimetric and fluorometric assays for alkaline phosphatase activity in cow's, goat's, and sheep's milk.  

Science.gov (United States)

Raw milk is a well-established vehicle for the carriage of human pathogens, and many regulatory bodies have consequently mandated compulsory pasteurization as a food safety intervention. The residual activity of alkaline phosphatase (ALP) has historically been used to verify the adequacy of pasteurization of cow's milk. However, there is uncertainty on how the current ALP standards and methods of analysis can be applied to sheep's and goat's milk, which naturally contain different levels of the enzyme than that found in cow's milk. The official ALP methods applied in Canada (colorimetric assay; MFO-3) and in the United States (Fluorophos) were assessed for their ability to detect enzyme activity in raw and pasteurized milk derived from cows, sheep, and goats. The detection limit and the limit of quantitation were 0.8 and 2.02 microg/ml phenol, respectively, for the MFO-3 method and 43 and 85 mU/liter, respectively, for the Fluorophos method. The average ALP levels in raw goat's, cow's, and sheep's milk were 165, 1,562, and 3,512 microg/ml phenol, respectively. Raw milk detection limits, which correspond to raw milk phosphatase levels, were 0.051, 0.485, and 0.023% in cow's, goat's, and sheep's milk, respectively, for the MFO-3 method and 0.007, 0.070, and 0.004%, respectively, for the Fluorophos method. Although both methods can be used for ALP determination in cow's, goat's, and sheep's milk, the Fluorophos assay was superior to the colorimetric MFO-3 method based on sensitivity and time required to complete the analysis. PMID:18810873

Klotz, V; Hill, Art; Warriner, K; Griffiths, M; Odumeru, J

2008-09-01

242

Collaborative evaluation of a fluorometric method for measuring alkaline phosphatase activity in cow's, sheep's, and goat's milk.  

Science.gov (United States)

Pasteurization of raw milk was introduced to extend product shelf life and destroy pathogens. The measurement of alkaline phosphatase (ALP) activity has been used as an indicator of proper pasteurization in dairy products for more than 65 years. This study was undertaken to evaluate six different fluid dairy products at lower phosphatase levels than previously verified using the Fluorophos Test System, a sensitive and precise method for ALP activity detection. Thirteen laboratories participated in this collaborative, international study to evaluate the fluorometric test at 20, 40, 100, 350, and 500 mU/liter and extend the scope of the method to include milk from not only cows but also goats and sheep. Initially, the statutory level of ALP measured fluorometrically was set to equivalent levels of colorimetric test standards (500 mU/liter). The European Union recently announced its intention of lowering the legal limit from 500 to 350 mU/liter and, in addition, setting a target value of 100 mU/liter, which if exceeded would trigger an investigation into the pasteurizer plant performance. At 500 mU/liter of ALP, this trial generated relative standard deviation of repeatability values of 6.48, 5.69, and 1.74% and relative standard deviation of reproducibility values of 14.66, 13.30, and 5.33% for all cow's, sheep's, and goat's milk samples, respectively. Data from this study are comparable to data from previous studies and indicate the suitability of the Fluorophos Test System method for measuring ALP activity in milk from cows, sheep, and goats not only at the current European statutory level of 500 mU/liter but also at much lower levels. PMID:15895740

Harding, Frank; Garry, Eileen

2005-05-01

243

EXPRESSION OF ALKALINE PHOSPHATASE DURING OSTEOGENIC DIFFERENTIATION OF RAT BONE MARROW STROMAL CELLS  

Digital Repository Infrastructure Vision for European Research (DRIVER)

Introduction: Bone marrow contains a population of stem cells capable of differentiating to osteoblast and forming the bone nodule by dexamethasone. Material and Methods: The stromal cells of bone marrow obtained from 4 to 6 weeks old Spruge-Dawely male rats were grown in primary culture for 7 days and subcultured for 18 days. The cells were cultured in either DMEM medium containing 15% fetal calf serum and antibiotics as the controls or the above medium supplemented with osteogenic supplemen...

Akbari, M.; Nikbakht, M.; Gh, Sobhani A.

2001-01-01

244

Assessment of pasteurisation of milk and cream produced by on-farm dairies using a fluorimetric method for alkaline phosphatase activity.  

Science.gov (United States)

The alkaline phosphatase test is used as an indicator of adequate pasteurisation of milk and cream. A proprietary fluorimetric technique (Fluorophos) is a sensitive and quantitative method for the determination of alkaline phosphatase (ALP) activity in milk products. Currently, adequate pasteurisation of milk products is regarded as confirmed in samples that contain a residual bovine ALP activity of 500 mU/litre was deemed as a failure and was found in 3.5% of whole milk, 2.4% semiskimmed milk, 5.0% of skimmed milk, and 39% of cream samples from on-farm dairies. Bovine ALP activity of >100 and <500 mU/litre was found in 18.4% of whole milk, 9.3% of semi-skimmed milk, 13.2% skimmed milk and 44.5% of cream samples from on-farm dairies. Results with skimmed milk samples showed significantly lower bovine ALP activity than whole milk. All 409 milk and cream samples from two large commercial dairies passed the fluorimetric test at less than 500 mU/litre of bovine ALP, and 99% of these milk and cream samples had bovine ALP activity of less than 100 mU/litre. The presence of residual bovine phosphatase indicates a failure and may be due to either inadequate pasteurisation or post pasteurisation contamination with raw milk. Residual bovine phosphatase was demonstrated in 108/114 (94.7%) of milk samples with a bovine ALP activity greater than 500 mU/litre, i.e. true failures. Of more concern is that residual bovine phosphatase was found in 395/401 (98.5%) of samples that gave bovine ALP activity greater than 100 mU/litre but equal to or less than 500 mU/litre. Residual bovine phosphatase was demonstrated in 37/108 (30.2%) of cream samples with bovine ALP activity greater than 500 mU/litre. Presence of reactivated bovine phosphatase is not an indication of a failure but can mask the presence of residual bovine phosphatase. Reactivated bovine phosphatase was found in 74/106 (69.8%) of cream samples. Our results confirm that the more sensitive fluorimetric method is suitable for testing pasteurised whole milk and semiskimmed milk, but for statutory purposes the acceptable level of residual bovine phosphatase should be <100 mU/litre. Our findings have highlighted a potential problem when testing skimmed milk and cream samples from on-farm dairies. To ensure public safety we need more stringent standards for the ALP test and new methods that will accurately confirm that pasteurisation of these products has been achieved. PMID:15259408

Allen, G; Bolton, F J; Wareing, D R A; Williamson, J K; Wright, P A

2004-06-01

245

Alkaline Phosphatase Activity : an overlooked player on the phosphate behavior in macrotidal estuaries  

Science.gov (United States)

The non-conservative behavior of phosphate within the estuarine salinity gradient is essentially assigned to physico-chemical processes, such as desorption at low salinity and to benthic exchanges. Microbial phosphatase activity (APA), generally related to phosphate deficiency, is seldom studied in phosphate rich estuarine waters. In order to address the impact of microbial activity (bacterial abundance, production BSP, APA) on phosphate behavior, we studied these activities on a seasonal basis within the salinity gradient of two macrotidal estuaries presenting different levels of suspended solids. Whatever the season the Charente estuary is characterized by high levels of Suspended Particulate Matter (SPM > 1g.L-1), particularly in the Maximum Turbidity Zone (MTZ) located at the 5-10 psu. In this area characterized by high BSP and APA there is a significant increase of PO4 levels especially during summer. In the Aulne estuary the particle load is significantly lower (1/10) but high BSP and APA are equally recorded. In the highly turbid waters of the Charente estuary, active phytoplankton is virtually absent as pheopigments constitute up to 80% of the total pigments, particularly in the MTZ, therefore APA may essentially have a bacterial origin. In the Aulne estuary attached bacteria are dominant, both in numbers and production, and their distribution along the haline gradient perfectly follows those of APA and phosphate levels. These observations, associated with the very close relationships observed between APA, SPM and BSP, suggest that APA derive mainly from bacterial (attached) origin and operate at the expense of particulate phosphorus and hence contribute to PO4 regeneration, especially in spring and summer. Finally, as APA increased as PO4, whereas the reverse is observed in both fresh and marine waters, an original scheme for APA regulation, related to the large dominance of attached bacteria can be described for the estuarine waters.

Delmas, Daniel; Labry, Claire; Youenou, Agnes; Quere, Julien; Auguet, Jean Christophe; Montanie, Helene

2014-05-01

246

Relation of fatty acid composition in lead-exposed mallards to fat mobilization, lipid peroxidation and alkaline phosphatase activity  

Science.gov (United States)

The increase of n-6 polyunsaturated fatty acids (PUFA) in animal tissues has been proposed as a mechanism of Pb poisoning through lipid peroxidation or altered eicosanoids metabolism. We have studied fatty acid (FA) composition in liver and brain of mallards (Anas platyrhynchos) feeding for three weeks on diets containing combinations of low or high levels of vitamin E (20 or 200 UI/kg) and Pb (0 or 2 g/kg). Saturated FA, n-6 PUFA and total concentrations of FA were higher in livers of Pb-exposed mallards, but not in their brains. The percentage of n-6 PUFA in liver and brain was slightly higher in Pb-exposed mallards. The increase of n-6 PUFA in liver was associated with increased triglycerides and cholesterol in plasma, thus could be in part attributed to feed refusal and fat mobilization. The hepatic ratios between adrenic acid (22:4 n-6) and arachidonic acid (20:4 n-6) or between adrenic acid and linoleic acid (18:2 n-6) were higher in Pb exposed birds, supporting the existing hypothesis of increased fatty acid elongation by Pb. Among the possible consequences of increased n-6 PUFA concentration in tissues, we found increased lipid peroxidation in liver without important histopathological changes, and decreased plasma alkaline phosphatase activity that may reflect altered bone metabolism in birds.

Mateo, R.; Beyer, W.N.; Spann, J.W.; Hoffman, D.J.

2003-01-01

247

Alkaline phosphatase, cytokeratin 7, cytokeratin 8 in the diagnosis of primary lung adenocarcinoma from 148 pleura fluids specimens.  

Directory of Open Access Journals (Sweden)

Full Text Available Adenocarcinomas are the most common cause of malignancy in pleura fluids. Usual primary sites include the lung, breast, gastrointestinal tract, and genitourinary tracts. Predicting the site of origin of an adenocarcinoma can be difficult due to overlapping morphologic characteristics. We investigated the use of alkaline phosphatase (AP, Cytokeratin7 (CK7 Cytokeratin8 (CK8 to distinguish adenocarcinomas of lung in 148 body cavity fluid samples. Overall results for primary lung adenocarcinomas, demonstrated CK8 reactivity in 106 (72% of 148 cases. 95 primary lung carcinoma samples (65% were positive for CK7. AP was expressed in 81% of primary lung adenocarcinomas. Positive immunoreactivity for AP was characterized by a red, diffusely apical cytoplasmic staining in tumor cells that ocurred singly or in groups. There was a significant difference between AP, CK 7 and CK 8 expressions in primary lung adenocarcinomas (P=0.02; Chi-squared test. The sensitivity of AP, CK8, CK7 as a marker for primary lung adenocarcinomas were 82%, 72%, 64%, respectively. Thus the AP positive staining largely confirmed the cytologic diagnosis of lung adenocarcinoma.

Temelli Ozlem

2009-05-01

248

Dissolved organic phosphorus utilization and alkaline phosphatase activity of the dinoflagellate Gymnodinium impudicum isolated from the South Sea of Korea  

Science.gov (United States)

This study investigated alkaline phosphatase (APase) activity and dissolved organic and inorganic phosphorus utilization by the harmful dinoflagellate Gymnodinium impudicum (Fraga et Bravo) Hansen et Moestrup isolated from the South Sea of Korea. Under conditions of limited phosphorus, observation of growth kinetics in batch culture yielded a maximum growth rate (?max) of 0.41 /day and a half saturation constant (Ks) of 0.71 ?M. In time-course experiments, APase was induced as dissolved inorganic phosphorus (DIP) concentrations fell below 0.83 ?M, a threshold near the estimated Ks; APase activity increased with further DIP depletion to a maximum of 0.70 pmol/cell/h in the senescent phase. Thus, Ks may be an important index of the threshold DIP concentration for APase induction. G. impudicum utilizes a wide variety of dissolved organic phosphorus compounds in addition to DIP. These results suggest that DIP limitation in the Southern Sea of Korea may have led to the spread of G. impudicum along with the harmful dinoflagellate Cochlodinium polykrikoides in recent years.

Oh, Seok Jin; Kwon, Hyeong Kyu; Noh, Il Hyeon; Yang, Han-Soeb

2010-09-01

249

New oxo-bridged peroxotungsten complexes containing biogenic co-ligand as potent inhibitors of alkaline phosphatase activity.  

Science.gov (United States)

Novel dinuclear peroxo complexes of tungsten with coordinated cystine of the type A(2)[W(2)O(3)(O(2))(4)(cystine)].4H(2)O, A = Na (1) or K (2) have been synthesized from the reaction of A(2)WO(4,)cysteine and 30% H(2)O(2)at pH 2.5. The synthesized compounds were characterized by elemental analysis, spectral and physico-chemical methods. The two W(VI) centres with side-on bound peroxo groups of the dinuclear complex species are bridged by an oxo group and a cystine ligand, formed from the oxidation of cysteine. Cystine occurring as zwitterion binds the metal centers of the complex ion through O(carboxylate) atoms leading to hepta co-ordination around each W(VI). The compounds exhibit high stability toward decomposition in solution of acidic as well as physiological pH and serve as weak substrates to catalase, undergoing degradation in presence of the enzyme at a rate much slower relative to H(2)O(2). The compounds efficiently oxidized GSH to GSSG, a reaction in which only two of the peroxide groups of the complex species were found to participate. The compounds induce strong inhibitory effect on alkaline phosphatase activity with a potency higher than that of the free cystine, tungstate, or peroxotungstate. PMID:16477386

Hazarika, Pankaj; Kalita, Diganta; Sarmah, Swapnalee; Islam, Nashreen S

2006-03-01

250

Yam (Dioscorea batatas) Root and Bark Extracts Stimulate Osteoblast Mineralization by Increasing Ca and P Accumulation and Alkaline Phosphatase Activity.  

Science.gov (United States)

Yam (Dioscorea batatas) is widely consumed as functional food for health promotion mainly in East Asia countries. We assessed whether yam root (tuber) or bark (peel) extracts stimulated the activity of osteoblasts for osteogenesis. MC3T3-E1 cells (mouse osteoblasts) were treated with yam root extracts (water or methanol) (study I) or bark extracts (water or hexane) (study II) within 0~10 ?g/mL during the periods of osteoblast proliferation (5~10 day), matrix maturation (11~15 day) and mineralization (16~20 day) as appropriate. In study I, both yam root water and methanol extracts increased cell proliferation as concentration-dependent manner. Cellular collagen synthesis and alkaline phosphatase (ALP) activity, both the indicators of bone matrix protein and inorganic phosphate production for calcification respectively, were also increased by yam root water and methanol extract. Osteoblast calcification as cell matrix Ca and P accumulation was also increased by the addition of yam root extracts. In study II, yam bark extracts (water and hexane) increased osteoblast proliferation and differentiation, as collagen synthesis and ALP activity and osteoblast matrix Ca and P deposition. The study results suggested that both yam root and bark extracts stimulate osteogenic function in osteoblasts by stimulating bone matrix maturation by increasing collagen synthesis, ALP activity, and matrix mineralization. PMID:25320717

Kim, Suji; Shin, Mee-Young; Son, Kun-Ho; Sohn, Ho-Yong; Lim, Jae-Hwan; Lee, Jong-Hwa; Kwun, In-Sook

2014-09-01

251

In vitro response to alkaline phosphatase coatings immobilized onto titanium implants using electrospray deposition or polydopamine-assisted deposition.  

Science.gov (United States)

Immobilization of biomolecules onto implant surfaces is one of the most straightforward strategies to control the interaction between an implant and its biological environment. Recently, it was shown that the enzyme alkaline phosphatase (ALP) could be efficiently immobilized onto titanium implants in a single step using polydopamine. We hypothesized that such polydopamine-ALP coatings can enhance the early attachment of cells and increase mineralization. Therefore, the current study aimed at immobilization of ALP onto titanium by means of either one- or two-step polydopamine-assisted immobilization or electrospray deposition, the comparative characterization of these experimental substrates and subsequent cell behavioral analysis using primary osteoblast-like cells. Uncoated titanium and ALP-free polydopamine coatings served as controls. Despite significant ALP surface activity and lower water contact for angles ALP-containing surface modifications, only marginal effects on early cell behavior (i.e., cell spreading) and osteogenic differentiation (i.e., proliferation, differentiation and mineralization) were observed in comparison to uncoated titanium. PMID:23640792

Nijhuis, Arnold W G; van den Beucken, Jeroen J J P; Jansen, John A; Leeuwenburgh, Sander C G

2014-04-01

252

A Phos-Tag-Based Fluorescence Quenching System for Activity Assay and Inhibitor Screening for Alkaline Phosphatase  

Directory of Open Access Journals (Sweden)

Full Text Available Fluorescence resonance energy transfer (FRET is a distance-dependent interaction between the electronic excited states of two dye molecules. Here we introduce a novel FRET-based fluorescence quenching system for assaying the activity of alkaline phosphatase (AP by using a phos-phate-binding tag molecule, Phos-tag {1,3-bis[bis(pyridine-2-ylmethylamino]propan-2-olato dizinc(II complex}, attached to a nonfluorescent 4-{[4-(dimethylaminophenyl]diazenyl}benzoyl (Dabcyl: ?max 475 nm dye group. The fluorogenic biomolecule riboflavin 5’-phosphate (FMN: ?em 525 nm was used as an AP substrate. The Dabcyl-labeled Phos-tag specifically captured FMN to form a stable 1:1 complex, resulting in efficient fluorescence quenching. The quenching efficiency was more than 95% for a mixture of 12 ?M FMN and 13.5 ?M Dabcyl-labeled Phos-tag in aqueous solution at pH 7.4 and 25°C. When FMN was dephosphorylated with AP, riboflavin was released into the solution and fluorescence from the flavin moiety appeared. By using this quenching system, we succeeded in detecting time- and dose-dependent dephosphorylation of FMN by AP under near-physiological conditions.

Emiko Kinoshita-Kikuta

2014-08-01

253

Small intestine of artificially reared rat pups: weight gain and changes in alkaline phosphatase, lactase and sucrase activities during development.  

Science.gov (United States)

Comparative studies on small intestinal development in artificially reared (AR) and in mother-fed (MF) rats were carried out. When pups at 12 days of age were reared by continuous intragastric infusing of a cow milk formula (CMF), they gained weight equal to MF rats until 20 days; their small intestine weight was approximately 40 and 23% greater than that of MF rats at 16 days and 22 days of age, respectively. Autoradiographic studies indicated that the intestinal DNA synthetic index was 74-85% higher in AR than in MF rats at 16 days; similarly intestinal crypt depth and duodenal villus length were significantly higher in AR rats. The developmental pattern of duodenal alkaline phosphatase activity was similar in both AR and MF rats. In contrast, AR rats showed precocious decrease of lactase activity in the ileum at 16 days and in the jejunum at 18 days; but total lactase activity in the intestine was not different from MF rats at 16 days. Jejunal sucrase activity in AR rats was precociously induced and elevated. It was concluded that the intestine of AR rats undergoes structural and enzymic modifications. The physiological significance of these changes remains to be elucidated. PMID:6410020

Yeh, K Y

1983-08-01

254

Effects of Colchicine on Localization of Alkaline Phosphatase in McA-RH 7777 Rat Hepatoma Cells  

Science.gov (United States)

We investigated the changes caused by microtubule disruption in cell contact-induced translocation of alkaline phosphatase (ALP) from the Golgi area to the plasma membrane in McA-RH 7777 cells. When the cells were treated with colchicine, the tubular structure of microtubules in the cytoplasm was lost. Colchicine treatment also resulted in the appearance of numerous dots containing mannosidase II (man II) throughout the cytoplasm. Moreover, ALP was distributed in small dots throughout the cytoplasm, as well as in all regions of the plasma membrane, although it was most concentrated at sites of intercellular contact. On the other hand, when the cells were incubated in basal medium after colchicine treatment, large spots containing ALP reappeared in the perinuclear cytoplasm more quickly than the accumulation of small dots containing man II. These findings suggest that colchicine causes disassembly of the Golgi complex into fragments, which scatter throughout the cytoplasm, but that it does not interfere with translocation of ALP to the plasma membrane. Furthermore, cytoplasmic ALP may be localized at sites other than the Golgi complex. PMID:19180199

Taguchi, Meiko; Chida, Kohsuke

2008-01-01

255

Change in Localization of Alkaline Phosphatase and Mannosidase II by Colchicine Treatment of Primary Cultures of Fetal Rat Hepatocytes  

Science.gov (United States)

We examined the changes in localization of alkaline phosphatase (ALP) and mannosidase II (man II), a Golgi marker, after colchicine treatment of primary cultures of fetal rat hepatocytes, using double immunofluorescence staining and confocal laser microscopy. In hepatocytes cultured in basal medium, ALP was localized in the perinuclear cytoplasm, and man II was observed in the Golgi region of the cytoplasm. When hepatocytes were cultured in dexamethasone-supplemented medium, ALP was also localized in the plasma membrane surrounding the bile canaliculus-like structure that was formed between adjacent cells. In hepatocytes cultured in the same medium containing colchicine, the structure of microtubules in the cytoplasm was lost, man II exhibited granular distribution scattering throughout the cytoplasm, and ALP was localized in coarse granular sites of the cytoplasm. However, ALP was not colocalized at the same sites as man II. The present study indicated that colchicine inhibits the dexamethasone-promoted translocation of ALP to the plasma membrane surrounding the bile canaliculus-like structure in primary cultures of fetal rat hepatocytes by disassembling microtubules and discomposing the Golgi complex. PMID:18320021

Chida, Kohsuke; Taguchi, Meiko

2008-01-01

256

Matrix gla protein and alkaline phosphatase are differently modulated in human dermal fibroblasts from PXE patients and controls.  

Science.gov (United States)

Mineralization of elastic fibers in pseudoxanthoma elasticum (PXE) has been associated with low levels of carboxylated matrix gla protein (MGP), most likely as a consequence of reduced vitamin K (vit K) availability. Unexpectedly, vit K supplementation does not exert beneficial effects on soft connective tissue mineralization in the PXE animal model. To understand the effects of vit K supplementation and in the attempt to interfere with pathways leading to the accumulation of calcium and phosphate within PXE-mineralized soft connective tissues, we have conducted in vitro studies on dermal fibroblasts isolated from control subjects and from PXE patients. Cells were cultured in standard conditions and in calcifying medium (CM) in the presence of vit K1 and K2, or levamisole, an alkaline phosphatase (ALP) inhibitor. Control and PXE fibroblasts were characterized by a similar dose-dependent uptake of both vit K1 and vit K2, thus promoting a significant increase of total protein carboxylation in all cell lines. Nevertheless, MGP carboxylation remained much less in PXE fibroblasts. Interestingly, PXE fibroblasts exhibited a significantly higher ALP activity. Consistently, the mineralization process induced in vitro by a long-term culture in CM appeared unaffected by vit K, whereas it was abolished by levamisole. PMID:23223140

Boraldi, Federica; Annovi, Giulia; Vermeer, Cees; Schurgers, Leon J; Trenti, Tommaso; Tiozzo, Roberta; Guerra, Deanna; Quaglino, Daniela

2013-04-01

257

The periplasmic space in Thermus thermophilus: evidence from a regulation-defective S-layer mutant overexpressing an alkaline phosphatase.  

Science.gov (United States)

The presence of a periplasmic space within the cell envelope of Thermus thermophilus was analyzed in a mutant (HB8(Delta)UTR1) defective in the regulation of its S-layer (surface crystalline layer). This mutant forms round multicellular bodies (MBs) surrounded by a common envelope as the culture approaches the stationary phase. Confocal microscopy revealed that the origin of the MBs is the progressive detachment of the external layers coupled with the accumulation of NH(2)-containing material between the external envelopes and the peptidoglycan. A specific pattern of proteins was found as soluble components of the intercellular space of the MBs by a single fractionation procedure, suggesting that they are periplasmic-like components. To demonstrate this, we cloned a gene ( phoA) from T. thermophilus HB8 encoding a signal peptide-wearing alkaline phosphatase (AP), and engineered it to be overexpressed in the mutant from a shuttle vector. Most of the AP activity (>80%) was found as a soluble component of the MBs' intercellular fraction. All these data indicate that Thermus thermophilus actually has a periplasmic space which is functionally similar to that of Proteobacteria. The potential application of the HB8(Delta)UTR1 mutant for the overexpression of periplasmic thermophilic proteins is discussed. PMID:12072958

Castán, Pablo; Zafra, Olga; Moreno, Renata; de Pedro, Miguel A; Vallés, Cristina; Cava, Felipe; Caro, Eddy; Schwarz, Heinz; Berenguer, José

2002-06-01

258

Disparity between circulating and marginated neutrophils: evidence from studies on the granulocyte alkaline phosphatase, a marker of cell maturity.  

Science.gov (United States)

Since circulating and marginated human granulocytes are in rapid kinetic equilibrium, cells of these two compartments have been considered to be a homogeneous population. Our studies on the relationship between neutrophil maturity and the granulocyte alkaline phosphatase (GAP) activity cast doubt on this assumption. After iv administration of hydrocortisone, 12 male volunteers showed an augmentation in circulating granulocytes of 5730 cells/mm3, accompanied by an increase of band neutrophils from 18% to 33% (p less than 0.001). During this influx phase, the GAP activity decreased by 73% when measured cytochemically and by 28% when assayed biochemically (p less than 0.001 and less than 0.01, respectively). When granulocytes were demarginated by epinephrine, the mean count increased by 38%, accompanied by a rise in the portion of segmented forms from 74% to 79% (p less than 0.005) and by an increase of the cytochemical GAP activity by 24% (p less than 0.01). Exact complementary results were obtained during an excessive transient margination, the hemodialysis neutropenia: bands increased from 24% to 54% (p less than 0.02), while the cytochemical GAP dropped by 40% (p less than 0.005). Thus, our analysis of three situations with an acute transient shift of granulocytes indicates that functionally or chronologially "older" cells have a higher GAP activity, and that the transfer of granulocyte from the circulating to the marginal pool is selective. PMID:546224

Fehr, J; Grossmann, H C

1979-01-01

259

Crevicular Alkaline Phosphatase Activity and Rate of Tooth Movement of Female Orthodontic Subjects under Different Continuous Force Applications  

Science.gov (United States)

Purpose. This study is aimed to compare the effects of two different orthodontic forces on crevicular alkaline phosphatase activity, rate of tooth movement, and root resorption. Materials and Methods. Twelve female subjects of class II division 1 malocclusion participated. Maxillary canines with bonded fixed appliances acted as the tested teeth, while their antagonists with no appliances acted as the controls. Canine retraction was performed using nickel titanium coil spring that delivered forces of 100?gm or 150?gm to either side. Crevicular fluid was analyzed for ALP activity, and study models were casted to measure tooth movements. Root resorption was assessed using periapical radiographs before and after the force application. Results. ALP activity at the mesial sites peaked at week 1 for 150?gm group with significant differences when compared with the 100?gm group. Cumulative canine movements were significantly greater in the 150?gm force (2.10 ± 0.50?mm) than in the 100?gm force (1.57 ± 0.44?mm). No root resorption was in the maxillary canines after retraction. Conclusions. A force of 150?gm produced faster tooth movements and higher ALP activity compared with the 100?gm group and had no detrimental effects such as root resorption. PMID:23737787

Megat Abdul Wahab, Rohaya; Md Dasor, Maryati; Senafi, Sahidan; Abang Abdullah, Asma Alhusna; Yamamoto, Zulham; Jemain, Abdul Aziz; Zainal Ariffin, Shahrul Hisham

2013-01-01

260

Use of an alkaline phosphatase-labeled synthetic oligonucleotide probe for detection of Campylobacter jejuni and Campylobacter coli.  

Science.gov (United States)

A commercially available synthetic nucleic acid probe (SNAP) conjugated to alkaline phosphatase was compared with standard culture techniques for detecting Campylobacter species. The SNAP was able to detect either 5 ng of C. jejuni DNA or 10(5) CFU of bacteria. The SNAP could also detect DNA extracted from 10(5) CFU in mock-infected stool samples. The SNAP detected C. jejuni and C. coli but showed no reactivity with C. laridis, C. fetus subsp. fetus, C. fetus subsp. venerealis, C. fennelliae, "C. upsaliensis," C. cinaedii, C. fecalis, C. hyointestinalis, C. mucosalis, or Helicobacter (Campylobacter) pylori. The SNAP also showed no cross-reactivity with other enteric pathogens. When applied to pure cultures, the SNAP detected 55 clinical isolates of C. jejuni and 11 clinical isolates of C. coli, with an accuracy of 100%. When applied directly to clinical specimens, the SNAP detected Campylobacter spp. in 19 of 23 culture-positive stool specimens (sensitivity, 82.6%; specificity, 100%). Pure cultures of the Campylobacter strains isolated from the four probe-negative, culture-positive stool specimens gave positive reactions with the SNAP. While the SNAP had excellent sensitivity and specificity for isolated bacterial colony isolates, the main limitation to the Campylobacter probe detection kit may be the sensitivity limit on direct detection of Campylobacter organisms in stools. PMID:2380381

Olive, D M; Johny, M; Sethi, S K

1990-07-01

 
 
 
 
261

Cissus quadrangularis extract enhances biomineralization through up-regulation of MAPK-dependent alkaline phosphatase activity in osteoblasts.  

Science.gov (United States)

Cissus quadrangularis Linn. has been implicated as therapeutic agent for enhancing bone healing. Though its osteogenic activity has been suggested, the underlying mechanism still remains unclear. In the present study, the effects of ethanol extract of C. quadrangularis (CQ-E) on osteoblast differentiation and function were analyzed using murine osteoblastic cells. The results indicated that mRNA expressions of osteoblast-related genes were not affected by the CQ-E treatment. However, alkaline phosphatase (ALP) activity and the extent of mineralized nodules were significantly increased in treated cells compared with controls. The addition of an extracellular regulated kinase 1/2 inhibitor, a Jun N-terminal kinase 1/2/3 inhibitor and a p38 mitogen-activated protein kinase (MAPK) inhibitor resulted in significantly decreased ALP activity, preferentially by p38 MAPK inhibitor. These results suggested that CQ-E may regulate osteoblastic activity by enhancing ALP activity and mineralization process, and the increased ALP activity effect of CQ-E is likely mediated by MAPK-dependent pathway. PMID:19057968

Parisuthiman, Duenpim; Singhatanadgit, Weerachai; Dechatiwongse, Thaweephol; Koontongkaew, Sitthichai

2009-01-01

262

Pyridoxamine-5-phosphate enzyme-linked immune mass spectrometric assay substrate for linear absolute quantification of alkaline phosphatase to the yoctomole range applied to prostate specific antigen.  

Science.gov (United States)

There is a need to measure proteins that are present in concentrations below the detection limits of existing colorimetric approaches with enzyme-linked immunoabsorbent assays (ELISA). The powerful enzyme alkaline phosphatase conjugated to the highly specific bacterial protein streptavidin binds to biotinylated macromolecules like proteins, antibodies, or other ligands and receptors with a high affinity. The binding of the biotinylated detection antibody, with resulting amplification of the signal by the catalytic production of reporter molecules, is key to the sensitivity of ELISA. The specificity and amplification of the signal by the enzyme alkaline phosphatase in ELISA together with the sensitivity of liquid chromatography electrospray ionization and mass spectrometry (LC-ESI-MS) to detect femtomole to picomole amounts of reporter molecules results in an ultrasensitive enzyme-linked immune mass spectrometric assay (ELIMSA). The novel ELIMSA substrate pyridoxamine-5-phosphate (PA5P) is cleaved by the enzyme alkaline phosphatase to yield the basic and hydrophilic product pyridoxamine (PA) that elutes rapidly with symmetrical peaks and a flat baseline. Pyridoxamine (PA) and (13)C PA were both observed to show a linear relationship between log ion intensity and quantity from picomole to femtomole amounts by liquid chromatography-electrospray ionization and mass spectrometry. Four independent methods, (i) internal (13)C isotope PA dilution curves, (ii) internal (13)C isotope one-point calibration, (iii) external PA standard curve, and (iv) external (13)C PA standard curve, all agreed within 1 digit in the same order of magnitude on the linear quantification of PA. Hence, a mass spectrometer can be used to robustly detect 526 ymol of the alkaline phosphatase streptavidin probe and accurately quantify zeptomole amounts of PSA against log linear absolute standard by micro electrospray on a simple ion trap. PMID:25259405

Florentinus-Mefailoski, Angelique; Marshall, John G

2014-11-01

263

NMR evaluation of the interactions between humic substances and three molecules of agrochemical interest: beta-D-glucosidase and alkaline phosphatase enzymes and glyphosate.  

Digital Repository Infrastructure Vision for European Research (DRIVER)

This doctoral thesis presents results concerning the investigation, by liquid-state Nuclear Magnetic Resonance (NMR) technique, on interactions occurring among humic substances and three molecules largely diffused in soil: the herbicide glyphosate, the enzymes beta-D-Glucosidase and Alkaline Phosphatase. These agrochemical molecules were examined as a function of the increasing addition of humic substances. In detail, glyphosate was treated with both humic and fulvic acids (pH 5 and 7.2); ...

Mazzei, Pierluigi

2011-01-01

264

Detection of mRNA by in situ hybridisation and in northern blot analysis using oligodeoxynucleotide probes labelled with alkaline phosphatase.  

Digital Repository Infrastructure Vision for European Research (DRIVER)

AIMS: To assess whether a reduction in intensity of signal observed using an alkaline phosphatase labelled oligodeoxynucleotide probe could be explained on the basis of procedural steps rather than reduced sensitivity. METHOD: Signal intensity was assessed on in situ hybridisation for pro-opiomelanocortin (POMC) mRNA in rat pituitary and for somatostatin mRNA in human pancreas and in northern blot analysis for POMC mRNA in the presence and absence of formamide. The direct effects of formamide...

Farquharson, M. A.; Harvie, R.; Kennedy, A.; Mcnicol, A. M.

1992-01-01

265

Pretreatment of Rats with ?-tocopherol Alter Liver and Kidney Protein, Alkaline Phosphatase Activity and Phospholipid Profile after 24 Hour Intoxication with Cadmium  

Digital Repository Infrastructure Vision for European Research (DRIVER)

Cell membrane composition and fluidity are altered in diseases and previous reports suggest that membrane lipid is altered in heavy metal toxicity. This study was carried out to assess the effect of cadmium alone and its combination with different doses (75, 150 and 750 mg) of ?-tocophenylacetate on the phospholipid profile and alkaline phosphatase activity in the kidney and liver of rats. The results obtained show that in these tissues, cadmium significantly (p<0.05) increased the levels...

Eriyamremu, G. E.; Adaikpoh, M. A.; Obi, F. O.

2006-01-01

266

Comparación del ultramicrométodo inmunocitoquímico (UMICIQ con el de la fosfatasa alcalina-anti fosfatasa alcalina (APAAP para la cuantificación de subpoblaciones linfocitarias T Comparison of the immunocytochemical ultramicromethod and the alkaline phosphatase - anti-alkaline phosphatase method for the quantification of T lymphocyte subsets  

Directory of Open Access Journals (Sweden)

Full Text Available Se realizó un estudio comparativo entre el método inmunoenzimático que emplea el complejo fofatasa alcalina-anti fofatasa alcalina (APAAP y el ultramicrométodo inmunocitoquímico (UMICIQ utilizado para la detección de marcadores antigénicos y cuantificación de subpoblaciones de linfocitos T. Se estudiaron los antígenos celulares CD3, CD4 y CD8 en 30 individuos adultos supuestamente sanos. Al compararse los resultados por ambas técnicas, se encontraron diferencias estadísticamente significativas para una p A comparative study was conducted on the immunoenzymatic method based on the alkaline phosphatase - anti-alkaline phosphatase complex and the immunocytochemical ultramicromethod used for detecting antigen markers and for the quantification of T-lymphocyte subsets Cell antigens CD3, CD4 and CD8 from 30 apparently healthy adults were studied. When comparing the outcome of both techniques, statistically significant differences were found, (p< 0,05. It was concluded that although the diagnostic efficiency of both methods are similar, the alkaline phosphatase - anti-alkaline phosphatase method is quicker, more economical and less laborious than the immunocytochemical ultramicromethod, so it is recommended as a procedure of choice for these cell studies

Beatriz Socarrás Ferrer

2002-04-01

267

Evaluation of alkaline phosphatase activity and availability of various P fractions for bean (Phaseolus vulgaris in some calcareous soils amended with municipal sewage sludge  

Directory of Open Access Journals (Sweden)

Full Text Available To evaluate the relationship of various P fractions and alkaline phosphatase activity with bean indices growing in 10 calcareous soils, amended with municipal sewage sludge from Chaharmahal-Va-Bakhtiari province, a greenhouse research was carried out. Soil samples were incubated for one month with sludge at a rate equivalent to 1% (w/w. Then, the P fractions, including P adsorbed by Fe and Al oxides (]NaOH+CB]-P, occluded P (CBD-P and P absorbed by Ca (HCl-P, were determined by Olsen and Summers' sequential fractionation procedure. Furthermore, total P, organic P and residual P were determined. Also, alkaline phosphatase activity was measured. A pot experiment in a completely randomized design with three replications in the ten soils was done to evaluate the bean plant indices. The results showed that the amount of P fractions decreased in the following order: HCl-P>residual-P>]NaOH+CB]-P > OP>CBD-P. The results also indicated that alkaline phosphatase activity was significantly correlated with CBD-P fraction, organic P and total P. In addition, significant correlations were found between ([NaOH+CB]-P and HCl-P and plant shoots. In general, the results of this research showed that P fractionation method appears to be a powerful tool to identify the P status and availability in the soils amended with sewage sludge.

T. Raeisi

2014-07-01

268

Pst I restriction fragment length polymorphism of human placental alkaline phosphatase gene: Mendelian in segregation and localization of mutation site in the gene  

International Nuclear Information System (INIS)

The pattern of inheritance of a Pst I restriction fragment length polymorphism (RFLP) of the human placental alkaline phosphatase gene was studied in nine nuclear families by Southern blot hybridization analysis of genomic DNA. The dimorphic RFLP is defined by the presence of allelic fragments 1.0 kilobase and 0.8 kilobase long. The results of this study show that the two alleles of the Pst I RFLP of the placental alkaline phosphatase gene segregate as codominant traits according to Mendelian expectations. For a polymorphism to be useful as a genetic marker the probability that an offspring is informative (PIC) must be at least 0.15. The allelic frequency of the 1.0-kilobase allele is 0.21, which correlates to a probability that an offspring is informative of 0.275 and is indicative of a useful polymorphism. By using probes derived from different regions of the placental alkaline phosphatase cDNA, the mutated Pst I site causing the RFLP was located in the penultimate intron 2497 base pairs downstream from the transcriptional initiation site

269

Changes in Expression of Connexin 32, Bile Canaliculus-Like Structures, and Localization of Alkaline Phosphatase in Primary Cultures of Fetal Rat Hepatocytes  

Science.gov (United States)

We devised an experimental design in primary cultures of fetal rat hepatocytes for studying hepatocyte differentiation over a short period. In the present study, hepatocytes were first cultured for 3 days in dexamethasone-supplemented medium and then for an additional 3 days in dexamethasone- or epidermal growth factor-supplemented medium. In hepatocytes cultured continuously in dexamethasone-supplemented medium, the expression of connexin 32 increased and bile canaliculus-like structures and localization of alkaline phosphatase in the plasma membrane around bile canaliculus-like structures were maintained. Few cells incorporated bromodeoxyuridine. On the other hand, in most of the hepatocytes cultured in epidermal growth factor-supplemented medium, the expression of connexin 32 was minimally recognized, bile canaliculus-like structures were shortened or eliminated, and alkaline phosphatase was localized as numerous fine spots throughout the cytoplasm. More than 20% of all hepatocytes incorporated bromodeoxyuridine. The present study suggests that in hepatocytes, there is a close relationship among connexin 32 expression, the maintenance of bile canaliculus-like structures, and the localization of alkaline phosphatase to the plasma membrane around the bile canaliculus-like structures, and this indicates that the present experimental model is useful for studying hepatocyte differentiation over a short period. PMID:23554539

Fukazawa, Shoko; Chida, Kohsuke; Taguchi, Meiko; Takeuchi, Akihiro; Ikeda, Noriaki

2013-01-01

270

The role of tissue-nonspecific alkaline phosphatase in the phosphate-induced activation of alkaline phosphatase and mineralization in SaOS-2 human osteoblast-like cells.  

Science.gov (United States)

Tissue-nonspecific alkaline phosphatase (TNAP) plays a key role in mineralization by degrading inorganic pyrophosphate and providing free inorganic phosphate. We have previously reported that TNAP is induced by beta-glycerophosphate and NaH(2)PO(4) in short-term cultures of SaOS-2 human osteoblast-like cells and that PHEX (phosphate-regulating gene with homologies to endopeptidase on the X chromosome) mRNA is also induced after TNAP induction. In the present study, we have investigated the effects of levamisole, a TNAP inhibitor, and phosphonoformic acid (PFA), a type III sodium-phosphate cotransporter inhibitor, on the phosphate-induced expression of TNAP and mineralization. Levamisole inhibited beta-glycerophosphate-induced mineralization, TNAP and PHEX expression, and the increase in enzymatic activity of NPP1 (5'-nucleotide pyrophosphatase phosphodiesterase 1), but did not inhibit NaH(2)PO(4)-induced mineralization. PFA completely inhibited NaH(2)PO(4)-induced mineralization and NPP1 enzymatic activation, and partly inhibited beta-glycerophosphate-induced mineralization, but did not affect the increase in TNAP activity. These results suggest that phosphate derived from TNAP-induced hydrolysis of beta-glycerophosphate yields signals that induce TNAP expression and mineralization, and that PHEX expression may be linked to TNAP expression. However, luciferase assays failed to detect any transcriptional activation of the promoter region of the human TNAP gene by beta-glycerophosphate or NaH(2)PO(4), suggesting that the effects of these phosphates may be indirect. PMID:18500657

Orimo, Hideo; Shimada, Takashi

2008-08-01

271

Utilidad de la fosfatasa alcalina urinaria como marcador precoz de lesión tubular renal Utility of urinary alkaline phosphatase as early marker of renal tubular failure  

Directory of Open Access Journals (Sweden)

Full Text Available El objetivo de este estudio fue determinar la actividad de la fosfatasa alcalina urinaria (FALur para evaluar precozmente lesión tubular y su utilidad diagnóstica. Los pacientes estudiados fueron: 20 Controles (C, 11 hipertensos (HTA, 23 diabéticos (DBT y 34 con insuficiencia renal de diverso origen (IRDO. Se realizaron las determinaciones de: creatinina, clearence de creatinina (Jaffé cinético, FAL sérica y urinaria (cinético DGKC, microalbuminuria (inmunoturbidimétrico, proteiunuria (turbidimétrico, uroproteinograma, SDS-PAGE (al 12,5% e isoenzimograma de FAL. La FAL sérica hallada fue normal, sin diferencia entre grupos y sin relación con el aumento de la FALur. El valor de corte recomendado para FALur fue de 8 UI/L. La FALur estuvo elevada en HTA e IRDO y normal en individuos con DBT. Los aumentos de FALur en IRDO se relacionaron con la lesión tubular estructural y en pacientes con HTA podrían relacionarse con alteración tubular precoz. Se propone la determinación de FALur para la detección temprana de lesión tubular ante falla renal establecida o en individuos con riesgo de desarrollarla, y se establece su utilidad en pacientes: - con DBT y HTA para seguimiento (junto a microalbuminuria y clearence de creatinina, - internados en riesgo de insuficiencia renal aguda: para orientar tratamientos, - con insuficiencia renal crónica: como indicador de lesión y pronóstico.The objective of this study was to determine the activity of urinary Alkaline Phosphatase (ALPur to evaluate early tubular failure and its diagnostic usefulness. The patients studied were: 20 Controls (C, 11 with Hipertensión (HTA, 23 Diabetic (DBT and 34 with renal Insufficiency of diverse origin (IRDO. The creatinine, creatinine clearence (kinetic Jaffé serum and urinary ALP (kinetic DGKC, microalbuminuria (Immunoturbidimetric, proteiunuria (Turbidimetric, uroproteinogram, SDS-PAGE (12.5% and ALP isoenzymes determinations were made. The results indicate that serum ALP was normal, without difference between groups, and no relation with the increase in ALPur. Recommended cut-off value of ALPur was 8 UI/L. ALPur was elevated in HTA and IRDO, and normal in DBT. Increases in ALPur in IRDO were related to the structural tubular injury, and those in HTA could be related to early tubular alteration. Determination of ALPur is proposed for early detection of tubular injury, before renal failure is established or when there is risk of developing it, establishing its usefulness in: - DBT and HTA patients: screening (together with microalbuminuria and creatinine clearence. - Hospitalized patients in risk of acute renal insufficiency: in order to orient treatments. - Patients with chronic renal insufficiency: as an indicator of injury and prognosis.

María Beatriz Di Carlo

2007-09-01

272

Utilidad de la fosfatasa alcalina urinaria como marcador precoz de lesión tubular renal / Utility of urinary alkaline phosphatase as early marker of renal tubular failure  

Scientific Electronic Library Online (English)

Full Text Available SciELO Argentina | Language: Spanish Abstract in spanish El objetivo de este estudio fue determinar la actividad de la fosfatasa alcalina urinaria (FALur) para evaluar precozmente lesión tubular y su utilidad diagnóstica. Los pacientes estudiados fueron: 20 Controles (C), 11 hipertensos (HTA), 23 diabéticos (DBT) y 34 con insuficiencia renal de diverso or [...] igen (IRDO). Se realizaron las determinaciones de: creatinina, clearence de creatinina (Jaffé cinético), FAL sérica y urinaria (cinético DGKC), microalbuminuria (inmunoturbidimétrico), proteiunuria (turbidimétrico), uroproteinograma, SDS-PAGE (al 12,5%) e isoenzimograma de FAL. La FAL sérica hallada fue normal, sin diferencia entre grupos y sin relación con el aumento de la FALur. El valor de corte recomendado para FALur fue de 8 UI/L. La FALur estuvo elevada en HTA e IRDO y normal en individuos con DBT. Los aumentos de FALur en IRDO se relacionaron con la lesión tubular estructural y en pacientes con HTA podrían relacionarse con alteración tubular precoz. Se propone la determinación de FALur para la detección temprana de lesión tubular ante falla renal establecida o en individuos con riesgo de desarrollarla, y se establece su utilidad en pacientes: - con DBT y HTA para seguimiento (junto a microalbuminuria y clearence de creatinina), - internados en riesgo de insuficiencia renal aguda: para orientar tratamientos, - con insuficiencia renal crónica: como indicador de lesión y pronóstico. Abstract in english The objective of this study was to determine the activity of urinary Alkaline Phosphatase (ALPur) to evaluate early tubular failure and its diagnostic usefulness. The patients studied were: 20 Controls (C), 11 with Hipertensión (HTA), 23 Diabetic (DBT) and 34 with renal Insufficiency of diverse orig [...] in (IRDO). The creatinine, creatinine clearence (kinetic Jaffé) serum and urinary ALP (kinetic DGKC), microalbuminuria (Immunoturbidimetric), proteiunuria (Turbidimetric), uroproteinogram, SDS-PAGE (12.5%) and ALP isoenzymes determinations were made. The results indicate that serum ALP was normal, without difference between groups, and no relation with the increase in ALPur. Recommended cut-off value of ALPur was 8 UI/L. ALPur was elevated in HTA and IRDO, and normal in DBT. Increases in ALPur in IRDO were related to the structural tubular injury, and those in HTA could be related to early tubular alteration. Determination of ALPur is proposed for early detection of tubular injury, before renal failure is established or when there is risk of developing it, establishing its usefulness in: - DBT and HTA patients: screening (together with microalbuminuria and creatinine clearence). - Hospitalized patients in risk of acute renal insufficiency: in order to orient treatments. - Patients with chronic renal insufficiency: as an indicator of injury and prognosis.

María Beatriz, Di Carlo; Alejandra Gabriela, Gomez; Leticia Bibiana, Madalena; María Laura, Facio; Marco Antonio, Pizzolato; Gustavo Alberto, Negri.

2007-09-01

273

Variaciones de la enzima fosfatasa alcalina en la pulpa dental / Variations of alkaline phosphatase enzyme in the dental pulp  

Scientific Electronic Library Online (English)

Full Text Available SciELO Cuba | Language: Spanish Abstract in spanish En las últimas décadas, numerosas investigaciones se han dedicado al estudio de los mecanismos potenciales implicados en el desarrollo de la caries dental y su prevención, sin embargo, a pesar de haber disminuido gradualmente el índice de caries en la población, son muchos los pacientes que necesita [...] n tratarse la caries dental, tal es así que continuamente se están utilizando diferentes materiales en la búsqueda de aquel que ante una agresión a la pulpa, ayude a una respuesta biológica de la misma, conservando de esta forma su integridad. De ahí la importancia de la actividad de la fosfatasa alcalina de la pulpa en el proceso carioso, como una reacción ante el hidróxido de calcio que continuamente se está usando en toda la red docente-asistencial del país. Se seleccionaron 50 dientes monorradiculares, con pulpa viva y con caries de segundo, tercer y cuarto grado y 50 dientes sanos de pacientes de diferentes edades. Se extrajo la pulpa de cada diente y se realizaron improntas (3 por cada muestra), una de las cuales se procesó para obtener orientación morfológica, y las otras 2 para valorar la actividad de la fosfatasa alcalina. Para esto se utilizaron 2 métodos: el de calcio cobalto y el de alpha naftol fosfato de Gomori. Como resultado, se obtuvo que la pulpa tiene más actividad enzimática en caries profunda y que la edad del paciente no determina el aumento o disminución de dicha actividad. Abstract in english In the last decades, numerous investigations have been made on the study of potential mechanisms involved in the development of dental caries and their prevention. However, in spite of the gradual reduction of dental caries in the population, a lot of patients need to have their dental caries treate [...] d and different materials are continuously used searching for one that before the aggression to the pulp helps it to give a biological response, conserving this way its integrity. That's why the activity of the alkaline phosphatase of the pulp in the caries process is important as a reaction to the calcium hydroxide that is constantly utilized in the teaching-health service network of the country. 50 monoradicular teeth with living pulp and with caries of second, third and fourth degree, and 50 sound teeth from patients of different ages were selected. The pulp of each tooth was extracted and impressions were made (3 per sample). One of them was processed to obtain morphological guidance and the other two to assess the activity of alkaline phosphatase. The cobalt calcium method and Gomori's alpha naphthol phosphate method were used to this end. As a result, it was proved that the pulp has a higher enzymatic activity in deep caries and that the age of the patient does not determine the increase or decrease of this activity.

Zoraida, Pons Pinillos; Nadia, Hernández Rodríguez.

2005-08-01

274

Variaciones de la enzima fosfatasa alcalina en la pulpa dental Variations of alkaline phosphatase enzyme in the dental pulp  

Directory of Open Access Journals (Sweden)

Full Text Available En las últimas décadas, numerosas investigaciones se han dedicado al estudio de los mecanismos potenciales implicados en el desarrollo de la caries dental y su prevención, sin embargo, a pesar de haber disminuido gradualmente el índice de caries en la población, son muchos los pacientes que necesitan tratarse la caries dental, tal es así que continuamente se están utilizando diferentes materiales en la búsqueda de aquel que ante una agresión a la pulpa, ayude a una respuesta biológica de la misma, conservando de esta forma su integridad. De ahí la importancia de la actividad de la fosfatasa alcalina de la pulpa en el proceso carioso, como una reacción ante el hidróxido de calcio que continuamente se está usando en toda la red docente-asistencial del país. Se seleccionaron 50 dientes monorradiculares, con pulpa viva y con caries de segundo, tercer y cuarto grado y 50 dientes sanos de pacientes de diferentes edades. Se extrajo la pulpa de cada diente y se realizaron improntas (3 por cada muestra, una de las cuales se procesó para obtener orientación morfológica, y las otras 2 para valorar la actividad de la fosfatasa alcalina. Para esto se utilizaron 2 métodos: el de calcio cobalto y el de alpha naftol fosfato de Gomori. Como resultado, se obtuvo que la pulpa tiene más actividad enzimática en caries profunda y que la edad del paciente no determina el aumento o disminución de dicha actividad.In the last decades, numerous investigations have been made on the study of potential mechanisms involved in the development of dental caries and their prevention. However, in spite of the gradual reduction of dental caries in the population, a lot of patients need to have their dental caries treated and different materials are continuously used searching for one that before the aggression to the pulp helps it to give a biological response, conserving this way its integrity. That's why the activity of the alkaline phosphatase of the pulp in the caries process is important as a reaction to the calcium hydroxide that is constantly utilized in the teaching-health service network of the country. 50 monoradicular teeth with living pulp and with caries of second, third and fourth degree, and 50 sound teeth from patients of different ages were selected. The pulp of each tooth was extracted and impressions were made (3 per sample. One of them was processed to obtain morphological guidance and the other two to assess the activity of alkaline phosphatase. The cobalt calcium method and Gomori's alpha naphthol phosphate method were used to this end. As a result, it was proved that the pulp has a higher enzymatic activity in deep caries and that the age of the patient does not determine the increase or decrease of this activity.

Zoraida Pons Pinillos

2005-08-01

275

Site-directed mutagenesis maps interactions that enhance cognate and limit promiscuous catalysis by an alkaline phosphatase superfamily phosphodiesterase.  

Science.gov (United States)

Catalytic promiscuity, an evolutionary concept, also provides a powerful tool for gaining mechanistic insights into enzymatic reactions. Members of the alkaline phosphatase (AP) superfamily are highly amenable to such investigation, with several members having been shown to exhibit promiscuous activity for the cognate reactions of other superfamily members. Previous work has shown that nucleotide pyrophosphatase/phosphodiesterase (NPP) exhibits a >10?-fold preference for the hydrolysis of phosphate diesters over phosphate monoesters, and that the reaction specificity is reduced 10³-fold when the size of the substituent on the transferred phosphoryl group of phosphate diester substrates is reduced to a methyl group. Here we show additional specificity contributions from the binding pocket for this substituent (herein termed the R' substituent) that account for an additional ~250-fold differential specificity with the minimal methyl substituent. Removal of four hydrophobic side chains suggested on the basis of structural inspection to interact favorably with R' substituents decreases phosphate diester reactivity 10?-fold with an optimal diester substrate (R' = 5'-deoxythymidine) and 50-fold with a minimal diester substrate (R' = CH?). These mutations also enhance the enzyme's promiscuous phosphate monoesterase activity by nearly an order of magnitude, an effect that is traced by mutation to the reduction of unfavorable interactions with the two residues closest to the nonbridging phosphoryl oxygen atoms. The quadruple R' pocket mutant exhibits the same activity toward phosphate diester and phosphate monoester substrates that have identical leaving groups, with substantial rate enhancements of ~10¹¹-fold. This observation suggests that the Zn²? bimetallo core of AP superfamily enzymes, which is equipotent in phosphate monoester and diester catalysis, has the potential to become specialized for the hydrolysis of each class of phosphate esters via addition of side chains that interact with the substrate atoms and substituents that project away from the Zn²? bimetallo core. PMID:24261692

Wiersma-Koch, Helen; Sunden, Fanny; Herschlag, Daniel

2013-12-23

276

Probing the origin of the compromised catalysis of E. coli alkaline phosphatase in its promiscuous sulfatase reaction.  

Science.gov (United States)

The catalytic promiscuity of E. coli alkaline phosphatase (AP) and many other enzymes provides a unique opportunity to dissect the origin of enzymatic rate enhancements via a comparative approach. Here, we use kinetic isotope effects (KIEs) to explore the origin of the 109-fold greater catalytic proficiency by AP for phosphate monoester hydrolysis relative to sulfate monoester hydrolysis. The primary 18O KIEs for the leaving group oxygen atoms in the AP-catalyzed hydrolysis of p-nitrophenyl phosphate (pNPP) and p-nitrophenylsulfate (pNPS) decrease relative to the values observed for nonenzymatic hydrolysis reactions. Prior linear free energy relationship results suggest that the transition states for AP-catalyzed reactions of phosphate and sulfate esters are "loose" and indistinguishable from that in solution, suggesting that the decreased primary KIEs do not reflect a change in the nature of the transition state but rather a strong interaction of the leaving group oxygen atom with an active site Zn2+ ion. Furthermore, the primary KIEs for the two reactions are identical within error, suggesting that the differential catalysis of these reactions cannot be attributed to differential stabilization of the leaving group. In contrast, AP perturbs the KIE for the nonbridging oxygen atoms in the reaction of pNPP but not pNPS, suggesting a differential interaction with the transferred group in the transition state. These and prior results are consistent with a strong electrostatic interaction between the active site bimetallo Zn2+ cluster and one of the nonbridging oxygen atoms on the transferred group. We suggest that the lower charge density of this oxygen atom on a transferred sulfuryl group accounts for a large fraction of the decreased stabilization of the transition state for its reaction relative to phosphoryl transfer. PMID:17411045

Catrina, Irina; O'Brien, Patrick J; Purcell, Jamie; Nikolic-Hughes, Ivana; Zalatan, Jesse G; Hengge, Alvan C; Herschlag, Daniel

2007-05-01

277

Pueraria mirifica extract and puerarin enhance proliferation and expression of alkaline phosphatase and type I collagen in primary baboon osteoblasts.  

Science.gov (United States)

Phytoestrogen-rich Pueraria mirifica (PM) tuberous extract is a promising candidate for the development of anti-osteoporosis drugs for postmenopausal women, but its action has never been validated in humans or in non-human primates, which are more closely related to humans than rodents. In vitro study of non-human primate osteoblasts is thus fundamental to prepare for in vivo studies of phytoestrogen effects on primate bone. This study aimed to establish a culture system of baboon primary osteoblasts and to investigate the effects of PM extract and its phytoestrogens on these cells. Primary osteoblasts from adult baboon fibulae exhibited osteoblast characteristics in regard to proliferation, differentiation, mineralization, and estrogen receptor expression. They responded to 17?-estradiol by increased proliferation rate and mRNA levels of alkaline phosphatase (ALP), type I collagen, and osteocalcin. After being exposed for 48 h to 100 ?g/ml PM extract, 1000 nM genistein, or 1000 nM puerarin, primary baboon osteoblasts markedly increased the rate of proliferation and mRNA levels of ALP and type I collagen without changes in Runx2, osterix, or osteocalcin expression. PM extract, genistein, and puerarin also decreased the RANKL/OPG ratio, suggesting that they could decrease osteoclast-mediated bone resorption. However, neither PM extract nor its phytoestrogens altered calcium deposition in osteoblast culture. In conclusion, we have established baboon primary osteoblast culture, which is a new tool for bone research and drug discovery. Furthermore, the present results provide substantial support for the potential of PM extract and its phytoestrogens to be developed as therapeutic agents against bone fragility. PMID:25442257

Tiyasatkulkovit, Wacharaporn; Malaivijitnond, Suchinda; Charoenphandhu, Narattaphol; Havill, Lorena M; Ford, Allen L; VandeBerg, John L

2014-10-15

278

Arginine Coordination in Enzymatic Phosphoryl Transfer: Evaluation of the Effect of Arg166 Mutations in Escherichia Coli Alkaline Phosphatase  

International Nuclear Information System (INIS)

Arginine residues are commonly found in the active sites of enzymes catalyzing phosphoryl transfer reactions. Numerous site-directed mutagenesis experiments establish the importance of these residues for efficient catalysis, but their role in catalysis is not clear. To examine the role of arginine residues in the phosphoryl transfer reaction, we have measured the consequences of mutations to arginine 166 in Escherichia coli alkaline phosphatase on hydrolysis of ethyl phosphate, on individual reaction steps in the hydrolysis of the covalent enzyme-phosphoryl intermediate, and on thio substitution effects. The results show that the role of the arginine side chain extends beyond its positive charge, as the Arg166Lys mutant is as compromised in activity as Arg166Ser. Through measurement of individual reaction steps, we construct a free energy profile for the hydrolysis of the enzyme-phosphate intermediate. This analysis indicates that the arginine side chain strengthens binding by ?3 kcal/mol and provides an additional 1-2 kcal/mol stabilization of the chemical transition state. A 2.1 (angstrom) X-ray diffraction structure of Arg166Ser AP is presented, which shows little difference in enzyme structure compared to the wild-type enzyme but shows a significant reorientation of the bound phosphate. Altogether, these results support a model in which the arginine contributes to catalysis through binding interactions and through additional transition state stabilization that mal transition state stabilization that may arise from complementarity of the guanidinum group to the geometry of the trigonal bipyramidal transition state

279

Design and characterization of a recombinant colorimetric SAG1-alkaline phosphatase conjugate to detect specific antibody responses against Toxoplasma gondii.  

Science.gov (United States)

The purpose of this study was to design a novel in vitro tool by using recombinant protein technology to detect specific antibody responses against Toxoplasma gondii in a simple, rapid and highly sensitive reagent. The surface T. gondii SAG1 protein is an important immunodominant target, which provides a great interest as a diagnostic antigen. To further exploit its immunodetection capacity, in the present study, the full length sag1 gene was inserted into the pLIP6 prokaryotic expression vector so as to produce a SAG1 antigen genetically fused to the bacterial alkaline phosphatase (AP). After expression optimization, the recombinant fusion protein folded correctly in soluble form in the periplasmic space and preserved both the AP enzymatic activity and the SAG1 immunoreactivity. Subsequently, direct-ELISA and dot-blot immunoassays were designed, using crude preparation SAG1-AP conjugate, to explore its value in serodiagnosis of human toxoplasmosis. We demonstrate that the recombinant SAG1-AP can detect specific T. gondii antibodies in one-step procedure and can successfully discriminate between T. gondii immune and non-immune patients, in agreement with the standard gold test. In conclusion, the present study shows that the genetic fusion protein provides a new tool for one-step T. gondii immunodetection, which was easily, quickly and reproducibly produced as homogeneous bi-functional reagent. Thus, this recombinant immunoconjugate is a promising marker for Toxoplasma serodiagnosis, requiring further evaluation on a larger series and could provide the basis for direct antibody capture enzyme-immunoassay for specific immunoglobulin M and G detection. PMID:23727049

Chahed Bel-Ochi, Nouha; Bouratbine, Aïda; Mousli, Mohamed

2013-08-30

280

In vitro modulation of alkaline phosphatase activity of Saccharomyces cerevisiae grown in low or high phosphate medium  

Directory of Open Access Journals (Sweden)

Full Text Available Our objective was to characterize the modulation of the activity of Saccharomyces cerevisiae alkaline phosphatases (ALPs by classic inhibitors of ALP activity, cholesterol and steroid hormones, in order to identify catalytic similarities between yeast and mammalian ALPs. S. cerevisiae expresses two ALPs, coded for by the PHO8 and PHO13 genes. The product of the PHO8 gene is repressible by Pi in the medium. ALP activity from yeast (grown in low or high phosphate medium homogenates was determined with p-nitrophenylphosphate as substrate, pH 10.4 (lPiALP or hPiALP, respectively. Activation of hPiALP was observed with 5 mM L-amino acids (L-homoarginine _ 186%, L-leucine _ 155% and L-phenylalanine - 168% and with 1 mM levamisole (122%; percentage values, in comparison to control, of recovered activity. EDTA (5 mM and vanadate (1 mM distinctly inhibited hPiALP (2 and 20%, respectively. L-homoarginine (5 mM had a lower activating effect on lPiALP (166% and was the strongest hPiALP activator. Corticosterone (5 mM inhibited hPiALP to 90%, but no effect was observed in low phosphate medium. Cholesterol, ß-estradiol and progesterone also had different effects on lPiALP and hPiALP. A concentration-dependent activation of lPiALP minus hPiALP was evident with all three compounds, most especially with ß-estradiol and cholesterol. These results do not allow us to identify similarities of the behavior of S. cerevisiae ALPs and any of the mammalian ALPs but allow us to raise the hypothesis of differential regulation of S. cerevisiae ALPs by L-homoarginine, ß-estradiol and cholesterol and of using these compounds to discriminate between S. cerevisiae lPiALP and hPiALP.

J Fernandes

2008-01-01

 
 
 
 
281

Leucine Aminopeptidase, ?-Glucosidase and Alkaline Phosphatase Activity Rates and Their Significance in Nutrient Cycles in Some Coastal Mediterranean Sites  

Directory of Open Access Journals (Sweden)

Full Text Available In aquatic microbial ecology, knowledge of the processes involved in the turnover of organic matter is of utmost importance to understand ecosystem functioning. Microorganisms are major players in the cycling of nutrients (nitrogen, phosphorus and carbon, thanks to their enzymatic activities (leucine aminopeptidase, LAP, alkaline phosphatase, AP, and ?-glucosidase, ?-GLU on organic polymers (proteins, organic phosphates and polysaccharides, respectively. Estimates of the decomposition rates of organic polymers are performed using fluorogenic compounds, whose hydrolysis rate allow us to obtain information on the “potential” metabolic activity of the prokaryotic community. This paper refers the enzyme patterns measured during recent oceanographic cruises performed in some coastal Mediterranean sites, not yet fully investigated in terms of microbial biogeochemical processes. Mean enzyme activity rates ranged from 5.24 to 5558.1 nM/h, from 12.68 to 244.73 nM/h and from 0.006 to 9.51 nM/h for LAP, AP and ?-GLU, respectively. The highest LAP and AP activity rates were measured in the Gulf of Milazzo (Tyrrhenian Sea and in the Straits of Messina, in association with the lowest bacterioplankton abundance; in contrast, the lowest ones were found in the northern Adriatic Sea. ?-GLU was more active in the Straits of Messina. Activity rates were analysed in relation to the main environmental variables. Along the northern Adriatic coastal side affected by the Po river, significant inverse relationships linked LAP and AP with salinity, pointing out that fluvial inputs provided organic substrates for microbial metabolism. Both in the Gulf of Manfredonia and in the Straits of Messina, LAP and AP levels were inversely related with the concentration of nitrate and inorganic phosphorus, respectively. In the Gulf of Milazzo, high cell-specific AP measured in spite of phosphorus availability suggested the role of this enzyme not only in phosphorus, but also in carbon release.

Gabriella Caruso

2010-03-01

282

Effects of metallic ions and diphosphonates on inhibition of pericardial bioprosthetic tissue calcification and associated alkaline phosphatase activity.  

Science.gov (United States)

This study focused on the association of extrinsic alkaline phosphatase (AP) activity with both early and advanced calcification of glutaraldehyde-pretreated bovine pericardial bioprosthetic (GPBP) tissue, and the inhibition of both calcification and AP activity by pre-incubation in diphosphonates (sodium-ethanehydroxydiphosphonate [NaEHDP], aminopropanehydroxydiphosphonate [APD]) and metallic salts (FeCl3, Ga(NO3)3, AICI3). GPBP specimens were implanted subcutaneously in 3 wk old male rats after pre-incubation. Following explantation of the tissue at 72 h and 21 d, calcification was assessed morphologically by light microscopy and chemically by atomic adsorption spectroscopy for calcium content and by molybdate complexation for phosphorus. AP activity was characterized by enzymatic hydrolysis of paranitrophenyl phosphate and by histochemical studies. In both control and pretreated groups, AP levels were greater in 72 h explants than 21 d retrievals, which demonstrated extensive calcification in control explants. All pre-incubations that resulted in inhibition of calcification after 21 d, except for APD, were associated with 72 h AP content which was lower than control specimens. The typical time of initiation of calcification was 72 h, as determined by previous studies with this model system. Covalently bound APD inhibited calcification. Increased AP activity in the APD group may be due to the toxicity of this agent with resultant acute inflammation, or other incompletely understood effects of diphosphonates on calcification and AP. Furthermore, EHDP and Ga3+ incubations were also associated with decreased GPBP AP at 72 h compared to control, but were not effective for inhibiting calcification after 21 d. We concluded that inhibition of peak GPBP AP activity is not necessarily associated with the prevention of GPBP mineralization. PMID:8507781

Hirsch, D; Schoen, F J; Levy, R J

1993-04-01

283

Tartrate-resistant acid phosphatase as a biomarker of bone turnover in dog Fosfatase ácida resistente ao tartarato como biomarcador do metabolismo ósseo no cão  

Digital Repository Infrastructure Vision for European Research (DRIVER)

Values of serum tartrate-resistant acid phosphatase ( TRAP) activity were obtained in adult dogs and its biological variability was assessed. Nine healthy skeletally mature Portuguese Podengo dogs were used for the determination of TRAP, total and bone alkaline phosphatase serum activities, and also to study their relationship with serum minerals, namely calcium (Ca), phosphorous (P), and magnesium (Mg). The serum TRAP activity was 2.19±0.56IU/mL, with intra-individual variation of 18.3% and...

Sousa, C. P.; Nery, F.; Azevedo, J. T.; Viegas, C. A.; Gomes, M. E.; Dias, I. R.

2011-01-01

284

The expression of placental alkaline phosphatase (PLAP) and PLAP-like enzymes in normal and neoplastic human tissues. An immunohistological survey using monoclonal antibodies  

DEFF Research Database (Denmark)

The immunohistological expression of placental alkaline phosphatase (PLAP) and PLAP-like enzyme was studied in frozen sections from a wide variety (n = 254) of normal and malignant tissues using monoclonal antibodies reactive with PLAP (H317) and PLAP/PLAP-like enzyme (H17E2; H315). PLAP/PLAP-like reactivity was seen in normal thymus, and foetal and neonatal testis, and in 21 out of 22 malignant germ cell tumours (GCTs), but was also found in normal endocervix, normal Fallopian tube and in 28 out of 167 non-GCTs (particularly in ovarian and proximal gastrointestinal tract tumours). Positivity for true PLAP (as demonstrated with H317) was seen in term placenta, in endocervix, and in Fallopian tube (but not in other normal tissues) and was commonly found in ovarian and proximal gastrointestinal tract tumours. Reactivity with H317 was unusual in malignant GCTs (2 out of 22 cases). These findings confirm that PLAP/PLAP-like positivity is a highly sensitive immunohistological marker for malignant GCTs, but one which by itself is of only moderate specificity. Furthermore, expression of true PLAP is rare in GCTs and favours instead an origin from the ovary or proximal gastrointestinal tract. The results also indicate that the predominant heat-stable alkaline phosphatase species in normal foetal and neonatal testis, and in thymus has a similar immunohistological profile to that found in malignant GCTs, and is a PLAP-like enzyme ("germ cell alkaline phosphatase") distinct from true PLAP. The occurrence of this marker in GCTs would appear to reflect increased eutopic production of an enzyme present in trace amount in corresponding normal tissues rather than a genuine example of ectopic expression.

Hamilton-Dutoit, Stephen Jacques; Lou, H

1990-01-01

285

iPhos: a toolkit to streamline the alkaline phosphatase-assisted comprehensive LC-MS phosphoproteome investigation  

Science.gov (United States)

Background Comprehensive characterization of the phosphoproteome in living cells is critical in signal transduction research. But the low abundance of phosphopeptides among the total proteome in cells remains an obstacle in mass spectrometry-based proteomic analysis. To provide a solution, an alternative analytic strategy to confidently identify phosphorylated peptides by using the alkaline phosphatase (AP) treatment combined with high-resolution mass spectrometry was provided. While the process is applicable, the key integration along the pipeline was mostly done by tedious manual work. Results We developed a software toolkit, iPhos, to facilitate and streamline the work-flow of AP-assisted phosphoproteome characterization. The iPhos tookit includes one assister and three modules. The iPhos Peak Extraction Assister automates the batch mode peak extraction for multiple liquid chromatography mass spectrometry (LC-MS) runs. iPhos Module-1 can process the peak lists extracted from the LC-MS analyses derived from the original and dephosphorylated samples to mine out potential phosphorylated peptide signals based on mass shift caused by the loss of some multiples of phosphate groups. And iPhos Module-2 provides customized inclusion lists with peak retention time windows for subsequent targeted LC-MS/MS experiments. Finally, iPhos Module-3 facilitates to link the peptide identifications from protein search engines to the quantification results from pattern-based label-free quantification tools. We further demonstrated the utility of the iPhos toolkit on the data of human metastatic lung cancer cells (CL1-5). Conclusions In the comparison study of the control group of CL1-5 cell lysates and the treatment group of dasatinib-treated CL1-5 cell lysates, we demonstrated the applicability of the iPhos toolkit and reported the experimental results based on the iPhos-facilitated phosphoproteome investigation. And further, we also compared the strategy with pure DDA-based LC-MS/MS phosphoproteome investigation. The results of iPhos-facilitated targeted LC-MS/MS analysis convey more thorough and confident phosphopeptide identification than the results of pure DDA-based analysis. PMID:25521246

2014-01-01

286

In vitro modulation of alkaline phosphatase activity of Saccharomyces cerevisiae grown in low or high phosphate medium  

Scientific Electronic Library Online (English)

Full Text Available SciELO Brazil | Language: English Abstract in english Our objective was to characterize the modulation of the activity of Saccharomyces cerevisiae alkaline phosphatases (ALPs) by classic inhibitors of ALP activity, cholesterol and steroid hormones, in order to identify catalytic similarities between yeast and mammalian ALPs. S. cerevisiae expresses two [...] ALPs, coded for by the PHO8 and PHO13 genes. The product of the PHO8 gene is repressible by Pi in the medium. ALP activity from yeast (grown in low or high phosphate medium) homogenates was determined with p-nitrophenylphosphate as substrate, pH 10.4 (lPiALP or hPiALP, respectively). Activation of hPiALP was observed with 5 mM L-amino acids (L-homoarginine _ 186%, L-leucine _ 155% and L-phenylalanine - 168%) and with 1 mM levamisole (122%; percentage values, in comparison to control, of recovered activity). EDTA (5 mM) and vanadate (1 mM) distinctly inhibited hPiALP (2 and 20%, respectively). L-homoarginine (5 mM) had a lower activating effect on lPiALP (166%) and was the strongest hPiALP activator. Corticosterone (5 mM) inhibited hPiALP to 90%, but no effect was observed in low phosphate medium. Cholesterol, ß-estradiol and progesterone also had different effects on lPiALP and hPiALP. A concentration-dependent activation of lPiALP minus hPiALP was evident with all three compounds, most especially with ß-estradiol and cholesterol. These results do not allow us to identify similarities of the behavior of S. cerevisiae ALPs and any of the mammalian ALPs but allow us to raise the hypothesis of differential regulation of S. cerevisiae ALPs by L-homoarginine, ß-estradiol and cholesterol and of using these compounds to discriminate between S. cerevisiae lPiALP and hPiALP.

J, Fernandes; R, Amorim; I, Azevedo; M.J, Martins.

2008-01-01

287

Presence and patterns of alkaline phosphatase activity and phosphorus cycling in natural riparian zones under changing nutrient conditions  

Directory of Open Access Journals (Sweden)

Full Text Available Phosphorus (P is an important limiting nutrient in aquatic ecosystems and knowledge of P cycling is fundamental for reducing harmful algae blooms and other negative effects in water. Despite their importance, the characteristics of P cycling under changing nutrient conditions in shallow lakes were poorly investigated. In this study, in situ incubation experiments were conducted in a natural riparian zone in the main diversion channel used for water transfer into Lake Taihu (Wangyu River. Variations in microbial biomass, dissolved P fractions (organic and inorganic, and alkaline phosphatase activity (bulk APA and specific APA were determined after incubation with and without the addition of P and nitrogen (N (4 total water treatments: +P, +N, +NP, and control. Experiments were conducted during two seasons (late spring and early fall to account for natural differences in nutrient levels that may occur in situ. Our results demonstrated that low levels of DRP may not necessarily indicate P limitation. Phytoplankton exhibited “serial N limitation with P stress” in May, such that chlorophyll a (Chl a increased significantly with N addition, while the limiting nutrient shifted to P in October and phytoplankton biomass increased with P addition. Phytoplankton contributed greatly to APA production and was significantly influenced by P bioavailability, yet high levels of bulk APA were also not necessarily indicative of P limitation. In contrast to phytoplankton, bacteria were less P stressed. As a consequence of enhanced utilization of dissolved reactive P (DRP and dissolved organic P (DOP, +N treatment elevated APA significantly. By contrast, APA could be repressed to low values and phytoplankton converted a large portion of DRP to DOP with P addition. But this was not consistent with bacteria APA (bact-APA in the absence or presence of abundant phytoplankton biomass. The correlation between bulk APA and DRP was good at separate sites and discrepant for the whole data set. Regulation of APA was demonstrated by an inverse hyperbolic relationship between bulk APA, specific APA, and DRP, with a transition from high to low activity occurring between 20 and 50 ?g L-1. This study provides a better understanding of how APA and P cycling change with nutrient perturbations in Lake Taihu system. The obtained results can help understanding the process of P cycling in water and providing a reference for nutrient control in the water transfer project.

Peifang Wang

2014-02-01

288

An evaluation on the activity level of Aspartate aminotransferase and Alkaline phosphatase nzymes in peri-implant sulcus fluid  

Directory of Open Access Journals (Sweden)

Full Text Available Statement of Problem: The correlation between the activity of aspartate aminotransferase (AST and alkaline phosphatase (ALP enzymes in gingival sulcular fluid (GCF with inflammation and periodontal attachment loss has been proved, however there are not adequate studies about dental implants. Purpose: The aim of present study was to investigate the presence and activity level of AST & ALP and their correlation with pocket depth (PD and bleeding of peri-implant slcular fluid (PISF, and to evaluate the possibility of using these assessments as a diagnostic index in oral implantology. Material and Methods: In this study, 41 implants as test group and 41 contralateral teeth as control group, in 21 patients were evaluated. At first visit, the general information about implants and the values of pocket probing depth (PPD, modified sulcus bleeding index (mSBl and modified plaque index (mPI were recorded. At the second visit, samples of GCF/PISF were collected. AST & ALP activity was determined spectrophotometrically and data were analyzed by "t", "Mann-Whitney" tests and Pearson Spearman correlation coefficient."nResults: The results showed that there was a significant difference in the activity of AST between two study groups (P<0.0001. The average activity of ALP in test group was more than control group but the difference was not significant. After elimination of the confounding variables, the average AST in test group was 54.6 (S£=2.3 and in control groups was 44.8 (SE=2.3 (P=0.004. The average ALP in test group (SE=2.2 and in control (SE=2.2 were 36.6 and 35.4, respectively. Values of AST and ALP were positively correlated with other clinical parameters such as PD and mSBI which was significant in test group."nConclusion: The present study suggests that PISF analysis could be considered as a proper diagnostic strategy in the evaluation of dental implant success.

Paknegad M. Assistant Professor

2003-07-01

289

Resveratrol Increases Bone Mineral Density and Bone Alkaline Phosphatase in Obese Men : A Randomized Placebo-Controlled Trial  

DEFF Research Database (Denmark)

Context: Metabolic syndrome (MetS) is associated with low-grade inflammation, which may harmfully affect bone. Resveratrol (RSV) possesses anti-inflammatory properties, and rodent studies suggest bone protective effects. Objective: This study sought to evaluate effects of RSV treatment on bone in men with MetS. Setting and Design: The study was conducted at Aarhus University Hospital as a randomized, double-blinded, placebo-controlled trial assessing changes in bone turnover markers, bone mineral density (BMD), and geometry. Participants: The study population comprised 74 middle-aged obese men with MetS recruited from the general community, of which 66 completed all visits. Mean age of participants was 49.3 ± 6.3 years and mean body mass index was 33.7 ± 3.6 kg/m(2). Intervention: Oral treatment with 1.000 mg RSV (RSVhigh), 150mg RSV (RSVlow), or placebo daily for 16 weeks. Main Outcome Measure: Prespecified primary endpoint was change in bone alkaline phosphatase (BAP). Results: BAP increased dose dependently with RSV (R = 0.471, P < .001), resulting in a significantly greater increase in BAP in the RSVhigh group compared with placebo at all time-points (week 4, 16.4 ± 4.2%, P < .001; week 8, 16.5 ± 4.1%, P < .001; week 16, 15.2 ± 3.7%, P < .001). Lumbar spine trabecular volumetric bone mineral density (LS vBMDtrab) also increased dose dependently with RSV (R = 0.268, P = .036), with a significant increase of 2.6 ± 1.3% in the RSVhigh group compared with placebo (P = .043). In addition, changes in BAP and LS vBMDtrab were positively correlated (R = 0.281, P = .027). No consistent changes were detected in bone density at the hip. Conclusions: Our data suggest that high-dose RSV supplementation positively affects bone, primarily by stimulating formation or mineralization. Future studies of longer duration comprising populations at risk of osteoporosis are needed to confirm these results.

Ornstrup, Marie Juul; HarslØf, Torben

2014-01-01

290

Relationship between bone formation markers bone alkaline phosphatase, osteocalcin and amino-terminal propeptide of type I collagen and bone mineral density in elderly men. Preliminary results.  

Science.gov (United States)

Bone remodeling is altered in all metabolic bone diseases, especially in post-menopausal women and in the elderly. Predicting changes in bone mineral density (BMD) is useful to manage the progression of such diseases and to potentially provide interventions in reducing fracture risk. Continuous bone formation and resorption processes can be monitored by measuring biochemical markers of bone turnover (BTMs) and a relationship between BMD and BTMs has been known for long. The aim of this study was to evaluate the relationship between BMD and serum BTMs bone alkaline phosphatase (BAP), osteocalcin and amino-terminal propeptide of type I collegen (PINP) in elderly (>65 years) men. We prospectively studied 18 elderly men (median age=69, range=65-77 years) with no history of fractures, angina, stroke, myocardial infarction or diabetes mellitus. Patients who had undergone corticosteroid, calcitonin, androgen or bisphosphonate therapy were excluded from the study, as well as those who were vitamin D and calcium supplementation users. All the patients underwent lumbar-spine (L2-L4) dual-energy x-ray absorbtiometry and BMD, BAP, osteocalcin and PINP measurements. The mean BMD and body mass index (BMI) were 0.963±0.04 g/cm(2) and 24.4±1.2 kg/m(2), respectively. BAP, osteocalcin and PINP were 27.8±11.3 U/l, 25.6±7.1 ng/ml and 36.0±7.5 ng/ml, respectively. No correlation was found between BMD and BAP (R=-0.28, p=0.25), osteocalcin (R=-0.18, p=0.48) and PINP (R=-0.21, p=0.39), nor between BMI and both age (R=0.05, p=0.83) and BMD (R=0.10, p=0.67). In conclusion, we did not find any relationship between bone formation markers BAP, osteocalcin and PINP and bone density. Thus, our preliminary data suggest that BTMs are not useful in monitoring the bone mineral status of elderly men. PMID:23160690

Lumachi, Franco; Orlando, Rocco; Fallo, Francesco; Basso, Stefano M M

2012-01-01

291

Effects of dietary vitamin E on mucosal maltase and alkaline phosphatase enzyme activities and on the amount of mucosal malonyldialdehyde in broiler chickens  

Digital Repository Infrastructure Vision for European Research (DRIVER)

The effects of dietary vitamin E levels on mucosal maltase and alkaline phosphatase (ALP) enzyme activities and on the amount of mucosal malonyldialdehyde (MDA) in broiler chickens were studied in the present study. One hundred and eighty of day old male broiler chicks (Ross 308 strain) were randomly assigned into five groups, each with three replicates and 12 chicks in each replicate. Chickens in group A were fed corn-soy- based diet, while those in groups B, C, D and E were fed the same die...

Seyed Hamid Farrokhifar; Ramezan Ali Jafari; Naeem Erfani Majd; Seyed Reza Fatemi Tabatabaee; Mansour Mayahi

2014-01-01

292

Aedes aegypti alkaline phosphatase ALP1 is a functional receptor of Bacillus thuringiensis Cry4Ba and Cry11Aa toxins  

Digital Repository Infrastructure Vision for European Research (DRIVER)

Bacillus thuringiensis subs. israelensis produces at least three Cry toxins (Cry4Aa, Cry4Ba, and Cry11Aa) that are active against Aedes aegypti larvae. Previous work characterized a GPI-anchored alkaline phosphatase (ALP1) as a Cry11Aa binding molecule from the gut of A. aegypti larvae. We show here that Cry4Ba binds ALP1, and that the binding and toxicity of Cry4Ba mutants located in loop 2 of domain II is correlated. Also, we analyzed the contribution of ALP1 towards the toxicity of Cry4Ba ...

Jime?nez, Alan I.; Reyes, Esmeralda Z.; Cancino-rodezno, Angeles; Bedoya-pe?rez, Leidy P.; Caballero-flores, Gustavo G.; Muriel-millan, Luis F.; Likitvivatanavong, Supaporn; Gill, Sarjeet S.; Bravo, Alejandra; Sobero?n, Mario

2012-01-01

293

Production of placental alkaline phosphatase (PLAP) and PLAP-like material by epithelial germ cell and non-germ cell tumours in vitro.  

Digital Repository Infrastructure Vision for European Research (DRIVER)

Placental and placental-like alkaline phosphatase (PLAP) levels in the culture media of 87 cell lines of neoplastic and 'normal' origin were measured by a conventional immunosorbent enzymatic assay (IAEA) and by a new immunoradiometric assay (IRMA). The IRMA detected immunoreactive PLAP in 37 of 80 (46%) human epithelial and germ cell cultures, while the IAEA detected PLAP in only 25 (33%). Of the 52 non-germ cell tumour cultures, the IRMA detected expression in 24 (46%) and the IAEA in only ...

Iles, R. K.; Ind, T. E.; Chard, T.

1994-01-01

294

Role of heat stable fraction of alkaline phosphatase as an adjunct to ca 125 in monitoring patients of epithelial ovarian carcinoma  

Digital Repository Infrastructure Vision for European Research (DRIVER)

Heat stable fraction (HSF) of alkaline phosphatase (ALP) was evaluated as an adjunct to CA 125 as a tumour marker for epithelial ovarian cancer in a follow-up study. In our study group 63.4% of patients had elevated HSF levels (?10U/L) and 93.3% had elevated CA 125 levels (>35U/mL). The sensitivity of CA 125 and HSF was 93.3% and 63.3% respectively. The decline in the activity of HSF, over the pre-op levels was highly significant after the first (p=0.001) chemotherapy cycle and significant ...

Nigam, P. K.; Jain, A.; Goyal, P.; Chitra, R.

2005-01-01

295

Subcellular localisation and properties of histone phosphate phosphatase in human polymorphonuclear leukocytes: alterations in pregnancy and chronic granulocytic leukaemia and relationship to alkaline phosphatase  

International Nuclear Information System (INIS)

Using [32P]histone as substrate, an assay for histone phosphate phosphatase was optimised for human polymorphonuclear leukocytes. Kinetic studies showed that the activity was optimal at pH 6.8, was stimulated by Mn2+ and Co2+, and inhibited by sodium sulphite and zinc chloride. The apparent Ksub(m) of the enzyme for histone phosphate was 0.89 ?mol/l. (Auth.)

296

The evaluation of 25-hydroxy vitamin D, calcium, phosphate and alkaline phosphatase levels in epileptic children under antiepileptic medication  

Directory of Open Access Journals (Sweden)

Full Text Available "n 800x600 Normal 0 false false false EN-US X-NONE AR-SA MicrosoftInternetExplorer4 st1":*{behavior:url(#ieooui } /* Style Definitions */ table.MsoNormalTable {mso-style-name:"Table Normal"; mso-tstyle-rowband-size:0; mso-tstyle-colband-size:0; mso-style-noshow:yes; mso-style-priority:99; mso-style-parent:""; mso-padding-alt:0in 5.4pt 0in 5.4pt; mso-para-margin:0in; mso-para-margin-bottom:.0001pt; mso-pagination:widow-orphan; font-size:10.0pt; font-family:"Times New Roman","serif";} Background: Epilepsy is a common disease in the pediatric neurology. There are frequent anti-epileptic drugs which are used in management of epilepsy. Anti-epileptic drugs may have some complications on bone and vitamin-D metabolism. In this study we aimed to evaluate vitamin-D metabolism in epileptic children."n"nMethods: The study was a prospective and cross sectional one. A total 89 epileptic children who were taking anti-epileptic drugs for longer than six months with no underlying disorder in Imam Khomeini and Bahrami Hospitals in Tehran, Iran were enrolled in our study"n"nResults: Forty nine boys and 40 girls were enrolled in this study; mean age of the patients was 7.8±2.1 years. Mean duration of anti-epileptic drug therapy was 2.3 years (SD=0.4, 70 of patients were under monotherapy and 19 were under polytherapy. None of the patients had signs of rickets. Serum calcium and phosphor levels were within normal ranges. Serum alkaline phosphates levels were increased more than two times in 43%. 42% had vitamin-D deficiency (25-OH Vit D<10 ng/ml and another 33% had vitamin-D insufficiency (10<25-oh Vit D<20 ng/ml. 29 patients (32% were taking prophylactic supplemental Vit D (200-400 IU/day. There was significant difference between patients taking supplemental vitamin-D as prophylaxis and patients who did not (p=0.04. There was no significant difference in vitamin-D levels between patients according to age, gender or different drugs."n"nConclusion: Periodic measurement of 25-hydroxy vitamin-D is recommended in epileptic children taking anti-epileptic dugs. Supplemental vitamin-D administration in such patients may be helpful.

Keyhani doost Z

2011-01-01

297

Interaction between nandrolone decanoate and calcitonin in bone formation markers (osteocalcin and bone specific alkaline phosphatase) and IGF-I in rats.  

Science.gov (United States)

Bone tissue has been shown to contain numerous cell-to-cell signaling peptides called growth factors. These growth factors are thought to have important regulating effects for bone remodeling, due to their potent effects on bone cell metabolism. Our investigation was intended to assess the effect of nandrolone decanoate and calcitonin treatment on biochemical markers of bone formation (bone alkaline phosphatase - osteocalcin) and insulin-like growth factor-I in rats. We studied 48 adult male rats. The animals were divided into four groups. Group (A) served as control. Animals in Group (B) were injected with 4 mg/kg/day nandrolone decanoate. Animals in Group (C) were injected with 400mU/rat/day calcitonin and Group (D) received combined therapy for seven days. Nandrolone decanoate and calcitonin have a mild but significant effect on insulin-like growth factor-I without affecting osteocalcin levels, while calcitonin alone decreases the BALP levels. The coadministration of two agents caused notable elevation on insulin-like growth factor-I, followed by a significant increase of osteocalcin and bone alkaline phosphatase. PMID:15758466

Saranteas, T; Mourouzis, C; Mezitis, M; Tesseromatis, C; Spyraki, C

2001-12-01

298

Serum sialic acid and CEA concentrations in human breast cancer.  

Digital Repository Infrastructure Vision for European Research (DRIVER)

The concentration of bound sialic acid in the sera of 56 normal subjects and 65 subjects with breast cancer was measured, in order to determine (1) whether serum sialic acid concentrations are raised in breast cancer and (2) whether the concentration of sialic acid in serum reflects tumour stage. The amount of sialic acid in serum was compared to serum carcinoembryonic antigen (CEA) values. Urinary hydroxyproline and serum alkaline phosphatase concentrations were used as indicators of bone an...

Hogan-ryan, A.; Fennelly, J. J.; Jones, M.; Cantwell, B.; Duffy, M. J.

1980-01-01

299

A phage-displayed chicken single-chain antibody fused to alkaline phosphatase detects Fusarium pathogens and their presence in cereal grains  

International Nuclear Information System (INIS)

Graphical abstract: A phage-displayed chicken scFv antibody, FvSG7, binds on the surface antigen of conidiospores and the mycelia of F. verticillioides. Its fusion with alkaline phosphatase (AP) through a 218 linker displayed a 4-fold higher affinity compared with the parent scFv antibody and efficiently detected toxigenic Fusarium pathogens in cereal grains. Highlights: ? Generation of a highly reactive scFv antibody against F. verticillioides. ? Localization of the antibody binding to the surface target of F. verticillioides. ? Expression of the antibody–alkaline phosphatase (AP) fusion linked by a 218 linker. ? The antibody–AP fusion has a higher affinity than the parental antibody. ? The antibody–AP fusion detects toxigenic Fusarium pathogens in cereal grains. -- Abstract: Fusarium and its poisonous mycotoxins are distributed worldwide and are of particular interest in agriculture and food safety. A simple analytical method to detect pathogens is essential for forecasting diseases and controlling mycotoxins. This article describes a proposed method for convenient and sensitive detection of Fusarium pathogens that uses the fusion of single-chain variable fragment (scFv) and alkaline phosphatase (AP). A highly reactive scFv antibody specific to soluble cell wall-bound proteins (SCWPs) of F. verticillioides was selected from an immunized chicken phagemid library by phage display. The antibody was verified to bind on the surface of ungerminated conidiospores and mycelia of F. verticillioides. The scFv–AP fusion was constructed, and soluble expression in bacteria was confirmed. Both the antibody properties and enzymatic activity were retained, and the antigen-binding capacity of the fusion was enhanced by the addition of a linker. Surface plasmon resonance measurements confirmed that the fusion displayed 4-fold higher affinity compared with the fusion's parental scFv antibody. Immunoblot analyses showed that the fusion had good binding capacity to the components from SCWPs of F. verticillioides, and enzyme-linked immunosorbent assays revealed that the detection limit of the fungus was below 10?2 ?g mL?1, superior to the scFv antibody. The fusion protein was able to detect fungal concentrations as low as 10?3 mg g?1 of maize grains in both naturally and artificially contaminated samples. Thus, the fusion can be applied in rapid and simple diagnosis of Fusarium contamination in field and stored grain or in food

300

Initiation of mineralization in bioprosthetic heart valves: studies of alkaline phosphatase activity and its inhibition by AlCl3 or FeCl3 preincubations.  

Science.gov (United States)

The principal cause of the clinical failure of bioprosthetic heart valves fabricated from glutaraldehyde-pretreated porcine aortic valves is calcification. Other prostheses composed of tissue-derived and polymeric biomaterials also are complicated by deposition of mineral. We have previously demonstrated that: (a) Failure due to calcification of clinical bioprosthetic valves can be simulated by either a large animal circulatory model or subdermal implants in rodents. (b) Calcification of bioprosthetic tissue has complex host, implant, and mechanical determinants. (c) The initial calcification event in the rat subdermal model is the mineral deposition in devitalized cells intrinsic to the bioprosthetic tissue within 48 to 72 h, followed later by collagen mineralization. (d) Initiation of bioprosthetic tissue mineralization, like that of physiological bone formation, has "matrix vesicles" as early nucleation sites. (e) Alkaline phosphatase (AP), an enzyme also associated with matrix vesicles involved in bone mineral nucleation, is present in both fresh and fixed bioprosthetic tissue at sites of initial mineralization. (f) Certain inhibitors of bioprosthetic tissue calcification (e.g., Al3+, Fe3+) are localized to the sites at which alkaline phosphatase is present. On the basis of these results, we hypothesize that alkaline phosphatase is a key element in the pathogenesis of mineralization of bioprosthetic tissue. In the present studies, we focused on the relationship of AP to early events in calcification, and the inhibition of both calcification and AP activity by FeCl3 and AlCl3 preincubations. Subdermal implants of glutaraldehyde pretreated bovine pericardium (GPBP) were done in 3-week-old rats. AP was characterized by enzymatic hydrolysis of paranitrophenyl phosphate (pnpp), and by histochemical studies. Calcification was evaluated chemically (by atomic adsorption spectroscopy) and morphologically (by light microscopy). The results of these studies are as follows: (a) Extractable AP activity is present in fresh but not glutaraldehyde-pretreated bovine pericardial tissue. However, histochemical studies reveal active AP within the intrinsic devitalized cells of GPBP, despite extended glutaraldehyde incubation. (b) Extrinsic AP is rapidly adsorbed following implantation, with peak activity at 72 h (424 +/- 67.2 nm pnpp/mg protein/min enzyme activity [units]), but markedly lesser amounts at 21 days (96.8 +/- 3.9 units). (c) Simultaneously to the AP activity maximum, bulk calcification is initiated, with GPBP calcium levels rising from 1.2 +/- 0.1 (unimplanted) to 2.4 +/- 0.2 micrograms/mg at 72 h, to 55.6 +/- 3.1 micrograms/mg at 21 days, despite a marked decline in AP activity at this later time.(ABSTRACT TRUNCATED AT 400 WORDS) PMID:1918108

Levy, R J; Schoen, F J; Flowers, W B; Staelin, S T

1991-08-01

 
 
 
 
301

Intestinal alkaline phosphatase activity as a molecular marker of enterotoxicity induced by single dose of 5-fluorouracil and protective role of orally administered glutamine  

Directory of Open Access Journals (Sweden)

Full Text Available Background. One of the critical limitations for the administration of the chemotherapy is the toxicity affecting normal tissue. The main target organs for 5-fluorouracil (5-FU toxicity in humans and experimental animals are the gastrointestinal tract, bone marrow, and skin. The cytotoxic effects of antimetabolite chemotherapy are based on their role as substrates for the same transport processes and enzymes involved in anabolism and catabolism as the natural substrates. The main goal of our study was to analyze the dose-dependent antiproliferative effects of 5-FU on intestinal mucosa, enterotoxic potential of 5-FU in experimental animals and to test possible protective role of glutamine. Methods. In our study, we used Sprague Dawley rats. The control group of rats included 50 animals, while the groups where either 5-fluorouracil (5-FU alone or 5-FU and glutamine were administered included 200 animals. All experimental animals were further stratified according to the experimental model (25 animals in each of 8 experimental subgroups of animals. The 5-FU was administered by intraperitoneal application in single dose of 0, 100, 200, 300, and 400 mg of 5-FU per kg of body weight. Water solution of 1% glutamine was prepared daily and administered orally, in volume of 200 ml, for 7 days continuously, after the 7th day of 5-FU administration. Experimental animals were sacrificed 7 days after the administration of 5-FU. The isolation of enterocytes was performed according to the method of Kralovansky et al. In cell homogenate obtained by described method, we determined the protein content using the Biuret method and the DNA content using the Burton reagent. The activities of enzymes alkaline phosphatase (ALP, glutathione S-transferase (GST, glutathione reductase (GR, and glutathione peroxidase (GPX were determined by kinetic method. All paraffin samples of the small intestine were stained by haematoxiline and eosine(HE method. All the experiments were done in duplicate and analyzed by standard statistical methods. All the experiments were done in duplicate and analyzed by standard statistical methods. Results: Our results of enterotoxicity induced by intraperitonealy administered 5-FU showed statistically significant decrease of DNA content in small intestine samples of experimental animals, decrease in activity of intestinal alkaline phosphatase enzyme and the increase in glutathione-dependent enzymes. The glutamine supplementation reduced 5-FU intestinal toxicity. Conclusion: Intestinal alkaline phosphatase is a good marker of the dose-dependent enterotoxicity induced by 5-fluorouracil.

Bajin-Kati? Katica

2006-01-01

302

A Rapid Lateral Flow Immunoassay for the Detection of Tyrosine Phosphatase-Like Protein IA-2 Autoantibodies in Human Serum  

Science.gov (United States)

Type 1 diabetes (T1D) results from the destruction of pancreatic insulin-producing beta cells and is strongly associated with the presence of islet autoantibodies. Autoantibodies to tyrosine phosphatase-like protein IA-2 (IA-2As) are considered to be highly predictive markers of T1D. We developed a novel lateral flow immunoassay (LFIA) based on a bridging format for the rapid detection of IA-2As in human serum samples. In this assay, one site of the IA-2As is bound to HA-tagged-IA-2, which is subsequently captured on the anti-HA-Tag antibody-coated test line on the strip. The other site of the IA-2As is bound to biotinylated IA-2, allowing the complex to be visualized using colloidal gold nanoparticle-conjugated streptavidin. For this study, 35 serum samples from T1D patients and 44 control sera from non-diabetic individuals were analyzed with our novel assay and the results were correlated with two IA-2A ELISAs. Among the 35 serum samples from T1D patients, the IA-2A LFIA, the in-house IA-2A ELISA and the commercial IA-2A ELISA identified as positive 21, 29 and 30 IA-2A-positive sera, respectively. The major advantages of the IA-2A LFIA are its rapidity and simplicity. PMID:25047039

Kikkas, Ingrid; Mallone, Roberto; Larger, Etienne; Volland, Hervé; Morel, Nathalie

2014-01-01

303

A rapid lateral flow immunoassay for the detection of tyrosine phosphatase-like protein IA-2 autoantibodies in human serum.  

Science.gov (United States)

Type 1 diabetes (T1D) results from the destruction of pancreatic insulin-producing beta cells and is strongly associated with the presence of islet autoantibodies. Autoantibodies to tyrosine phosphatase-like protein IA-2 (IA-2As) are considered to be highly predictive markers of T1D. We developed a novel lateral flow immunoassay (LFIA) based on a bridging format for the rapid detection of IA-2As in human serum samples. In this assay, one site of the IA-2As is bound to HA-tagged-IA-2, which is subsequently captured on the anti-HA-Tag antibody-coated test line on the strip. The other site of the IA-2As is bound to biotinylated IA-2, allowing the complex to be visualized using colloidal gold nanoparticle-conjugated streptavidin. For this study, 35 serum samples from T1D patients and 44 control sera from non-diabetic individuals were analyzed with our novel assay and the results were correlated with two IA-2A ELISAs. Among the 35 serum samples from T1D patients, the IA-2A LFIA, the in-house IA-2A ELISA and the commercial IA-2A ELISA identified as positive 21, 29 and 30 IA-2A-positive sera, respectively. The major advantages of the IA-2A LFIA are its rapidity and simplicity. PMID:25047039

Kikkas, Ingrid; Mallone, Roberto; Larger, Etienne; Volland, Hervé; Morel, Nathalie

2014-01-01

304

Typing and subtyping of haptoglobin from native serum using disc gel electrophoresis in alkaline buffer: application to routine screening.  

Science.gov (United States)

A method with which the six common phenotypes of human haptoglobin can be identified using unseparated serum is described. In contrast to other reported methods, both typing and subtyping of haptoglobin can be performed by polyacrylamide gel electrophoresis in alkaline buffer using 0.1-4.0 microliter of native serum with hemoglobin added. Haptoglobin-hemoglobin complexes are visualized by their peroxidase activity using benzidine and barium peroxide. This relatively inexpensive and fast method seems particularly well suited for the typing and subtyping of haptoglobin from minute amounts in large series of sera and other body fluids and thus may be useful in medical genetics and forensic medicine. PMID:6496936

Linke, R P

1984-08-15

305

Multiwalled carbon nanotube modified screen-printed electrodes for the detection of p-aminophenol: Optimisation and application in alkaline phosphatase-based assays  

International Nuclear Information System (INIS)

Carboxylated multiwalled carbon nanotubes (MWCNT-COOH) were used to modify the working electrode surface of different screen-printed electrodes. The effect of this modification on the electrodic characteristics (double layer capacitance, electroactive area and heterogeneous rate constants for the electron transfer) was evaluated and optimized for the cyclic voltammetric determination of p-aminophenol. The enzymatic hydrolysis of p-aminophenylphosphate was employed for the quantification of alkaline phosphatase, one of the most important label enzymes in immunoassays. Finally, ELISA assays were carried out to quantify pneumolysin using this enzymatic system. Results obtained indicated that low superficial densities of MWCNT-COOH (0.03-0.06 ?g mm-2) yielded the same electrodic improvements but with better analytical properties

306

Microchannel conductivity measurements in microchip for on line monitoring of dephosphorylation rates of organic phosphates using paramagnetic-beads linked alkaline phosphatase.  

Science.gov (United States)

This paper presents the use of polymer coated microelectrodes for the realtime conductivity monitoring in a microchannel photoablated through the polymer without contact. Based on this strategy, a small conductometry sensor has been developed to record in time conductivity variation when an enzymatic reaction occurs through the channel. The rate constant determination, k2, for the dephosphorylation of organic phosphate-alkaline phosphatase-superparamagnetic beads complex using chemically different substrates such as adenosine monoesterphosphate, adenosine diphosphate and adenosine triphosphate was taken as an example to demonstrate selectivity and sensivity of the detection scheme. The k2 value measured for each adenosine phosphate decreases from 39 to 30s(-1) in proportion with the number (3, 2 and 1) of attached phosphate moiety, thus emphasizing the steric hindrance effect on kinetics. PMID:25476378

Kechadi, Mohammed; Sotta, Bruno; Gamby, Jean

2015-01-15

307

Variation of pH in lysed platelet concentrates influence proliferation and alkaline phosphatase activity in human osteoblast-like cells.  

Science.gov (United States)

Activated platelets release a multifaceted blend of growth factors that has stimulatory effects on mesenchymal cells, both in vitro and in vivo, which imply beneficial effects on wound repair and tissue regeneration. Previous studies on fibroblast cultures have revealed that more potent growth factors, with respect to cell proliferation, are released in acidic preparations of lysed platelet concentrates in comparison with neutral and alkaline preparations. The current study was intended to investigate the influence of pH on lysed platelet concentrates with respect to release of growth factors, cell proliferation and alkaline phosphatase (ALP) activity in human osteoblast-like cells (hFOB 1.19). Cell proliferation was assessed with the MTT kit, ALP activity by conventional enzymatic reaction kinetics and growth factors platelet-derived growth factor (PDGF) and transforming growth factor-beta (TGF-beta) by enzyme-linked immunosorbent assays. Osteoblast-like cells were stimulated with lysed platelet concentrates preincubated at pH 4.4, 5.4, 7.4, and 7.6. A 3-13-fold increase of cell proliferation was found in comparison with controls and the most evident increase was observed with platelets activated at pH 5.4. The highest ALP activity was observed in preparations at pH 7.6. Platelets incubated in an acidic environment (pH 5.4) induced a higher proliferation compared with preincubation at neutral or alkaline pH and the level of PDGF was also found to be higher in acidic preincubations. The level of TGF-beta was, in contrast, lowest at pH 4.4. We suggest, based on these experimental findings, that acidic milieu influence platelets to release growth factors more potent to stimulate osteoblast proliferation than neutral and alkaline platelet preparations. Lysed platelet concentrates prepared at an alkaline pH might release additional components with stimulating effects resulting in other features than cell proliferation. This is the first report, to our knowledge, about a pH dependent stimulatory effect of lysed platelet concentrates on human osteoblast-like cell proliferation. Lysed platelet concentrates, preincubated in acidic or alkaline buffers, may benefit fracture healing, implant fixation and might also be advantageous in the treatment of wounds with platelet constituents; however, this has to be investigated in extended experimental and clinical settings. PMID:17365859

Wahlström, Ola; Linder, Cecilia; Kalén, Anders; Magnusson, Per

2007-03-01

308

Improvement of the skeletal and dental hypophosphatasia phenotype in Alpl(-/-) mice by administration of soluble (non-targeted) chimeric alkaline phosphatase.  

Science.gov (United States)

Hypophosphatasia (HPP) results from ALPL gene mutations, which lead to a deficiency of tissue-nonspecific alkaline phosphatase (TNAP), and accumulation of inorganic pyrophosphate, a potent inhibitor of mineralization that is also a natural substrate of TNAP, in the extracellular space. HPP causes mineralization disorders including soft bones (rickets or osteomalacia) and defects in teeth and periodontal tissues. Enzyme replacement therapy using mineral-targeting recombinant TNAP has proven effective in preventing skeletal and dental defects in TNAP knockout (Alpl(-/-)) mice, a model for life-threatening HPP. Here, we show that the administration of a soluble, intestinal-like chimeric alkaline phosphatase (ChimAP) improves the manifestations of HPP in Alpl(-/-) mice. Mice received daily subcutaneous injections of ChimAP at doses of 1, 8 or 16mg/kg, from birth for up to 53days. Lifespan and body weight of Alpl(-/-) mice were normalized, and vitamin B6-associated seizures were absent with 16mg/kg/day of ChimAP. Radiographs, ?CT and histological analyses documented improved mineralization in cortical and trabecular bone and secondary ossification centers in long bones of ChimAP16-treated mice. There was no evidence of craniosynostosis in the ChimAP16-treated mice and we did not detect ectopic calcification by radiography and histology in the aortas, stomachs, kidneys or lungs in any of the treatment groups. Molar tooth development and function improved with the highest ChimAP dose, including enamel, dentin, and tooth morphology. Cementum remained deficient and alveolar bone mineralization was reduced compared to controls, though ChimAP-treated Alpl(-/-) mice featured periodontal attachment and retained teeth. This study provides the first evidence for the pharmacological efficacy of ChimAP for use in the treatment of skeletal and dental manifestations of HPP. PMID:25433339

Gasque, Kellen C S; Foster, Brian L; Kuss, Pia; Yadav, Manisha C; Liu, Jin; Kiffer-Moreira, Tina; van Elsas, Andrea; Hatch, Nan; Somerman, Martha J; Millán, José Luis

2015-03-01

309

Characterization of the proteomic profiles of the brown tide alga Aureoumbra lagunensis under phosphate- and nitrogen-limiting conditions and of its phosphate limitation-specific protein with alkaline phosphatase activity.  

Science.gov (United States)

The persistent bloom of the brown tide alga Aureoumbra lagunensis has been reported in coastal embayments along southern Texas, but the molecular mechanisms that sustain such algal bloom are unknown. We compared the proteome and physiological parameters of A. lagunensis grown in phosphate (P)-depleted, P- and nitrogen (N)-depleted, and nutrient-replete cultures. For the proteomic analysis, samples from three conditions were subjected to two-dimensional electrophoresis and tandem mass spectrometry analysis. Because of the paucity of genomic resources in this species, a de novo cross-species protein search was used to identify the differentially expressed proteins, which revealed their involvement in several key biological processes, such as chlorophyll synthesis, antioxidative protection, and protein degradation, suggesting that A. lagunensis may adopt intracellular nutrient compensation, extracellular organic nutrient regeneration, and damage protection to thrive in P-depleted environments. A highly abundant P limitation-specific protein, tentatively identified as a putative alkaline phosphatase, was further characterized by enzyme activity assay on nondenaturing gel and confocal microscopy, which confirmed that this protein has alkaline phosphatase activity, is a cytoplasmic protein, and is closely associated with the cell membrane. The abundance, location, and functional expression of this alkaline phosphatase all indicate the importance of organic P utilization for A. lagunensis under P limitation and the possible role of this alkaline phosphatase in regenerating phosphate from extra- or intracellular organic phosphorus. PMID:22247172

Sun, Ming-Ming; Sun, Jin; Qiu, Jian-Wen; Jing, Hongmei; Liu, Hongbin

2012-03-01

310

Localization of alkaline and acid phosphatase activity in the intestine of healthy breams (Abramis brama l.) and those infected with plerocercoid of tapeworm Ligula intestinalis (Linné 1758).  

Science.gov (United States)

The examination included 40 breams 4 to 7 years old (18 infected and 22 uninfected). Alkaline and acid phosphatase activity, as shown by the azo-coupling method, has been localized in the oesophagus and the intestine, being the strongest in the epithelium. It was distinctly less intensive in the Lamina propria mucosae and in the intermuscular connective tissue of the oesophagus, in the submucosa, in the cells of AUERBACH plexus and in the blood vessel walls. Besides only the acid phosphatase activity was noted in single (sometimes rather numerous) spherical cells - visible within the epithelium and Lamina propria mucosae. The cells are known as the components of so called "yellow bodies" (melanine macrophage centers) entering particular numerously in the spleen and in the pronephric kidney of infected breams. The activity of both enzymes in the epithelium was considerably weaker in the last third of the intestine, and none in cloaca and in Tunica muscularis all over the length of the intestine (and oesophagus) except for some cells of the connective tissue separating the layers of muscle fibres. No perceptible differences in the activity and localization of both enzymes in the intestine were observed between infected and uninfected fishes examined in different seasons of the year. PMID:409038

Jara, Z; Olech, W; Witala, B

1977-01-01

311

Estado nutricional, consumo de lácteos y niveles séricos de calcio, fósforo y fosfatasas alcalinas en escolares de Mérida / Nutritional status, consumption of dairy products and levels sericos of calcium, phosphorus, and alkaline phosphatases in schoolchildren of Mérida  

Scientific Electronic Library Online (English)

Full Text Available SciELO Venezuela | Language: Spanish Abstract in spanish Se realizó una investigación de Campo de Tipo Descriptiva Correlacional y de corte transversal para determinar el estado nutricional, consumo de lácteos y niveles séricos de calcio, fósforo, y fosfatasas alcalinas en escolares del 1er, 3er y 5to grado de la U.E "Rafael Antonio González" de la comuni [...] dad de Mesa Bolívar en el año 2007. La población estuvo conformada por la matricula escolar de 171 estudiantes. Se determinó la muestra con el método estratificado aleatorio simple, obteniéndose 47% de la matricula escolar, correspondiendo 80 niños distribuidos por grado: 21 niños en 1ero, 28 en 3ero y 31 en 5to, en edades comprendidas entre 6 a 12 años. Se determinó la cantidad y la frecuencia de consumo de productos lácteos para lo cual, se diseñó un cuestionario "ad hoc" contentivo de 10 ítems relacionados con la frecuencia de consumo, cantidad y tipo de lácteos. Se realizó evaluación nutricional a través de la Combinación de Indicadores (Peso para la Talla y Talla para la Edad) utilizando las tablas de Evaluación de la Organización Mundial de la Salud. Se determinaron los valores séricos de calcio, fósforo y fosfatasas alcalinas. Los escolares presentan 32,6% de malnutrición; tanto los niños (6-10 años y 11-12 años) como las niñas (8-12 años) presentaron un porcentaje de adecuación diario de calcio bajo (77,16%, 28,57% y 38,96%) respectivamente y 60% tienen hipocalcemia. Existe significancia estadística entre los niveles séricos de calcio y fósforo con el consumo diario promedio de calcio (p 0,05 y p 0,04). No hubo relación estadísticamente significativa entre el consumo de productos lácteos y el estado nutricional de los escolares. El estado nutricional de los escolares no depende del consumo diario de productos lácteos, sin embargo, dicho consumo si afecta los niveles séricos de calcio y fósforo. Abstract in english A cross-sectional descriptive correlational field research was conducted in order to determine the nutritional status, consumption of milk and serum levels of calcium, phosphorus, and alkaline phosphatase in students of 1st, 3rd and 5th grades of the "Rafael Antonio Gonzalez "school in Mesa Bolívar [...] in 2007. The population consisted of 171 students. We determined the sample with a simple random stratified method, yielding 47% of school enrollment, corresponding to 80 children distributed by grade: 21 children in 1st, 28 in 3rd, 31 in 5th, aged 6 to 12 years old. The amount and frequency of consumption of dairy products, with an "ad hoc" questionnaire designed containing 10 items related to the frequency of consumption, quantity and type of dairy product. Nutritional assessment was carried out by means of the combination of indicators (weight for height and height for age) using the tables of evaluation of the World Health Organization. Values were determined in serum calcium, phosphorus and alkaline phosphatase. The students had 32,6% of malnutrition, both boys (6-10 years and 11-12 years) and girls (8-12 years) had an adequate percentage of low calcium daily intake(77.16%, 28. 57% and 38.96%, respectively) and 60% had hypocalcemia. There is statistical significance between serum calcium and phosphorus with an average daily intake of calcium (p 0.05 and p 0.04). There was no statistically significant relationship between dairy products consumption and nutritional status of schoolchildren. The nutritional status of schoolchildren does not depend on daily consumption of dairy products, however, that consumption does affect serum calcium and phosphorus.

Lizbeth, Rojas; Gladys, Bastardo; Belquis, Sanz; G. Beatriz, Da Silva; Yurimay, Quintero de Rivas; Coromoto, Angarita; Maribel, Prada Briceño.

2011-12-01

312

SEAP expression in transiently transfected mammalian cells grown in serum-free suspension culture  

Digital Repository Infrastructure Vision for European Research (DRIVER)

A transient transfection process was established using a novel 'in-house' developed transfection reagent, Ro-1539. It allows rapid production of large quantities of various recombinant proteins. Here we describe the transient expression of the secreted human placental alkaline phosphatase (SEAP) by HEK293EBNA and CHO cells in serum-free suspension culture. Unexpectedly, high expression levels of SEAP (150 ?g/ml) were found 3–4 days post-transfection when placental alkaline phosphatase (AP)...

Schlaeger, Ernst-ju?rgen; Kitas, Eric A.; Dorn, Arnulf

2003-01-01

313

Targeting the active site of the placental isozyme of alkaline phosphatase by phage-displayed scFv antibodies selected by a specific uncompetitive inhibitor  

Directory of Open Access Journals (Sweden)

Full Text Available Abstract Background The isozymes of alkaline phosphatase, the tissue non-specific, intestinal and placental, have similar properties and a high degree of identity. The placental isozyme (PLAP is an oncofetal antigen expressed in several malignancies including choriocarcinoma, seminoma and ovarian carcinoma. We had earlier attempted to isolate PLAP-specific scFv from a synthetic human immunoglobulin library but were unable to do so, presumably because of the similarity between the isozymes. In this work, we have employed a PLAP-specific uncompetitive inhibitor, L-Phe-Gly-Gly, to select isozyme specific scFvs. An uncompetitive inhibitor binds to the enzyme in the presence of substrate and stabilizes the enzyme-substrate complex. Several uncompetitive inhibitors have varying degrees of isozyme specificity for human alkaline phosphatase isozymes. A specific uncompetitive inhibitor would be able to unmask conformational differences between the otherwise very similar molecules. Also, such inhibitors would be directed to regions at/close to the active site of the enzyme. In this work, the library was first incubated with PLAP and the bound clones then eluted by incubation with L-Phe-Gly-Gly along with the substrate, para-nitro phenyl phosphate (pNPP. The scFvs were then studied with regard to the biochemical modulation of their binding, isozyme specificity and effect on enzyme activity. Results Of 13 clones studied initially, the binding of 9 was inhibited by L-Phe-Gly-Gly (with pNPP and 2 clones were inhibited by pNPP alone. Two clones had absolute and 2 clones had partial specificity to PLAP. Two clones were cross-reactive with only one other isozyme. Three scFv clones, having an accessible His6-tag, were purified and studied for their modulation of enzyme activity. All the three scFvs inhibited PLAP activity with the kinetics of competitive inhibition. Cell ELISA could demonstrate binding of the specific scFvs to the cell surface expressed PLAP. Conclusion The results demonstrate the biochemical modulation of scFv binding. Also, the scFvs bound to the active site and denied the access to the substrate. The selection strategy could generate specific anti-enzyme antibodies to PLAP that can potentially be used for targeting, for modulating enzyme activity in in vitro and in vivo and as probes for the active site. This strategy also has a general application in selecting antibodies from combinatorial libraries to closely related molecules and conformations.

Kala Mrinalini

2005-12-01

314

Correlation between the extent of metastatic lesions in whole body bone scintigraphy of patients with prostatic cancer and prostatic acid phosphatase levels in serum with Eiken PAP RIA kit  

International Nuclear Information System (INIS)

The whole body bone scintigraphy of thirteen patients whose prostatic cancer were histologically confirmed, was processed in four colors, and the bone metastases were quantitatively estimated. On the basis of this estimation, the extent of bone metastases was classified into 4 divisions (grades 0, 1, 2 and 3). And then, the correlation between the extent of bone metastases and prostatic acid phosphatase (PAP), acid phosphatase (AcP) and alkali phosphatase (AlP) levels in serum were investigated

315

Identification of a selective inhibitor of murine intestinal alkaline phosphatase (ML260) by concurrent ultra-high throughput screening against human and mouse isozymes.  

Science.gov (United States)

Alkaline phosphatase (AP) isozymes are present in a wide range of species from bacteria to man and are capable of dephosphorylation and transphosphorylation of a wide spectrum of substrates in vitro. In humans, four AP isozymes have been identified-one tissue-nonspecific (TNAP) and three tissue-specific-named according to the tissue of their predominant expression: intestinal (IAP), placental (PLAP) and germ cell (GCAP) APs. Modulation of activity of the different AP isozymes may have therapeutic implications in distinct diseases and cellular processes. For instance, changes in the level of IAP activity can affect gut mucosa tolerance to microbial invasion due to the ability of IAP to detoxify bacterial endotoxins, alter the absorption of fatty acids and affect ectopurinergic regulation of duodenal bicarbonate secretion. To identify isozyme selective modulators of the human and mouse IAPs, we developed a series of murine duodenal IAP (Akp3-encoded dIAP isozyme), human IAP (hIAP), PLAP, and TNAP assays. High throughput screening and subsequent SAR efforts generated a potent inhibitor of dIAP, ML260, with specificity for the Akp3-, compared to the Akp5- and Akp6-encoded mouse isozymes. PMID:24412070

Ardecky, Robert J; Bobkova, Ekaterina V; Kiffer-Moreira, Tina; Brown, Brock; Ganji, Santhi; Zou, Jiwen; Pass, Ian; Narisawa, Sonoko; Iano, Flávia Godoy; Rosenstein, Craig; Cheltsov, Anton; Rascon, Justin; Hedrick, Michael; Gasior, Carlton; Forster, Anita; Shi, Shenghua; Dahl, Russell; Vasile, Stefan; Su, Ying; Sergienko, Eduard; Chung, Thomas D Y; Kaunitz, Jonathan; Hoylaerts, Marc F; Pinkerton, Anthony B; Millán, José Luis

2014-02-01

316

Copper(II) complexes with cyanoguanidine and o-phenanthroline: Theoretical studies, in vitro antimicrobial activity and alkaline phosphatase inhibitory effect  

Science.gov (United States)

Calculations based on density functional methods are carried out for two Cu(II) complexes with cyanoguanidine (cnge) and o-phenanthroline (o-phen): [Cu(o-phen)2(cnge)](NO3)2?2H2O (1) and [Cu(o-phen)(cnge)(H2O)(NO3)2] (2). The calculated geometrical parameters are in agreement with the experimental values. The results of Atoms in Molecules (AIM) topological analysis of the electron density indicate that the Cu-N(phen) bonds in complex (1) have lower electron density, suggesting that those bonds are stronger in complex (2). Moreover, the ionic character of the Cu-N bond in the complex (1) is slightly stronger than that in the complex (2) and this situation would explain the fact that only complex (2) was stable in water solution. For this reason, the antimicrobial and enzymatic assays were performed using complex (2). It is well known that the increased use of antibiotics has resulted in the development of resistant bacterial and fungal strains. In this context, the study of novel antimicrobial agents has an enormous importance and metal complexes represent an interesting alternative for the treatment of infectious diseases. The aim of this work is to prove the modification of some biological properties like antimicrobial activity or alkaline phosphatase inhibitory activity upon copper complexation.

Martínez Medina, Juan J.; Islas, María S.; López Tévez, Libertad L.; Ferrer, Evelina G.; Okulik, Nora B.; Williams, Patricia A. M.

2014-01-01

317

Effects of dietary vitamin E on mucosal maltase and alkaline phosphatase enzyme activities and on the amount of mucosal malonyldialdehyde in broiler chickens  

Science.gov (United States)

The effects of dietary vitamin E levels on mucosal maltase and alkaline phosphatase (ALP) enzyme activities and on the amount of mucosal malonyldialdehyde (MDA) in broiler chickens were studied in the present study. One hundred and eighty of male day old broiler chicks (Ross 308 strain) were randomly assigned into five groups, each with three replicates and 12 chicks in each replicate. Chickens in group A were fed corn-soy- based diet, while those in groups B, C, D and E were fed the same diet with 20, 60, 180, and 540 mg kg-1 vitamin E supplement (d-alpha tocopherol), respectively. Six birds were randomly chosen from each group, and were euthanized on days 10, 21, 32, and 42 of age. One segment of small intestine outset was homogenized and mucosal ALP and maltase activity were measured. Moreover, mucosal lipid peroxidate amount was measured to reveal the impact of vitamin E on oxidative stress. Maltase activity was increased with the increase of vitamin E up to 60 mg kg-1 of diet while with further levels, it was decreased. Addition of 60 mg kg-1 of vitamin E to the diet significantly increased ALP enzyme activity (p ? 0.001). Addition of 540 mg kg-1 of vitamin E supplement to the diet led to the minimum amount of MDA at 32 days of age. It may be concluded that supplementation of broiler's diet with 60 mg kg-1 of vitamin E can increase mucosal maltase and ALP enzyme activity.

Farrokhifar, Seyed Hamid; Ali Jafari, Ramezan; Erfani Majd, Naeem; Fatemi Tabatabaee, Seyed Reza; Mayahi, Mansour

2013-01-01

318

Development of a nanobody-alkaline phosphatase fusion protein and its application in a highly sensitive direct competitive fluorescence enzyme immunoassay for detection of ochratoxin a in cereal.  

Science.gov (United States)

A rapid and sensitive direct competitive fluorescence enzyme immunoassay (dc-FEIA) for ochratoxin A (OTA) based on a nanobody (Nb)-alkaline phosphatase (AP) fusion protein was developed. The VHH (variable domain of heavy chain antibody) gene of Nb28 was subcloned into the expression vector pecan45 containing the AP double-mutant gene. The Nb28-AP construct was transformed into Escherichia coli BL21(DE3)plysS, and soluble expression in bacteria was confirmed by sodium dodecyl sulfate-polyacrylamide gel electrophoresis and Western blot. Both the Nb properties and AP enzymatic activity were validated by colorimetric and fluorometric analysis. The 50% inhibitory concentration and the detection limit of the dc-FEIA were 0.13 and 0.04 ng/mL, respectively, with a linear range of 0.06-0.43 ng/mL. This assay was compared with LC-MS/MS, and the results indicated the reliability of Nb-AP fusion protein-based dc-FEIA for monitoring OTA contamination in cereal. PMID:25531426

Liu, Xing; Xu, Yang; Wan, De-Bin; Xiong, Yong-Hua; He, Zhen-Yun; Wang, Xian-Xian; Gee, Shirley J; Ryu, Dojin; Hammock, Bruce D

2015-01-20

319

One-Step Detection of Aflatoxin-B(1) Using scFv-Alkaline Phosphatase-Fusion Selected from Human Phage Display Antibody Library  

DEFF Research Database (Denmark)

A unique human phage display library was used to successfully generate a scFv to the highly carcinogenic toxin aflatoxin B1. Such an antibody has major potential applications in therapy and diagnostics. To further exploit its analytical capacity, the scFv was genetically fused to alkaline phosphatase, thereby generating a novel and highly sensitive self-indicating reagent. The performance of this reagent was further characterized, demonstrating its efficacy. The sensitivity of scFv-AP fusion was three-fold better than that of the scFv form. The ability of this human library to generate antibodies to a small hapten was clearly demonstrated and this is linked to its intrinsic diversity, which exceeds many existing conventional human libraries. Our results indicate that demography may influence the diversity of the repertoire of the library in terms of its capacity to generate antibodies to specific targets. Equally, the approach demonstrated should also be applicable for other haptens and larger antigens.

Rangnoi, Kuntalee; Jaruseranee, Nanthnit

2011-01-01

320

Bacillus thuringiensis Cry1AbMod toxin counters tolerance associated with low cadherin expression but not that associated with low alkaline phosphatase expression in Manduca sexta.  

Science.gov (United States)

To exert their toxic effect, Bacillus thuringiensis Cry1Ab toxin undergoes a sequential binding mechanism with different larval gut proteins including glycosyl-phosphatidyl-inositol anchored proteins like aminopeptidase-N (APN) or alkaline-phosphatase (ALP) and a transmembrane cadherin to form pre-pore structures that insert into the membrane. Cadherin binding induces oligomerization of the toxin by facilitating removal of the N-terminal region, while APN/ALP binding helps in oligomer membrane insertion. Cry1AbMod toxin was engineered to lack N-terminal region of the toxin and shown to counter resistance linked to cadherin mutations. In this manuscript we determined the toxicity of Cry1AbMod to Manduca sexta larvae silenced in the expression of cadherin, ALP or APN receptors. As previously reported Cry1Ab toxicity relied principally in ALP and cadherin in comparison to APN. Our data shows that Cry1AbMod counters resistance associated with low cadherin expression but was not effective against ALP silenced larvae. These results show that Cry1AbMod could be effective against resistance insects linked to mutations on binding molecules involved in toxin oligomerization but not against resistant insects linked to mutations on binding molecules involved in oligomer membrane insertion. PMID:25239508

Gómez, Isabel; Flores, Biviana; Bravo, Alejandra; Soberón, Mario

2014-09-17

 
 
 
 
321

Differences in growth and alkaline phosphatase activity between Microcystis aeruginosa and Chlorella pyrenoidosa in response to media with different organic phosphorus  

Directory of Open Access Journals (Sweden)

Full Text Available The growth of Microcystis aeruginosa and Chlorella pyrenoidosa in three dissolved organic phosphorus sources (glucose-1- phosphate, adenosine triphosphate, cyclic-adenosine monophosphate were studied in cultures separated by a dialysis membrane. Results showed that M. aeruginosa and C. pyrenoidosa could utilize those three forms of organic phosphorus, but their growth rates and cell abundances were low in comparison with those in the orthophosphate control. M. aeruginosa had a higher growth rate than C. pyrenoidosa in glucose-1-phosphate, and then became dominate in the separate cultures. In contrast, those two algal species didn’t show any significant differences in the growth rate and cell abundance in the medium with adenosine triphosphate and cyclicadenosine monophosphate. Alkaline phosphatase was an important enzyme for hydrolyzing glucose-1-phosphate, adenosine triphosphate and cyclic-adenosine monophosphate, the activity of which was positively correlated with the growth rate of algae. Considering the big proportion of glucose-1-phosphate in the Lake Taihu, the capability of M. aeruginosa to efficiently utilize this type of organic phosphorus source might be one of reason that why M. aeruginosa is the dominant species in this hyper-eutrophic lake.

Yang YU

2011-02-01

322

Novel bioluminescent assay of alkaline phosphatase using adenosine-3'-phosphate-5'-phosphosulfate as substrate and the luciferin-luciferase reaction and its application.  

Science.gov (United States)

This paper describes a novel bioluminescent assay of alkaline phosphatase (ALP) utilizing ATP-sulfurylase and the luciferin-luciferase reaction. The principle governing the assay is as follows. Adenosine-3'-phosphate-5'-phosphosulfate, which serves as the substrate for ALP, is hydrolyzed enzymatically to produce adenosine-5'-phosphosulfate (APS). APS is converted into ATP by ATP-sulfurylase in the presence of pyrophosphate. The ATP produced is detected by the luciferin-luciferase reaction. The measurable range was 1 zmol to 100 fmol/assay and the detection limit at blank+3 SD was 10 zmol/assay. The coefficient of variation (CV, n=5) was examined at each point of the standard curve; the mean CV percentage was 4.47% (n=6). This assay system was applied to enzyme immunoassay of human chorionic gonadotropin and allele-specific PCR enzyme-linked immunosorbent assay of verotoxin gene using ALP as the label enzyme; 10(-2) mIU/mL hCG in urine and 5 pg of Escherichia coli O157 DNA could be assayed directly and with high sensitivity by the proposed method. PMID:12654306

Arakawa, H; Shiokawa, M; Imamura, O; Maeda, M

2003-03-15

323

Aedes aegypti alkaline phosphatase ALP1 is a functional receptor of Bacillus thuringiensis Cry4Ba and Cry11Aa toxins.  

Science.gov (United States)

Bacillus thuringiensis subs. israelensis produces at least three Cry toxins (Cry4Aa, Cry4Ba, and Cry11Aa) that are active against Aedes aegypti larvae. Previous work characterized a GPI-anchored alkaline phosphatase (ALP1) as a Cry11Aa binding molecule from the gut of A. aegypti larvae. We show here that Cry4Ba binds ALP1, and that the binding and toxicity of Cry4Ba mutants located in loop 2 of domain II is correlated. Also, we analyzed the contribution of ALP1 toward the toxicity of Cry4Ba and Cry11Aa toxins by silencing the expression of this protein though RNAi. Efficient silencing of ALP1 was demonstrated by real-time quantitative PCR (qPCR) and Western blot. ALP1 silenced larvae showed tolerance to both Cry4Ba and Cry11Aa although the silenced larvae were more tolerant to Cry11Aa in comparison to Cry4Ba. Our results demonstrate that ALP1 is a functional receptor that plays an important role in the toxicity of the Cry4Ba and Cry11Aa proteins. PMID:22728570

Jiménez, Alan I; Reyes, Esmeralda Z; Cancino-Rodezno, Angeles; Bedoya-Pérez, Leidy P; Caballero-Flores, Gustavo G; Muriel-Millan, Luis F; Likitvivatanavong, Supaporn; Gill, Sarjeet S; Bravo, Alejandra; Soberón, Mario

2012-09-01

324

A novel role of alkaline phosphatase in the ERK1/2 dephosphorylation in renal cell carcinoma cell lines: A new plausible therapeutic target.  

Science.gov (United States)

Extracellular regulated kinase 1/2 (ERK1/2) has been shown to be activated in renal cell carcinoma (RCC). Previously, we have reported aberrant expression/activity of Liver/Bone/Kidney alkaline phosphatase (L/B/K ALP) in RCC. The present study was conducted to find out whether L/B/K ALP has any role in the dephosphorylation of ERK1/2 in renal cancer cell lines. Two renal cancer cell lines viz. ACHN and A498 were transfected with full length L/B/K ALP cDNA. ALP expression/activity and ERK1/2 phosphorylation were evaluated. Increased L/B/K ALP expression/activity was associated with significantly reduced (P = 0.001) phosphorylation status of ERK1/2 in ALP cDNA transfected cells in comparison to that of control. This is the first study that suggests deactivation of ERK1/2 by stimulation of ALP in renal cancer cell lines which can be used as a therapeutic target of RCC. PMID:25241253

Sharma, Ujjawal; Pal, Deeksha; Prasad, Rajendra

2014-12-01

325

Expanding the spectrum of phenotypes associated with germline PIGA mutations: a child with developmental delay, accelerated linear growth, facial dysmorphisms, elevated alkaline phosphatase, and progressive CNS abnormalities.  

Science.gov (United States)

Phosphatidyl inositol glycan (PIG) enzyme subclasses are involved in distinct steps of glycosyl phosphatidyl inositol anchor protein biosynthesis. Glycolsyl phosphatidyl inositol-anchored proteins have heterogeneous functions; they can function as enzymes, adhesion molecules, complement regulators and co-receptors in signal transduction pathways. Germline mutations in genes encoding different members of the PIG family result in diverse conditions with (severe) developmental delay, (neonatal) seizures, hypotonia, CNS abnormalities, growth abnormalities, and congenital abnormalities as hallmark features. The variability of clinical features resembles the typical diversity of other glycosylation pathway deficiencies such as the congenital disorders of glycosylation. Here, we report the first germline missense mutation in the PIGA gene associated with accelerated linear growth, obesity, central hypotonia, severe refractory epilepsy, cardiac anomalies, mild facial dysmorphic features, mildly elevated alkaline phosphatase levels, and CNS anomalies consisting of progressive cerebral atrophy, insufficient myelinization, and cortical MRI signal abnormalities. X-exome sequencing in the proband identified a c.278C>T (p.Pro93Leu) mutation in the PIGA gene. The mother and maternal grandmother were unaffected carriers and the mother showed 100% skewing of the X-chromosome harboring the mutation. These results together with the clinical similarity of the patient reported here and the previously reported patients with a germline nonsense mutation in PIGA support the determination that this mutation caused the phenotype in this family. PMID:24259184

van der Crabben, Saskia N; Harakalova, Magdalena; Brilstra, Eva H; van Berkestijn, Frédérique M C; Hofstede, Floris C; van Vught, Adrianus J; Cuppen, Edwin; Kloosterman, Wigard; Ploos van Amstel, Hans Kristian; van Haaften, Gijs; van Haelst, Mieke M

2014-01-01

326

High frequencies of elevated alkaline phosphatase activity and rickets exist in extremely low birth weight infants despite current nutritional support  

Directory of Open Access Journals (Sweden)

Full Text Available Abstract Background Osteopenia and rickets are common among extremely low birth weight infants (ELBW, Methods We evaluated all ELBW infants admitted to Texas Children's Hospital NICU in 2006 and 2007. Of 211 admissions, we excluded 98 patients who were admitted at >30 days of age or did not survive/stay for >6 weeks. Bone radiographs obtained in 32 infants were reviewed by a radiologist masked to laboratory values. Results In this cohort of 113 infants, P-APA was found to have a significant inverse relationship with BW, gestational age and serum phosphorus. In paired comparisons, P-APA of infants Conclusion Elevation of P-APA >600 IU/L was very common in ELBW infants. BW was significantly inversely related to both P-APA and radiologic rickets. No single value of P-APA was related to radiological findings of rickets. Given the very high risk of osteopenia and rickets among ELBW infants, we recommend consideration of early screening and early mineral supplementation, especially among infants

Parker Bruce R

2009-07-01

327

Signal-on electrochemical immunoassay for APE1 using ionic liquid doped Au nanoparticle/graphene as a nanocarrier and alkaline phosphatase as enhancer.  

Science.gov (United States)

In this paper, the Au nanoparticles decorated graphene nanosheets (AuNPs/Gr) were prepared as nanocarriers using ionic liquid (IL) as linker reagent. Then the alkaline phosphatase (ALP) and the ferrocene tagged detection antibodies (Fc-Ab2) were loaded on the IL doped AuNPs/Gr as a trace label for ultrasensitive measurements of human apurinic/apyrimidinic endonuclease 1 (APE1), which is a multifunctional protein in the DNA base excision repair pathway relating to various types of cancer. Several labeling protocols were investigated for the determination of the APE1 protein concentration and improved analytical features were obtained with the proposed carriers of IL doped AuNPs/Gr which were labeled with Fc-Ab2 and ALP (ALP/Fc-Ab2/AuNPs/IL/Gr). The reason may be that the IL doped AuNPs/Gr carriers (AuNPs/IL/Gr) could not only enhance the immobilized amount of ALP and Fc-Ab2, but also promote the electron transfer rate. Thus, through the specific recognition of antigen-antibody, numerous ALP/Fc-Ab2/AuNPs/IL/Gr, which are captured onto every single immunocomplex, could further catalyze the ascorbic acid 2-phosphate (AA-p) reaction to amplify the electrochemical signal. Transmission electron microscopy (TEM) images of the AuNPs/IL/Gr nanocomposites revealed the formation of a functionalized surface network structure. The resulting immunosensor exhibited a linear response to APE1 in the concentration range of 0.1-80 pg mL(-1) with a detection limit of 0.04 pg mL(-1), indicating potential applications in clinical diagnostics. PMID:25356934

Zhong, Zhaoyang; Li, Mengxia; Qing, Yi; Dai, Nan; Guan, Wei; Liang, Wei; Wang, Dong

2014-12-21

328

Chicken single-chain antibody fused to alkaline phosphatase detects Aspergillus pathogens and their presence in natural samples by direct sandwich enzyme-linked immunosorbent assay.  

Science.gov (United States)

A sensitive and specific analytical method to detect ubiquitous aflatoxigenic Aspergillus pathogens is essential for monitoring and controlling aflatoxins. Four highly reactive chicken single-chain variable fragments (scFvs) against soluble cell wall proteins (SCWPs) from Aspergillus flavus were isolated by phage display. The scFv antibody AfSA4 displayed the highest activity toward both A. flavus and A. parasiticus and specifically recognized a surface target of their cell walls as revealed by immunofluorescence localization. Molecular modeling revealed a unique compact motif on the antibody surface mainly involving L-CDR2 and H-CDR3. As measured by surface plasmon resonance, AfSA4 fused to alkaline phosphatase had a higher binding capability and 6-fold higher affinity compared with AfSA4 alone. Immunoblot analyses showed that the fusion had good binding capacity to SCWP components from the two fungal species. Direct sandwich enzyme-linked immunosorbent assays with mouse antiaspergillus monoclonal antibody mAb2A8 generated in parallel as a capture antibody revealed that the detection limit of the two fungi was as low as 10(-3) ?g/mL, 1000-fold more sensitive than that reported previously (1 ?g/mL). The fusion protein was able to detect fungal concentrations below 1 ?g/g of maize and peanut grains in both artificially and naturally contaminated samples, with at least 10-fold more sensitivity than that reported (10 ?g/g) thus far. Thus, the fusion can be applied in rapid, simple, and specific diagnosis of Aspergillus contamination in field and stored food/feed commodities. PMID:24128348

Xue, Sheng; Li, He-Ping; Zhang, Jing-Bo; Liu, Jin-Long; Hu, Zu-Quan; Gong, An-Dong; Huang, Tao; Liao, Yu-Cai

2013-11-19

329

Short-term moderate hypothermia stimulates alkaline phosphatase activity and osteocalcin expression in osteoblasts by upregulating Runx2 and osterix in vitro.  

Science.gov (United States)

Exposure of Normal Human Osteoblast cells (NHOst) to a period of hypothermia may interrupt their cellular functions, lead to changes in bone matrix and disrupt the balance between bone formation and resorption, resulting in bone loss or delayed fracture healing. To investigate this possibility, we exposed NHOst cells to moderate (35 °C) and severe (27 °C) hypothermia for 1, 12, 24 and 72 h. The effects of hypothermia with respect to cell cytoskeleton organization, metabolic activity and the expression of cold shock chaperone proteins, osteoblast transcription factors and functional markers, were examined. Our findings showed that prolonged moderate hypothermia retained the polymerization of the cytoskeletal components. NHOst cell metabolism was affected differently according to hypothermia severity. The osteoblast transcription factors Runx2 and osterix were necessary for the transcription and translation of bone matrix proteins, where alkaline phosphatase (Alp) activity and osteocalcin (OCN) bone protein were over expressed under hypothermic conditions. Consequently, bone mineralization was stimulated after exposure to moderate hypothermia for 1 week, indicating bone function was not impaired. The cold shock chaperone protein Rbm3 was significantly upregulated (pRbm3 has a dual function: one as a chaperone protein that stabilizes mRNA transcripts and a second one in enhancing the transcription of Alp and Ocn genes. Our studies demonstrated that hypothermia permitted the in vitro maturation of NHOst cells probably through an osterix-dependent pathway. For that reason, we suggest that moderate hypothermia can be clinically applied to counteract heat production at the fracture site that delays fracture healing. PMID:24928274

Aisha, M D; Nor-Ashikin, M N K; Sharaniza, A B; Nawawi, H M; Kapitonova, M Y; Froemming, G R A

2014-08-01

330

A signal "on" photoelectrochemical biosensor for assay of protein kinase activity and its inhibitor based on graphite-like carbon nitride, Phos-tag and alkaline phosphatase.  

Science.gov (United States)

A highly sensitive and selective photoelectrochemical (PEC) biosensor is fabricated for the detection of protein kinase activity based on visible-light active graphite-like carbon nitride (g-C3N4) and the specific recognition utility of Phos-tag for protein kinase A (PKA)-induced phosphopeptides. For assembling the substrate peptides, g-C3N4 and gold nanoparticles (g-C3N4-AuNPs) complex is synthesized and characterized. When the immobilized peptides on g-C3N4-AuNPs modified ITO electrode are phosphorylated under PKA catalysis, they can be specifically identified and binded with biotin functionalized Phos-tag (Phos-tag-biotin) in the presence of Zn(2+). Then, through the specific interaction between biotin and avidin, avidin functionalized alkaline phosphatase (avidin-ALP) is further assembled to catalyze its substrate of l-ascorbic acid-2-phosphate trisodium salt (AAP) to produce electron donor of ascorbic acid (AA), resulting an increased photocurrent compared with the absence of phosphorylation event. Based on the specific identification effect of Phos-tag, the fabricated biosensor presents excellent selectivity for capturing the phosphorylated serine residues in the substrate peptides. With the good photoactivity of g-C3N4 and ALP-catalyzed signal amplification, the fabricated biosensor presents high sensitivity and low detection limit (0.015 unit/mL, S/N = 3) for PKA. The applicability of this PEC biosensor is further testified by the evaluation of PKA inhibition by HA-1077 with the IC50 value of 1.18?M. This new strategy is also successfully applied to detect the change of PKA activity in cancer cell lysate with and without drug stimulation. Therefore, the developed PEC method has great potential in screening of kinase inhibitors and highly sensitive detection of kinase activity. PMID:25286353

Yin, Huanshun; Sun, Bing; Dong, Linfeng; Li, Bingchen; Zhou, Yunlei; Ai, Shiyun

2015-02-15

331

A search for trace expression of placental-like alkaline phosphatase in non-malignant human tissues: demonstration of its occurrence in lung, cervix, testis and thymus.  

Science.gov (United States)

A search for placental or placental-like human alkaline phosphatase (ALP) was made in human tissues. The tissue extracts were assayed for ALP before and after heating at 65 degrees C for 1 h. Trace amounts of heat-stable ALP activity (greater than 0.01 IU/g) were found in lung, testis, cervix and thymus. The heat-stable ALP in these four tissues gave in Ouchterlony double diffusion plates lines of apparent identity with placental ALP when a rabbit anti-human placental antiserum was used. Inhibition studies with L-phenyl-alanine (Phe), L-homoarginine (Har), L-phenylalanylglycylglycine (Pgg), L-leucine (Leu) and levamisole (Leva), were carried out on the heat-stable ALP and on the total ALP. The heat-stable ALPs from cervix and lung gave [I]50 values with each inhibitor comparable to those of placental ALP. The heat-stable ALPs froM testis and thymus gave [I]50 values for Leu and Pgg which were significantly different from the placental isoenzyme. Electrophoresis of heat-stable lung ALP from different individuals showed polymorphic differences similar to those seen with placental ALPs. Such differences were not seen with heat-stable testis ALP. We conclude that human non-malignant testis, cervix, lung and thymus tissues contain small amounts of placental or placental-like ALPs. The heat-stable ALPs in cervix and lung appear to have the same characteristics as placental ALP and are probably encoded by the same gene locus. The heat stable ALPs in testis and thymus, though immunologically very similar to placental ALP differ from it in inhibition profile and electrophoretically. The significance of the results in relation to the "ectopic' expression of placental and placental-like ALPs in malignancy is discussed. PMID:6814793

Goldstein, D J; Rogers, C; Harris, H

1982-10-13

332

Leucine aminopeptidase, beta-glucosidase and alkaline phosphatase activity rates and their significance in nutrient cycles in some coastal Mediterranean sites.  

Science.gov (United States)

In aquatic microbial ecology, knowledge of the processes involved in the turnover of organic matter is of utmost importance to understand ecosystem functioning. Microorganisms are major players in the cycling of nutrients (nitrogen, phosphorus) and carbon, thanks to their enzymatic activities (leucine aminopeptidase, LAP, alkaline phosphatase, AP, and beta-glucosidase, beta-GLU) on organic polymers (proteins, organic phosphates and polysaccharides, respectively). Estimates of the decomposition rates of organic polymers are performed using fluorogenic compounds, whose hydrolysis rate allow us to obtain information on the "potential" metabolic activity of the prokaryotic community. This paper refers the enzyme patterns measured during recent oceanographic cruises performed in some coastal Mediterranean sites, not yet fully investigated in terms of microbial biogeochemical processes. Mean enzyme activity rates ranged from 5.24 to 5558.1 nM/h, from 12.68 to 244.73 nM/h and from 0.006 to 9.51 nM/h for LAP, AP and beta-GLU, respectively. The highest LAP and AP activity rates were measured in the Gulf of Milazzo (Tyrrhenian Sea) and in the Straits of Messina, in association with the lowest bacterioplankton abundance; in contrast, the lowest ones were found in the northern Adriatic Sea. beta-GLU was more active in the Straits of Messina. Activity rates were analysed in relation to the main environmental variables. Along the northern Adriatic coastal side affected by the Po river, significant inverse relationships linked LAP and AP with salinity, pointing out that fluvial inputs provided organic substrates for microbial metabolism. Both in the Gulf of Manfredonia and in the Straits of Messina, LAP and AP levels were inversely related with the concentration of nitrate and inorganic phosphorus, respectively. In the Gulf of Milazzo, high cell-specific AP measured in spite of phosphorus availability suggested the role of this enzyme not only in phosphorus, but also in carbon release. PMID:20479960

Caruso, Gabriella

2010-01-01

333

Mosaic expression of pluripotency-related proteins oct-3/4 and alkaline phosphatase in human pancreatic carcinoma cell PANC-1  

Directory of Open Access Journals (Sweden)

Full Text Available Most current research on cancer stem cells (CSCs associated with human tumorshas focused on the molecular and cellular analysis of hematopoietic lineagemarkers (e.g., CD44, CD138, and CD 133, which can also serve as important CSC markers in a variety of cancers. However, these markers are generallyexpressed at late stages in embryonic development. Oct-3/4, a member of thefamily of POU-domain transcriptional factors, and alkaline phosphatase (ALP areknown to be expressed in the inner cell mass of blastocysts, germ cells, andpluripotent embryonic stem cells. We thus consider Oct-3/4 and ALP to bepromising markers for CSC. Herein, we examined expression of Oct-3/4 and ALPusing 6 established human pancreatic carcinoma cell lines. RT-PCR analysisrevealed the presence of Oct-3/4 and ALP mRNA in those cells.Immunocytochemical and cytochemical staining revealed that both Oct-3/4 andALP proteins are present as mosaics in PANC-1 cell line, one of those 6 cell lines(23% and 19%, respectively. However, Oct-3/4-positive PANC-1 cells did notexhibit overt ATP-binding cassette transporter G2 (ABCG2 activity, as revealedby Hoechst 33342 dye exclusion assay. Transfection of PANC-1 cells with anOct-3/4 promoter-directed, enhanced green fluorescent protein (EGFP constructconfirmed the presence of Oct-3/4-positive cells. These findings indicate that inPANC-1 cells there are at least 2 subset populations, namely Oct-3/4-positive andALP-positive cells. However, it remains unknown whether expression of these 2markers overlaps. Enrichment of Oct-3/4- or ALP-positive cells by gene transferand subsequent drug selection will be helpful for further characterization of thesecells as possible CSCs.

Masahiro Sato

2013-01-01

334

Low serum levels of 1.25-dihydroxyvitamin D and histomorphometric evidence of osteomalacia after jejunoileal bypass for obesity.  

Digital Repository Infrastructure Vision for European Research (DRIVER)

Twenty-seven unselected patients were investigated three to eight years after jejunoileal bypass for morbid obesity. The serum levels of calcium, magnesium, and phosphorus, and the renal excretions of calcium and magnesium were reduced. The serum alkaline phosphatase levels were increased. The serum levels of the two vitamin D metabolites 25-hydroxyvitamin D (25-OHD) and 1.25-dihydroxyvitamin D (1.25-(OHD)2D) were reduced and inversely related to the increased serum levels of immunoreactive p...

Mosekilde, L.; Melsen, F.; Hessov, I.; Christensen, M. S.; Lund, B. J.; Lund, B. I.; Sørensen, O. H.

1980-01-01

335

Development of a single-chain variable fragment-alkaline phosphatase fusion protein and a sensitive direct competitive chemiluminescent enzyme immunoassay for detection of ractopamine in pork  

International Nuclear Information System (INIS)

Graphical abstract: Detection model of dc-CLEIA based on anti-RAC scFv-AP fusion protein. Highlights: ? The scFv-AP fusion protein against ractopamine (RAC) was produced. ? A dc-CLEIA for RAC was developed based on the purified scFv-AP fusion protein. ? The sensitivity of dc-CLEIA was 10 times as sensitive as dc-ELISA for RAC. ? Recovery tests from pork samples were studied. ? Good accuracy was obtained. - Abstract: A rapid, sensitive chemiluminescent enzyme immunoassay (CLEIA) for ractopamine (RAC) based on a single-chain variable fragment (scFv)-alkaline phosphatase (AP) fusion protein was developed. The scFv gene was prepared by cloning the heavy- and light-chain variable region genes (VH and VL) from hybridoma cell line AC2, which secretes antibodies against RAC, and assembling VH and VL genes with a linker by means of splicing overlap extension polymerase chain reaction. The resulting scFv gene was inserted into the expression vector pLIP6/GN containing AP to produce the fusion protein in Escherichia coli strain BL21. The purified scFv-AP fusion protein was used to develop a direct competitive CLEIA (dcCLEIA) protocol for detection of RAC. The average concentration required for 50% inhibition of binding and the limit of detection of the assay were 0.25 ± 0.03 and 0.02 ± 0.004 ng mL?1, respectively, and the linear response range extended from 0.05 to 1.45 ng mL?1. The assay was 10 ti/sup>. The assay was 10 times as sensitive as the corresponding enzyme-linked immunosorbent assay based on the same fusion protein. Cross-reactivity studies showed that the fusion protein did not cross react with RAC analogs. DcCLEIA was used to analyze RAC spiked pork samples, and the validation was confirmed by high-performance liquid chromatography–tandem mass spectrometry (HPLC–MS). The results showed a good correlation between the data of dc-CLEIA and HPLC–MS (R2 > 0.99), indicating that the assay was an efficient analytical method for monitoring food safety.

336

Development of a single-chain variable fragment-alkaline phosphatase fusion protein and a sensitive direct competitive chemiluminescent enzyme immunoassay for detection of ractopamine in pork  

Energy Technology Data Exchange (ETDEWEB)

Graphical abstract: Detection model of dc-CLEIA based on anti-RAC scFv-AP fusion protein. Highlights: Black-Right-Pointing-Pointer The scFv-AP fusion protein against ractopamine (RAC) was produced. Black-Right-Pointing-Pointer A dc-CLEIA for RAC was developed based on the purified scFv-AP fusion protein. Black-Right-Pointing-Pointer The sensitivity of dc-CLEIA was 10 times as sensitive as dc-ELISA for RAC. Black-Right-Pointing-Pointer Recovery tests from pork samples were studied. Black-Right-Pointing-Pointer Good accuracy was obtained. - Abstract: A rapid, sensitive chemiluminescent enzyme immunoassay (CLEIA) for ractopamine (RAC) based on a single-chain variable fragment (scFv)-alkaline phosphatase (AP) fusion protein was developed. The scFv gene was prepared by cloning the heavy- and light-chain variable region genes (V{sub H} and V{sub L}) from hybridoma cell line AC2, which secretes antibodies against RAC, and assembling V{sub H} and V{sub L} genes with a linker by means of splicing overlap extension polymerase chain reaction. The resulting scFv gene was inserted into the expression vector pLIP6/GN containing AP to produce the fusion protein in Escherichia coli strain BL21. The purified scFv-AP fusion protein was used to develop a direct competitive CLEIA (dcCLEIA) protocol for detection of RAC. The average concentration required for 50% inhibition of binding and the limit of detection of the assay were 0.25 {+-} 0.03 and 0.02 {+-} 0.004 ng mL{sup -1}, respectively, and the linear response range extended from 0.05 to 1.45 ng mL{sup -1}. The assay was 10 times as sensitive as the corresponding enzyme-linked immunosorbent assay based on the same fusion protein. Cross-reactivity studies showed that the fusion protein did not cross react with RAC analogs. DcCLEIA was used to analyze RAC spiked pork samples, and the validation was confirmed by high-performance liquid chromatography-tandem mass spectrometry (HPLC-MS). The results showed a good correlation between the data of dc-CLEIA and HPLC-MS (R{sup 2} > 0.99), indicating that the assay was an efficient analytical method for monitoring food safety.

Dong Jiexian; Li Zhenfeng; Lei Hongtao; Sun Yuanming [Guangdong Provincial Key Laboratory of Food Quality and Safety, South China Agricultural University, Guangzhou 510642 (China); Ducancel, Frederic [CEA, iBiTec-S, Service de Pharmacologie et d' Immnoanalyse (SPI), CEA Saclay, F-91191 Gif sur Yvette (France); Xu Zhenlin [Guangdong Provincial Key Laboratory of Food Quality and Safety, South China Agricultural University, Guangzhou 510642 (China); Boulain, Jean-Claude [CEA, iBiTec-S, Service de Pharmacologie et d' Immnoanalyse (SPI), CEA Saclay, F-91191 Gif sur Yvette (France); Yang Jinyi; Shen Yudong [Guangdong Provincial Key Laboratory of Food Quality and Safety, South China Agricultural University, Guangzhou 510642 (China); Wang Hong, E-mail: gzwhongd@63.com [Guangdong Provincial Key Laboratory of Food Quality and Safety, South China Agricultural University, Guangzhou 510642 (China)

2012-07-29

337

Polymer-anchored peroxo compounds of vanadium(V) and molybdenum(VI): synthesis, stability, and their activities with alkaline phosphatase and catalase.  

Science.gov (United States)

We generated a series of new polymer-bound peroxo complexes of vanadium(V) and molybdenum(VI) of the type [VO(O(2))(2)(sulfonate)]-PSS [PSS = poly(sodium 4-styrene sulfonate)] (PV(3)), [V(2)O(2)(O(2))(4)(carboxylate)VO(O(2))(2)(sulfonate)]-PSSM [PSSM = poly(sodium styrene sulfonate-co-maleate)] (PV(4)), [Mo(2)O(2)(O(2))(4)(carboxylate)]-PA [PA = poly(sodium acrylate)] (PMo(1)), [MoO(O(2))(2)(carboxylate)]-PMA [PMA = poly(sodium methacrylate)] (PMo(2)), and [MoO(O(2))(2)(amide)]-PAm [PAm = poly(acrylamide)] (PMo(3)) by reacting V(2)O(5) (for PV(3) and PV(4)) or H(2)MoO(4) (for PMo(1), PMo(2), and PMo(3)) with H(2)O(2) and the respective water-soluble macromolecular ligand at pH 5-6. The compounds were characterized by elemental analysis (CHN and energy-dispersive X-ray spectroscopy), spectral studies (UV-vis, IR, (13)C NMR, (51)V NMR, and (95) Mo NMR), thermal (TGA) as well as scanning electron micrographs (SEM), and EDX analysis. It has been demonstrated that compounds retain their structural integrity in solutions of a wide range of pH values and are approximately 100 times weaker as substrate to the enzyme catalase relative to H(2)O(2), its natural substrate. The effect of the title compounds, along with previously reported compounds [V(2)O(2)(O(2))(4)(carboxylate)]-PA (PV(1)) and [VO(O(2))(2)(carboxylate)]-PMA (PV(2)) on rabbit intestine alkaline phosphatase (ALP) has been investigated and compared with the effect induced by the free diperoxometallates viz. Na[VO(O(2))(2)(H(2)O)] (DPV), [MoO(O(2))(2)(glycine)(H(2)O)] (DMo(1)), and [MoO(O(2))(2)(asparagine)(H(2)O)] (DMo(2)). It has been observed that although all the compounds tested are potent inhibitors of the enzyme, the polymer-bound and neat complexes act via distinct mechanisms. Each of the macromolecular compounds is a classical noncompetitive inhibitor of ALP. In contrast, the action of neat pV and heteroligand pMo compounds on the enzyme function is consistent with a mixed type of inhibition. PMID:21786762

Boruah, Jeena Jyoti; Kalita, Diganta; Das, Siva Prasad; Paul, Saurav; Islam, Nashreen S

2011-09-01

338

Effects of Aqueous Stem-Bark Extract of Momordica balsamina Linn on Some Serum Enzymes in Normal and Ethanol Fed Rats  

Digital Repository Infrastructure Vision for European Research (DRIVER)

Aqueous stem-bark extract of Momordica balsamina Linn was administered using stomach tubes to normal and alcohol fed rats to study the effect of the extract on organs and tissues by estimating the level of some serum enzymes. The extract was administered for two weeks at a dose of 0.56 mg/100 g body weight. The parameters studied include some serum enzymes (Prostatic and total acid phosphatase, alkaline phosphatase, alanine aminotransferase (ALAT) and aspartate aminotransferase (ASAT))...

Geidam, M. A.; Dauda, E.; Hamza, H. G.

2007-01-01

339

Serum tartrate-resistant acid phosphatase 5b activity as a prognostic marker of survival in breast cancer with bone metastasis  

Directory of Open Access Journals (Sweden)

Full Text Available Abstract Background Serum tartrate-resistant acid phosphatase 5b (TRACP 5b activity is a marker of osteoclast number and is elevated in breast cancer (BC patients with extensive bone metastasis, which might in turn reflect the tumour burden. We tested the hypothesis that baseline serum TRACP 5b activity and its interval change are potential prognostic markers of survival in BC patients with bone metastasis. Methods We analyzed the data from previous prospective studies. A total of 100 patients with newly diagnosed bone metastasis were included. Cox proportional regression model was used to evaluate the correlation between the overall survival time (OS and baseline serum TRACP 5b activity and its interval changes. The least significant change (LSC of TRACP 5b was calculated from data obtained from 15 patients with early BC. Results Estrogen receptor status (Hazard Ratio (HR = 0.397; p = 0.003 and visceral metastasis (HR = 0.492; p = 0.0045 were significantly correlated with OS. The OS was significantly shorter in those patients with higher baseline TRACP 5b activity based on a cut-off value to delineate the highest tertile (HR = 3.524; p p = 0.0015. Conclusions We found that TRACP 5b activity and its interval change after treatment bore a prognostic role in BC patients with bone metastasis and a high baseline serum TRACP 5b activity. Further prospective phase II study is necessary to confirm these results.

Liu Hsin-Yi

2010-04-01

340

Serum and pancreatic juice carcinoembryonic antigen in pancreatic and biliary disease.  

Digital Repository Infrastructure Vision for European Research (DRIVER)

Serum and pancreatic juice carcinoembryonic antigen (CEA) concentrations were studied in a group of 144 patients undergoing endoscopic retrograde cholangiopancreatography (ERCP) with a variety of benign and malignant pancreatic and biliary diseases. Serum CEA was found to be a poor diagnostic and discriminating marker for pancreatic disorders and was raised in obstructive jaundice from various causes correlating with serum alkaline phosphatase. A pancreatic juice CEA concentration of greater ...

Carr-locke, D. L.

1980-01-01

 
 
 
 
341

Fosfatasa alcalina intestinal: una enzima con propiedades antiinflamatorias / Intestinal alkaline phosphatase: an enzyme with anti-inflammatory properties / Fosfatasse alcalina intestinal: uma enzima com propriedades anti-inflamatórias  

Scientific Electronic Library Online (English)

Full Text Available SciELO Colombia | Language: Spanish Abstract in portuguese Resumo Uma das principais funções da Fosfatasse Alcalina Intestinal (FAI) é a detoxificação dos lipopolissacarídeos (LPS) bacterianos para controlar a inflamação intestinal. Recentes publicações indicam que a FAI participa na detoxificação de outros compostos bacterianos (flagelina e motivos CpG do [...] DNA) e de muitos nucleotídeos libres (ATP, UDP). A FAI está involucrada de forma direita na recuperação tissular da inflamação pela Resolvina E1 (RvE1). A ação antiinflamatória da FAI melhora indiretamente a função da barreira intestinal e impacta a diversidade e a composição da microbiota. Diversas doenças intestinais, incluindo enterocolitis necrótica, doença celíaca e a inflamação crônica intestinal (inflammatory bowel disease, IBD) estão relacionados com diminuições na expressão e atividade da FAI. De outro jeito, uma elevada atividade da FAI no cólon é sinônimo de processos inflamatórios, devido a elevada concentração da isoforma tissular da Fosfatasse Alcalina não especifica (FANE), e a infiltração tissular pelos neutrófilos (que também contém FANE). A administração exógena da FAI reduz a inflamação intestinal/sistêmica (dependendo da via de administração) incluindo uns poucos testes no homem. Em conclusão, a homeostase intestinal e a preservação da saúde dependem em grande medida da capacidade da FAI para detoxificar os LPS e suprimir a inflamação metabólica induzida por estes. Embora, é preciso realizar pesquisas bem feitas sobre como os costumes alimentares podem modificar a detoxificação dos diferentes compostos proinflamatórios bacterianos e maximizar a Abstract in spanish Resumen Una de las principales funciones de la Fosfatasa Alcalina Intestinal (FAI) es la detoxificación de los lipopolisacáridos (LPS) bacterianos para controlar la inflamación intestinal. Recientes publicaciones indican que FAI participa en la detoxificación de otros compuestos bacterianos (flageli [...] na y motivos CpG de DNA) y de muchos nucleótidos libres (ATP, UDP). FAI está involucrada de manera directa en la recuperación tisular de la inflamación por la Resolvina E1. La acción antiinflamatoria de FAI mejora indirectamente la función de la barrera intestinal e impacta la diversidad y la composición de la microbiota. Diversas enfermedades intestinales, incluyendo enterocolitis necrótica, enfermedad celíaca y la inflamación crónica intestinal (o inflammatory bowel disease, IBD) están relacionadas con disminuciones en la expresión y actividad de FAI. Por otro lado, una elevada actividad de FAI en colon es sinónimo de procesos inflamatorios, debido a la elevada concentración de la isoforma tisular de Fosfatasa Alcalina no específica (FANE), y a la infiltración tisular por los neutrófilos (que también contienen FANE). En algunos ensayos en humanos se ha observado que la administración exógena de FAI reduce la inflamación intestinal/sistémica (dependiendo de la vía de administración). En conclusión, la homeóstasis intestinal y la preservación de la salud dependen en gran medida de la capacidad de FAI para detoxificar los LPS y suprimir la inflamación metabólica inducida por estos. Sin embargo, es necesario realizar investigaciones a fondo sobre como los hábitos alimenticios pueden modificar la detoxificación de los diferentes compuestos proinflamatorios bacterianos y maximizar la actividad de FAI Abstract in english Abstract One of the main functions of Intestinal Alkaline Phosphatase (FAI) is to detoxify bacterial lipopolysaccharides (LPS) to control intestinal inflammation. Recent data indicate that FAI participates in the detoxification of other bacterial compounds (flagellin and DNA CpG motifs) and many fre [...] e nucleotides (ATP, UDP). FAI is directly involved in the resolution of tissue inflammation mediated by Resolvin E1. The anti-inflammatory action of FAI indirectly improves the intestinal barrier function and affects the diversity of microbiota. Var

Jean-Paul, Lallès; Jaime Parra, Suescún.

2014-06-01

342

iagnostic accuracy study comparing total alkaline phosphatase with intact parathyroid hormone 1-84 for the diagnosis of high turnover renal osteodystrophy in chronic renal failure on hemodialysis  

Directory of Open Access Journals (Sweden)

Full Text Available INTRODUCTION High turnover renal osteodystrophy (HTRO is a highly prevalent complication in patients with chronic kidney disease and mineral bone disease (CKD-MBD, causing pain and significant fracture-associated morbidity and mortality. The diagnostic gold standard test is bone biopsy but there are other, more widely available screening tests such as 1-84 intact parathormone (1-84 iPTH and nonspecific markers such as total alkaline phosphatase (tALP. PURPOSE To determine the diagnostic value (ROC curve, predictive values and likelihood ratios of 1-84 iPTH and tALP for HTRO screening. METHODS A diagnostic accuracy study was performed on a sample of CKD-MDB patients, grouping them according to bone biopsy results and analyzing the results of the diagnostic tests as descriptive variables. RESULTS The study group comprised 188 patients with CKD-MDB, 36 of which had biopsy-confirmed HTRO (19.15%. The average age was 50.2 years in the biopsy group, and 53.4 years in the non-biopsy group (p=0.2385, most were male (63.8% and diabetic (80.5%. The mean time in dialysis was 5.02 years in the biopsy group, and 2.61 years for the non-biopsy group (p<0.001. The mean Kt/V was 1.44 in the biopsy group, and 1.40 in the non-biopsy group (p=0.5354. The mean tALP was 398.02 IU/L in the group with HTRO versus 141.76 IU/L in the group without HTRO (p<0.001. The best cut-off value for tALP was 300-350 IU/L with a near 80% post-test probability, but also with a 15-20% probability for HTRO if the test is negative. The mean 1-84 iPTH was 1248.01 pg/ml in the group with HTRO versus 350.76 pg/ml in the group without HTRO (p<0.001. The 1-84 iPTH cut-off reference value of 300 pg/ml was associated with a post-test probability of 30% for HTRO diagnosis and had a lower overall performance. The best cut-off value for iPTH 1-84 was 600 pg/ml with a post-test probability for HTRO of 70% if positive and less than 5% if the test results are negative. DISCUSSION Both markers show good correlation with bone biopsy findings. tALP elevation detects presence of HTRO in selected patients but does not rule it out. tALP does not perform as well as 1-84 iPTH as a screening test for HTRO.

Andrés Marcelo Rojas González

2014-09-01

343

Altered alkaline phosphatase activity in obese Zucker rats liver respect to lean Zucker and Wistar rats discussed in terms of all putative roles ascribed to the enzyme  

Directory of Open Access Journals (Sweden)

Full Text Available Biliary complications often lead to acute and chronic liver injury after orthotopic liver transplantation (OLT. Bile composition and secretion depend on the integrated action of all the components of the biliary tree, starting from hepatocytes. Fatty livers are often discarded as grafts for OLT, since they are extremely vulnerable to conventional cold storage (CS. However, the insufficiency of donors has stimulated research to improve the usage of such marginal organs as well as grafts. Our group has recently developed a machine perfusion system at subnormothermic temperature (20°C; MP20 that allows a marked improvement in preservation of fatty and even of normal rat livers as compared with CS. We sought to evaluate the response of the biliary tree of fatty liver to MP20, and a suitable marker was essential to this purpose. Alkaline phosphatase (AlkP, EC 3.1.3.1, frequently used as marker of membrane transport in hepatocytes and bile ducts, was our first choice. Since no histochemical data were available on AlkP distribution and activity in fatty liver, we have first settled to investigate AlkP activity in the steatotic liver of fatty Zucker rats (fa/fa, using as controls lean Zucker (fa/+ and normal Wistar rats. The AlkP reaction in Wistar rats was in accordance with the existing data and, in particular, was present in bile canaliculi of hepatocytes in the periportal region and midzone, in the canals of Hering and in small bile ducts but not in large bile ducts. In lean ZR liver the AlkP reaction in Hering canals and small bile ducts was similar to Wistar rat liver but hepatocytes had lower canalicular activity and besides presented moderate basolateral reaction. The difference between lean Zucker and Wistar rats, both phenotypically normal animals, could be related to the fact that lean Zucker rats are genotypically heterozygous for a recessive mutated allele. In fatty liver, the activity in ductules and small bile ducts was unchanged, but most hepatocytes were devoid of AlkP activity with the exception of clusters of macrosteatotic hepatocytes in the mid-zone, where the reaction was intense in basolateral domains and in distorted canaliculi, a typical pattern of cholestasis. The interpretation of these data was hindered by the fact that the physiological role of AlkP is still under debate. In the present study, the various functions proposed for the role of the enzyme in bile canaliculi and in cholangiocytes are reviewed. Independently of the AlkP role, our data suggest that AlkP does not seem to be a reliable marker to study the initial step of bile production during OLT of fatty livers, but may still be used to investigate the behaviour of bile ductules and small bile ducts.

V. Bertone

2011-02-01

344

Estudio de la fosfatasa ácida y alcalina en suelos de la Región Pampeana Norte del área sojera argentina Study of acid and alkaline phosphatase in soils of the Pampean North Region from argentine soybean area  

Directory of Open Access Journals (Sweden)

Full Text Available La transformación de los compuestos de fósforo orgánico (Po a fósforo inorgánico (Pi soluble, es denominada mineralización y es llevada a cabo por un grupo de enzimas conocidas como fosfatasas. En este trabajo, se estudió la actividad fosfatasa de cinco lotes de la región pampeana norte del área sojera argentina, mediante: la evaluación de la actividad fosfatasa del suelo, y el recuento de las comunidades bacterianas y fúngicas con esa actividad, y de esta manera se obtuvo información sobre el potencial de los mismos para movilizar el Po. Se determinó el número de bacterias aeróbicas heterotróficas cultivables (BAHC así como el de hongos cultivables (HC, además del número de productores de fosfatasas ácidas y alcalinas. El número de bacterias con actividad fosfatasa fue en promedio 6,85 10(5 UFC g-1 de suelo para la fosfatasa ácida; mientras que para la fosfatasa alcalina fue en promedio 5,80 10(5 UFC g-1 de suelo. En cambio, el valor medio de hongos con actividad fosfatasa ácida fue 1,78 10³ UFC g-1 de suelo y para la enzima alcalina 1,77 10³ UFC g-1 de suelo. No se encontraron diferencias estadísticamente significativas entre el número de bacterias y el de hongos con fosfatasa ácida o alcalina entre los distintos suelos. Por otro lado, el nivel de actividad de la fosfatasa alcalina osciló entre 5,72 y 15,5 mg p-nitrofenol kg-1 suelo h-1, mientras la ácida varió entre 27,4 y 105 mg p-nitrofenol kg-1 suelo h-1. Se encontraron diferencias significativas en la actividad enzimática entre los cinco suelos, siendo mayor la actividad ácida que la alcalina. Los resultados de este trabajo demostraron que los suelos estudiados presentan una actividad mineralizadora de fuentes de Po que coincide con otros trabajos de suelos cultivados y que el recuento de los organismos productores de fosfatasas complementa la información obtenida a partir de la determinación de la actividad fosfatasa del suelo. Mediante la utilización de ambos métodos, es posible estudiar la fosfatasa ácida y alcalina de un suelo y obtener información sobre el potencial del mismo para movilizar Po.Transformation of organic phosphorus (Po into soluble inorganic phosphorus (Pi is called mineralization and is carried out by phosphatase enzymes. The present research focuses on the study of the phosphatase activity of five soils from the soybean area of the Northern Pampean region, by evaluating the phosphatase activity in soil samples and the number of bacteria and fungi with that activity. Soil samples were collected and the total number and phosphatase activity of cultivated heterotrophic aerobic bacteria (CHAB and cultivated fungi (CF was assessed. No significant differences were observed in the numbers of CHAB and CH between the studied soils. The number of bacteria with acid phosphatase activity was 6.85 10(5 CFU g-1 soil, while alkaline activity was 5.80 10(5 CFU g-1 soil. In contrast, the number of fungi with acid phosphatase activity was 1.78 10³ CFU g-1 soil and with alkaline activity was 1.77 10³ CFU g-1 soil. No significant differences were observed in the number of bacteria and fungi with both enzymes. However, acid activity was higher than alkaline activity in soil samples. Alkaline phosphatase activity ranged from 5.72 to 15.5 mg p- nitrofenol kg-1 soil h-1 while acid activity varied from 27.4 to 10(5 mg p-nitrofenol kg-1 soil h-1. There were significant differences in phosphatase activity between the soybean soils. Our results show that the mineralization activities of Po sources are in agreement with other cultivated soils. On the other hand, the number of bacteria and fungi complements the information on soil phosphatase activity. Clearly, both methods allow the study of alkaline and acid phosphatase activity in soil and give information about the soil potential to mobilize Po.

Leticia Andrea Fernández

2008-07-01

345

Estudio de la fosfatasa ácida y alcalina en suelos de la Región Pampeana Norte del área sojera argentina / Study of acid and alkaline phosphatase in soils of the Pampean North Region from argentine soybean area  

Scientific Electronic Library Online (English)

Full Text Available SciELO Argentina | Language: Spanish Abstract in spanish La transformación de los compuestos de fósforo orgánico (Po) a fósforo inorgánico (Pi) soluble, es denominada mineralización y es llevada a cabo por un grupo de enzimas conocidas como fosfatasas. En este trabajo, se estudió la actividad fosfatasa de cinco lotes de la región pampeana norte del área s [...] ojera argentina, mediante: la evaluación de la actividad fosfatasa del suelo, y el recuento de las comunidades bacterianas y fúngicas con esa actividad, y de esta manera se obtuvo información sobre el potencial de los mismos para movilizar el Po. Se determinó el número de bacterias aeróbicas heterotróficas cultivables (BAHC) así como el de hongos cultivables (HC), además del número de productores de fosfatasas ácidas y alcalinas. El número de bacterias con actividad fosfatasa fue en promedio 6,85 10(5) UFC g-1 de suelo para la fosfatasa ácida; mientras que para la fosfatasa alcalina fue en promedio 5,80 10(5) UFC g-1 de suelo. En cambio, el valor medio de hongos con actividad fosfatasa ácida fue 1,78 10³ UFC g-1 de suelo y para la enzima alcalina 1,77 10³ UFC g-1 de suelo. No se encontraron diferencias estadísticamente significativas entre el número de bacterias y el de hongos con fosfatasa ácida o alcalina entre los distintos suelos. Por otro lado, el nivel de actividad de la fosfatasa alcalina osciló entre 5,72 y 15,5 mg p-nitrofenol kg-1 suelo h-1, mientras la ácida varió entre 27,4 y 105 mg p-nitrofenol kg-1 suelo h-1. Se encontraron diferencias significativas en la actividad enzimática entre los cinco suelos, siendo mayor la actividad ácida que la alcalina. Los resultados de este trabajo demostraron que los suelos estudiados presentan una actividad mineralizadora de fuentes de Po que coincide con otros trabajos de suelos cultivados y que el recuento de los organismos productores de fosfatasas complementa la información obtenida a partir de la determinación de la actividad fosfatasa del suelo. Mediante la utilización de ambos métodos, es posible estudiar la fosfatasa ácida y alcalina de un suelo y obtener información sobre el potencial del mismo para movilizar Po. Abstract in english Transformation of organic phosphorus (Po) into soluble inorganic phosphorus (Pi) is called mineralization and is carried out by phosphatase enzymes. The present research focuses on the study of the phosphatase activity of five soils from the soybean area of the Northern Pampean region, by evaluating [...] the phosphatase activity in soil samples and the number of bacteria and fungi with that activity. Soil samples were collected and the total number and phosphatase activity of cultivated heterotrophic aerobic bacteria (CHAB) and cultivated fungi (CF) was assessed. No significant differences were observed in the numbers of CHAB and CH between the studied soils. The number of bacteria with acid phosphatase activity was 6.85 10(5) CFU g-1 soil, while alkaline activity was 5.80 10(5) CFU g-1 soil. In contrast, the number of fungi with acid phosphatase activity was 1.78 10³ CFU g-1 soil and with alkaline activity was 1.77 10³ CFU g-1 soil. No significant differences were observed in the number of bacteria and fungi with both enzymes. However, acid activity was higher than alkaline activity in soil samples. Alkaline phosphatase activity ranged from 5.72 to 15.5 mg p- nitrofenol kg-1 soil h-1 while acid activity varied from 27.4 to 10(5) mg p-nitrofenol kg-1 soil h-1. There were significant differences in phosphatase activity between the soybean soils. Our results show that the mineralization activities of Po sources are in agreement with other cultivated soils. On the other hand, the number of bacteria and fungi complements the information on soil phosphatase activity. Clearly, both methods allow the study of alkaline and acid phosphatase activity in soil and give information about the soil potential to mobilize Po.

Leticia Andrea, Fernández; Marcelo Antonio, Sagardoy; Marisa Anahí, Gómez.

2008-07-01

346

Effects of Aqueous Stem-Bark Extract of Momordica balsamina Linn on Some Serum Enzymes in Normal and Ethanol Fed Rats  

Directory of Open Access Journals (Sweden)

Full Text Available Aqueous stem-bark extract of Momordica balsamina Linn was administered using stomach tubes to normal and alcohol fed rats to study the effect of the extract on organs and tissues by estimating the level of some serum enzymes. The extract was administered for two weeks at a dose of 0.56 mg/100 g body weight. The parameters studied include some serum enzymes (Prostatic and total acid phosphatase, alkaline phosphatase, alanine aminotransferase (ALAT and aspartate aminotransferase (ASAT, serum glucose, albumin and total protein. Results obtained shows that the stem-bark extract has hypoglycaemic effect in rats. The extract alone was observed to have significant effect on alkaline phosphatase. The level of albumin was insignificantly increased as well as those of ALAT and ASAT. Prostatic and total acid phosphatases were observed to be significantly increased in ethanol fed rats alone also.

M.A. Geidam

2007-01-01

347

Application of Scharer's quantitative method for the determination of residual alkaline phosphatase activity in standard Minas / Aplicação do método modificado de Scharer para a determinação quantitativa da atividade de fosfatase alcalina residual em queijo minas padrão  

Scientific Electronic Library Online (English)

Full Text Available SciELO Brazil | Language: English Abstract in portuguese A pasteurização do leite é um ponto crítico na indústria de laticínios, e falhas nessa etapa comprometem a segurança do produto. O método enzimático de Scharer é tradicionalmente utilizado na verificação da eficiência da pasteurização e baseia-se na pesquisa da atividade de fosfatase alcalina residu [...] al em leite. Embora vários métodos estejam disponíveis para avaliar a eficiência da pasteurização, há um número reduzido de dados publicados baseados na quantificação da atividade da fosfatase alcalina em queijo. Neste estudo, o método modificado de Scharer foi utilizado para determinar os níveis de fosfatase alcalina residual em queijo minas padrão, antes e após 20 dias de maturação. Os queijos foram feitos com leite cru ou com leite pasteurizado com adição de diferentes concentrações de leite cru (0, 0,05%, 0,10%, 0,20% e 0,50%). Nas amostras de queijo fresco, o método apresentou sensibilidade apenas com 0,50% de adição de leite cru ao leite pasteurizado utilizado na fabricação de queijo. Em níveis de adição de até 0,20% de leite cru no leite pasteurizado, as concentrações de fenol se mostraram inferiores a 1?g de fenol/g de produto lácteo, que é o valor preconizado como indicador de pasteurização adequada. Abstract in english Milk pasteurization is a critical issue in the dairy industry, and failures in this process can affect final product safety. Scharer's enzymatic method is still traditionally used to verify pasteurization efficiency compliance, and it is based on screening for residual alkaline phosphatase in milk. [...] Although several methods are used to quantify enzymatic activity to assess milk pasteurization efficiency, there is a small amount of published data regarding the use of these methods to quantify alkaline phosphatase in cheese. In this study, the Scharer's modified method was used to determine the levels of residual alkaline phosphatase in standard minas cheese, before and after 20 days of ripening. The cheeses were made using raw or pasteurized milk with the addition of different concentrations of raw milk (0; 0.05%; 0.10%; 0.20%; and 0.50%). In the fresh cheese samples, the method showed a sensitivity of only 0.50% with the addition of raw milk to the pasteurized milk used to make cheese. In addition, levels of up 0.20% of raw milk in pasteurized milk, the concentrations of phenol was inferior to 1?g phenol/g of dairy product which is the preconized indicator value for adequate pasteurization.

C.F., Soares; L.M., Fonseca; M.O., Leite; M.C.P.P., Oliveira.

1223-12-01

348

SERUM CHEMISTRIES OF COTURNIX JAPONICA GIVEN DIETARY MANGANESE OXIDE (MN3O4)  

Science.gov (United States)

Plasma creatinine and inorganic phosphorus were increased in manganese oxide (Mn3O4)-treated adult male Coturnix quail, but BUN, BUN/creatinine ratio, uric acid, and total calcium were decreased. 2. Serum enzymes (alkaline phosphatase glutamic oxaloacetic transaminase, glutamic p...

349

Prostatic acid phosphatase in serum and semen of dogs / Fosfatasa ácida prostática en suero y semen de perros  

Scientific Electronic Library Online (English)

Full Text Available SciELO Chile | Language: English Abstract in spanish La incidencia de cáncer de próstata ha incrementado el uso de los marcadores celulares para detectar el cáncer en este tejido. Antígenos específicos del tejido o antígenos de diferenciación se encuentran en la superficie de las células normales. Clínicamente, estos antígenos son importantes para el [...] diagnóstico de alteraciones en estos tejidos y para la inmunoterapia. Este estudio trata de evaluar la importancia de la fosfatasa ácida prostática en la próstata canina e investigar su concentración en el suero y en el plasma seminal de perros saludables de diferentes edades. La concentración de fosfatasa ácida prostática en el plasma seminal y en el suero fue evaluada por espectrofotometría, utilizando un kit comercial. Los niveles de la fosfatasa ácida prostática (PAP) no fueron diferentes de acuerdo con la edad y no presentaron correlación con la edad o con las dimensiones de las próstatas verificadas por ecografía. Los valores de concentración de PAP presentaron una gran variación en cada grupo. Sin embargo, son necesarios más estudios para evaluar el papel de la fosfatasa ácida prostática en la próstata canina y su importancia como una prueba de diagnóstico para los trastornos de la próstata. Abstract in english The incidence of prostatic malignancy has increased the use of tissue markers to detect cancer. Tissue specific antigens or differentiation antigens are found on the surface of normal cells. Clinically, these antigens are important to diagnose alterations in the tissues and for immunotherapy. The ob [...] jective of the present study was to evaluate the prostatic acid phosphatase concentration in blood and seminal plasma of intact and healthy dogs at different ages. The evaluation was carried out by spectrophotometer, using a commercial kit. The prostatic acid phosphatase (PAP) levels did not differ according to the age and did not correlate with age or prostatic dimensions verified by ultrasonography. The PAP concentration values varied greatly within each group. However, more studies are necessary to evaluate the role of prostatic acid phosphatase in the canine prostate and its importance as a diagnostic test for prostate disorders.

CRF, Gadelha; WRR, Vicente; APC, Ribeiro; M, Apparicio; GJ, Covizzi; ACN, Campos.

350

Core structure of glycyrrhizan GA, the main polysaccharide from the stolon of Glycyrrhiza glabra var. glandulifera; anti-complementary and alkaline phosphatase-inducing activities of the polysaccharide and its degradation products.  

Science.gov (United States)

The controlled Smith degradation and limited hydrolysis of glycyrrhizan GA, a representative polysaccharide with remarkable phagocytosis-enhancing activity isolated from the stolon of Glycyrrhiza glabra L. var. glandulifera Reg. et Herd. were carried out. Methylation analyses of the primary and the secondary Smith degradation products and of the limited hydrolysis product indicated that the core structural features of glycyrrhizan GA include a backbone chain composed of beta-1,3-linked D-galactose residues. Three-fifths of the galactose units in the backbone carry side chains composed of beta-1,3- and beta-1,6-linked D-galactosyl residues at position 6. Anti-complementary and alkaline phosphatase-inducing activities of the polysaccharide, periodate oxidation-reduction and the controlled Smith degradation products were investigated, and the controlled Smith degradation product showed significant activity. PMID:1446371

Takada, K; Tomoda, M; Shimizu, N

1992-09-01

351

Experimental study on the usefulness of magnetotherapy in bone fractures (tibial osteotomy in the rat). Accumulation of 99 mTc MDP - tests of tensile strength - determination of alkaline phosphatase  

International Nuclear Information System (INIS)

Non-directional magnetic field therapy using a flux density of 60 G and a frequency of 25 Hz was carried out over 12 hours daily in rats in order to ascertain its influence on the healing process following osteotomy of the tibia with internal splint fixation of the fractured bone being carried out as an additional measure. The results thus achieved were compared to those seen in control animals, were no magnetotherapy was carried out, on the basis of scintiscan studies using 99 mTc MDP (degree of density in the callus formed around the fracture zone), the plasma levels of alkaline phosphatase and tests of tensile strength. The follow-up observations of the healing process were additionally based on radiological and histological evaluations of the animals. Beneficial effects of magnetotherapy on the healing process could not be confirmed with any statistical significance. (TRV)

352

Serum Enzymes Levels in HIV/AIDS Patients in Maiduguri, North-Eastern Nigeria  

Directory of Open Access Journals (Sweden)

Full Text Available The serum enzymes (i.e., Aspartate and Alanine amino transaminases and Alkaline phosphatase levels in HIV-1 infected symptomatic/AIDS patients in Maiduguri, North- Eastern Nigeria was studied. The sample population used in this study comprised forty AIDS patients and forty healthy subjects as healthy controls. Five milliliter blood sample was collected by aseptic means after obtaining the consent of the patients and subjects. The two groups were screened for HIV. The 40 AIDS patients were confirmed HIV positive and the forty healthy controls were confirmed HIV negative. The serum enzyme levels in both the AIDS patients and the healthy subjects were assessed by serum assays of Aspartate and alanine amino transaminases and alkaline phosphatase. The results show that there is an elevation in serum enzyme levels in HIV/AIDS patients compared to the healthy subjects. This study is aimed at providing information which will help in managing HIV/AIDS patients.

A.C. Ene

2006-01-01

353

Effect of Refined Petroleum Product (Kerosene Flame and Fumes on Serum Enzyme Characteristics of Broiler Chickens  

Directory of Open Access Journals (Sweden)

Full Text Available Enzyme serology of broiler chickens exposed to refined petroleum product (kerosene flame and fumes at varying distances over a period of 16 hrs daily for 56 days in a poultry house was evaluated. The burning of kerosene was simulated in a designed burner. The measured distances were 4m, 8m and 12m from the flame point. The control birds were located in a separate poultry building without the flame treatment. Proprietary broiler starter and finisher diets were fed ad libitum. Blood samples were taken at the 4th and 8th weeks for enzymological assay from each treatment. The enzymes assayed were Serum Glutamic Oxaloacetic Transaminase (SGOT, Serum Glutamic Pyruvic Transaminase (SGPT and Alkaline Phosphatase (ALP. No significant (P > 0.05 differences were observed in Serum Glutamic Pyruvic Transaminase (SGPT and Alkaline Phosphatase (ALP. Results also showed that there were significant differences (P < 0.05 in Serum Glutamic Oxaloacetic Transaminase (SGOT.

A.O. Amakiri

2008-01-01

354

Diagnosis of testicular carcinoma in situ, (intratubular- and micro-invasive) seminoma and embryonal carcinoma using direct enzymatic alkaline phosphatase reactivity on frozen histological sections.  

Digital Repository Infrastructure Vision for European Research (DRIVER)

Abstract Aims: Testis-sparing surgery might benefit quality of life, but can only be applied with histological examination for seminoma and embryonal carcinoma, and carcinoma in situ (CIS). Diagnosis is based on paraffin-embedded tissue, therefore a delay in further surgery is mostly unavoidable. Methods and Results: A total of 4,093 snap frozen samples and paraffin tissue of 1,500 patients were included. Besides standard H & e staining, the direct enzymatic alkaline phosphatas...

Stoop, Hans; Kirkels, Wim; Dohle, Gert; Gillis, Ad; Den Bakker, Michael; Biermann, Katharina; Oosterhuis, J. Wolter; Looijenga, Leendert

2011-01-01

355

Variation of pH in lysed platelet concentrates influence proliferation and alkaline phosphatase activity in human osteoblast-like cells.  

Digital Repository Infrastructure Vision for European Research (DRIVER)

Activated platelets release a multifaceted blend of growth factors that has stimulatory effects on mesenchymal cells, both in vitro and in vivo, which imply beneficial effects on wound repair and tissue regeneration. Previous studies on fibroblast cultures have revealed that more potent growth factors, with respect to cell proliferation, are released in acidic preparations of lysed platelet concentrates in comparison with neutral and alkaline preparations. The current study was intended to in...

Wahlstro?m, Ola; Linder, Cecilia; Kale?n, Anders; Magnusson, Per

2007-01-01

356

Activation of thiol-dependent antioxidant activity of human serum albumin by alkaline pH is due to the B-like conformational change.  

Science.gov (United States)

Antioxidant activity of human serum albumin (HSA) increased steeply as the reaction mixture was shifted from neutral to alkaline pH. The antioxidant activity was also remarkably increased by Ca(2+) or a cationic detergent (cetyltrimethylammonium chloride). Carboxyl group modification of HSA resulted in about 40-fold increase of the antioxidant activity. The chemical modification study indicated that in addition to functional cysteine(s), cationic amino acid residues such as histidine, arginine and lysine appeared to involve in the antioxidant reaction. HSA also exhibited alkaline-pH dependent peroxidase activity to remove fatty acid hydroperoxide. At neutral pH, only two thiols of Cys-289 and free Cys-34 of HSA were modified by a thiol-specific modification reagent, 5-((((2-iodoacetyl)amino)ethy)amino)naphthalene-1-sulfonic acid (I14), regardless of the presence or absence of dithiothreitol (DTT), and the resultant antioxidant activity was not decreased, suggesting that Cys-289 and Cys-34 did not participate in the antioxidant reaction. At alkaline pH, I14 modified several additional HSA thiols in the presence, but did not in the absence of DTT. The antioxidant activity of the modified HSA was remarkably decreased to as much as 30% of the antioxidant activity given by the unmodified HSA in the absence of DTT. The HPLC pattern for tryptic peptides containing modified cysteine(s) derived from the I14-treated c-HSA (carboxyl group-modified HSA) at pH 7.0 with DTT was very similar to that of the I14-modified HSA at pH 8.0 with DTT. Taken together, these results suggest that activation of thiol-dependent antioxidant activity of HSA at alkaline pH is due to the conformational change favorable for the functional cysteine(s)-mediated catalysis. PMID:10933886

Lee, H; Cha, M K; Kim, I H

2000-08-15

357

A study on serum enzyme levels in various liver diseases  

Digital Repository Infrastructure Vision for European Research (DRIVER)

Patients with chronic liver diseases are asymptomatic or have only vague non-specific symptoms. Effective medical treatments for chronic liver disease (before cirrhosis is established) are becoming increasingly available and since abnormal LFTs may be the only indication of these diseases. Aims: Enzymes study of various liver diseases. Discussion: serum Alkaline phosphatase (ALP), Gamma Glutamyl transferase (Gamma GT), Alanine and Aspartate amino transferases were estimated in viral Hepatiti...

Salma Mahaboob R, Jayarami Reddy U.

2013-01-01

358

The Predictivity of Serum Biochemical Markers in Acute Biliary Pancreatitis  

Digital Repository Infrastructure Vision for European Research (DRIVER)

Background and Aim. There are no accurate methods of differentiating acute biliary pancreatitis. Obstructions of biliary ducts, idiopathic pancreatitis may be related with biliary origin which needs identification for acute treatment. We searched for the predictivity of biochemical markers in early acute biliary pancreatitis. Patients and Methods. Serum levels of AST (Aspartate Transaminase),ALT (Alanine Transaminase), ALP (Alkaline Phosphatase), GGT (Gamma Glutamyl Transferase), total bilir...

Bülent Güngör; Amp Layan, Kas Amp M. Amp A.; Cafer Polat; Deniz ?eren; Kenan Erzurumlu; Zafer Malazgirt

2011-01-01

359

Associations between Serum C-reactive Protein and Serum Zinc, Ferritin, and Copper in Guatemalan School Children  

Digital Repository Infrastructure Vision for European Research (DRIVER)

Inflammation affects trace nutrient concentrations, but research on copper and particularly in children is limited. We assessed associations between serum C-reactive protein (CRP) and zinc, iron, copper, and other biomarkers (alkaline phosphatase, hemoglobin, and albumin), in 634 healthy 6- to 11-year-old Guatemalan schoolchildren. CRP was measured by a standardized, high-sensitive method. For significant associations with CRP, we stratified nutrient concentrations across categories of CRP an...

Bui, Vinh Q.; Stein, Aryeh D.; Digirolamo, Ann M.; Ramakrishnan, Usha; Flores-ayala, Rafael C.; Ramirez-zea, Manuel; Grant, Frederick K.; Villalpando, Salvador; Martorell, Reynaldo

2012-01-01

360

Yu Ping Feng San, an Ancient Chinese Herbal Decoction, Regulates the Expression of Inducible Nitric Oxide Synthase and Cyclooxygenase-2 and the Activity of Intestinal Alkaline Phosphatase in Cultures  

Science.gov (United States)

Yu Ping Feng San (YPFS), a Chinese herbal decoction comprising Astragali Radix (AR; Huangqi), Atractylodis Macrocephalae Rhizoma (AMR; Baizhu), and Saposhnikoviae Radix (SR; Fangfeng), has been used clinically to treat inflammatory bowel diseases (IBD). Previously, we demonstrated a dual role of YPFS in regulating cytokine release in cultured macrophages. In this study, we elucidated the anti-inflammatory effect of YPFS that is mediated through modulating the expression of three key enzymes involved in IBD: inducible nitric oxide synthase (iNOS), cyclooxygenase-2 (COX-2), and intestinal alkaline phosphatase (IALP). In a lipopolysaccharide (LPS)-induced chronic-inflammation model of cultured murine macrophages, YPFS treatment suppressed the activation of iNOS and COX-2 expression in a dose-dependent manner. Conversely, application of YPFS in cultured small intestinal enterocytes markedly induced the expression of IALP in a time-dependent manner, which might strengthen the intestinal detoxification system. A duality of YPFS in modulating the expression of iNOS and COX-2 was determined here. The expression of iNOS and COX-2 in macrophages was induced by YPFS, and this activation was partially blocked by the NF-?B-specific inhibitor BAY 11-7082, indicating a role of NF-?B signaling. These YPFS-induced changes in gene regulation strongly suggest that the anti-inflammatory effects of YPFS are mediated through the regulation of inflammatory enzymes. PMID:24967898

Du, Crystal Y. Q.; Choi, Roy C. Y.; Dong, Tina T. X.; Lau, David T. W.; Tsim, Karl W. K.

2014-01-01

 
 
 
 
361

A Chronoamperometric Screen Printed Carbon Biosensor Based on Alkaline Phosphatase Inhibition for W(IV) Determination in Water, Using 2-Phospho-l-Ascorbic Acid Trisodium Salt as a Substrate.  

Science.gov (United States)

This paper presents a chronoamperometric method to determine tungsten in water using screen-printed carbon electrodes modified with gold nanoparticles and cross linked alkaline phosphatase immobilized in the working electrode. Enzymatic activity over 2-phospho-l-ascorbic acid trisodium salt, used as substrate, was affected by tungsten ions, which resulted in a decrease of chronoamperometric current, when a potential of 200 mV was applied on 10 mM of substrate in a Tris HCl buffer pH 8.00 and 0.36 M of KCl. Calibration curves for the electrochemical method validation, give a reproducibility of 5.2% (n = 3), a repeatability of 9.4% (n = 3) and a detection limit of 0.29 ± 0.01 µM. Enriched tap water, purified laboratory water and bottled drinking water, with a certified tungsten reference solution traceable to NIST, gave a recovery of 97.1%, 99.1% and 99.1% respectively (n = 4 in each case) and a dynamic range from 0.6 to 30 µM. This study was performed by means of a Lineweaver-Burk plot, showing a mixed kinetic inhibition. PMID:25621602

Alvarado-Gámez, Ana Lorena; Alonso-Lomillo, María Asunción; Domínguez-Renedo, Olga; Arcos-Martínez, María Julia

2015-01-01

362

A Chronoamperometric Screen Printed Carbon Biosensor Based on Alkaline Phosphatase Inhibition for W(IV Determination in Water, Using 2-Phospho-l-Ascorbic Acid Trisodium Salt as a Substrate  

Directory of Open Access Journals (Sweden)

Full Text Available This paper presents a chronoamperometric method to determine tungsten in water using screen-printed carbon electrodes modified with gold nanoparticles and cross linked alkaline phosphatase immobilized in the working electrode. Enzymatic activity over 2-phospho-l-ascorbic acid trisodium salt, used as substrate, was affected by tungsten ions, which resulted in a decrease of chronoamperometric current, when a potential of 200 mV was applied on 10 mM of substrate in a Tris HCl buffer pH 8.00 and 0.36 M of KCl. Calibration curves for the electrochemical method validation, give a reproducibility of 5.2% (n = 3, a repeatability of 9.4% (n = 3 and a detection limit of 0.29 ± 0.01 µM. Enriched tap water, purified laboratory water and bottled drinking water, with a certified tungsten reference solution traceable to NIST, gave a recovery of 97.1%, 99.1% and 99.1% respectively (n = 4 in each case and a dynamic range from 0.6 to 30 µM. This study was performed by means of a Lineweaver–Burk plot, showing a mixed kinetic inhibition.

Ana Lorena Alvarado-Gámez

2015-01-01

363

Effects of Aqueous Stem Bark Extract of Cissus populnea on Some Serum Enzymes in Normal and Alloxan Induced Diabetic Rats  

Digital Repository Infrastructure Vision for European Research (DRIVER)

This study was undertaken to assess the effect of 100 mg kg-1 body weight of aqueous stem bark extract of Cissus populnea on serum enzyme levels in normal and alloxan induced diabetic rats. The four weeks experimental protocol involving intragastric administration of the extract revealed a significant increase (P<0.05) in the level of serum alkaline and total acid phosphatase only as a result of diabetes induction. The treatment with Cissus has also revealed a sign...

Geidam, M. A.; Adoga, G. I.; Sanda, F. A.

2004-01-01

364

Introducción del método inmunocitoquímico de la fosfatasa alcalina-antifosfatasa alcalina para la clasificación inmunológica de los Síndromes Linfo y Mieloproliferativos Agudos Introduction of the alkaline phosphatase-alkaline antiphosphatase immunocytochemical method for the immunological classification of the acute lympho-and myeloproliferative syndromes  

Directory of Open Access Journals (Sweden)

Full Text Available Se realizó el inmunofenotipaje celular en 30 pacientes con el diagnóstico de síndromes linfo y mieloproliferativos agudos por el método inmunoenzimático fosfatasa alcalina-antifosfatasa alcalina (APAAp introducido en nuestro laboratorio. Los marcadores estudiados fueron: CD3, CD5, CD7, CD10, CD13, CD15, CD22, CD33, CD34 y CD41 mediante los anticuerpos monoclonales correspondientes, según cada caso. De las leucemias agudas, 16 resultaron ser leucemias linfoides (LLA (53,3 % y 12 mieloides (LMA (40 %. Entre las LLA, el 50 % fue del fenotipo B y del resto, 1 caso del tipo T (LLA-T (3,33 %. Un paciente se diagnosticó como leucemia aguda híbrida (LAH (3,33 % y el otro se clasificó como leucemia aguda indiferenciada (LAI (3,33 %. Se concluye que el APAAP es un método más rápido y tan eficaz como otros métodos enzimáticos para la clasificación inmunológica de los síndromes linfo y mieloproliferativosThe cellular immunophenotyping was carried out in 30 patients with the diagnosis of acute lympho- and myeloproliferative syndromes by the alkaline phosphatase-alkaline antiphosphatase immunoenzimatic method (APAA introduced in our laboratory. The CD3, CD5, CD7, CD10; CDl3, CDl5, CD22, CD33 and CD41 markers were studied by using the corresponding monoclonal antibodies, according to each case. Of the acute leukemias, 16 were lymphoid leukemias (ALL (53.3 % and 12 were myeloid leukemias (AML (40 %. Among the ALL, 50 % were phenotype B and of the rest, 1 case was type T (ALL-T (3.33 %. A patient was diagnosed acute hybrid leukemia (AHL (3.33 % and the other was classified as acute undifferentiated leukemia (AUL (3.33 %. It is concluded that the APAA is faster and as efficient as other enzimatic methods for the immunologic classification of the lympho- and myeloproliferative syndromes

Berta B Socarrás Ferrer

2001-04-01

365

Production of human alkaline phosphatase, a secreted, glycosylated protein, from a baculovirus expression system and the attachment-dependent cell line Trichoplusia ni BTI-Tn 5B1-4 using a split-flow, air-lift bioreactor.  

Science.gov (United States)

A split-flow, air-lift bioreactor for the cultivation of insect cells to produce recombinant protein is described. It can be used advantageously with attached cell systems. This bioreactor incorporates two sections: a rise and a downcomer. Trichoplusia ni BTI-Tn 5B1-4 cells are grown on a support material of glass beads or microcarriers placed in the downcomer. This cell line is more productive than other commonly used insect cell lines, but it has the disadvantage of being difficult to use at large volumes since it is not easily adaptable to suspension culture. Adequate oxygen demand is supplied by sparging without direct exposure of cells to air bubbles. Nutrients are supplied convectively to the attached cells on support material as the fluid flows through the downcomer. The split-flow, air-lift bioreactor appears to be suitable for insect cell culture and is potentially scalable. It can provide a high surface-to-volume ratio and can be operated in batch or continuous mode. A lab-scale prototype bioreactor has been constructed and tested for the production of a secreted, glycosylated recombinant protein (human alkaline phosphatase) or seAP using an Autographa californica multiple nuclear polyhedrosis virus (AcMNPV) vector. With a ratio of riser cross-sectional area to downcomer cross-sectional area of 1, an aspect ratio of 4.4, an air-flow rate of 54 mL/min in the riser, and a bed of 2400 3-min nonporous glass beads, 10.7 micrograms/mL of seAP was produced using an MOI (multiplicity of infection or the ratio of plaque-forming units to cells) of 10.(ABSTRACT TRUNCATED AT 250 WORDS) PMID:7764358

Chung, I S; Taticek, R A; Shuler, M L

1993-01-01

366

Up-regulation of alkaline phosphatase expression in human primary osteoblasts by cocultivation with primary endothelial cells is mediated by p38 mitogen-activated protein kinase-dependent mRNA stabilization.  

Science.gov (United States)

For the regeneration of bone in tissue engineering applications, it is essential to provide cues that support neovascularization. This can be achieved by cell-based therapies using mature endothelial cells (ECs) or endothelial progenitor cells. In this context, ECs were used in various in vivo studies in combination with primary osteoblasts to enhance neovascularization of bone grafts. In a previous study, we have shown that cocultivation of human primary ECs and human primary osteoblasts (hOBs) leads to a cell contact-dependent up-regulation of alkaline phosphatase (ALP) expression in osteoblasts, indicating that cocultivated ECs may support osteogenic differentiation and osteoblastic cell functions. In the present study, we investigated this effect in more detail, revealing a time and cell number dependency of EC-mediated up-regulation of the early osteoblastic marker ALP, whereas osteocalcin, a late marker of osteogenesis, was down-regulated. The effect on ALP expression was bidirectional specific for both cell types. Functional inhibition of gap junctional communication between ECs and hOBs by 18alpha-glycyrrhetinic acid had only a weak suppressive effect on EC-mediated ALP up-regulation. In contrast, inhibition of p38 mitogen-activated protein kinase nearly completely prevented the EC-mediated stimulation of osteoblastic ALP expression. To investigate the molecular mechanism underlying the ALP up-regulation, we examined the effect of EC cocultivation on osteoblastic ALP promoter activity as well as mRNA stability. Cocultivation of ECs with hOBs significantly elevated the half-life of osteoblastic ALP mRNA without affecting its promoter activity. In summary, our data show that EC-mediated up-regulation of osteoblastic ALP expression is cell-type specific and is posttranscriptionally regulated via p38 mitogen-activated protein kinase-dependent mRNA turn-over. PMID:19409035

Hager, Sven; Lampert, Florian M; Orimo, Hideo; Stark, G Björn; Finkenzeller, Günter

2009-11-01

367

The intestinal epithelial cell differentiation marker intestinal alkaline phosphatase (ALPi) is selectively induced by histone deacetylase inhibitors (HDACi) in colon cancer cells in a Kruppel-like factor 5 (KLF5)-dependent manner.  

Science.gov (United States)

The histone deacetylase inhibitor (HDACi) sodium butyrate promotes differentiation of colon cancer cells as evidenced by induced expression and enzyme activity of the differentiation marker intestinal alkaline phosphatase (ALPi). Screening of a panel of 33 colon cancer cell lines identified cell lines sensitive (42%) and resistant (58%) to butyrate induction of ALP activity. This differential sensitivity was similarly evident following treatment with the structurally distinct HDACi, MS-275. Resistant cell lines were significantly enriched for those harboring the CpG island methylator phenotype (p = 0.036, Chi square test), and resistant cell lines harbored methylation of the ALPi promoter, particularly of a CpG site within a critical KLF/Sp regulatory element required for butyrate induction of ALPi promoter activity. However, butyrate induction of an exogenous ALPi promoter-reporter paralleled up-regulation of endogenous ALPi expression across the cell lines, suggesting the presence or absence of a key transcriptional regulator is the major determinant of ALPi induction. Through microarray profiling of sensitive and resistant cell lines, we identified KLF5 to be both basally more highly expressed as well as preferentially induced by butyrate in sensitive cell lines. KLF5 overexpression induced ALPi promoter-reporter activity in resistant cell lines, KLF5 knockdown attenuated butyrate induction of ALPi expression in sensitive lines, and butyrate selectively enhanced KLF5 binding to the ALPi promoter in sensitive cells. These findings demonstrate that butyrate induction of the cell differentiation marker ALPi is mediated through KLF5 and identifies subsets of colon cancer cell lines responsive and refractory to this effect. PMID:25037223

Shin, Joongho; Carr, Azadeh; Corner, Georgia A; Tögel, Lars; Dávaos-Salas, Mercedes; Tran, Hoanh; Chueh, Anderly C; Al-Obaidi, Sheren; Chionh, Fiona; Ahmed, Naseem; Buchanan, Daniel D; Young, Joanne P; Malo, Madhu S; Hodin, Richard A; Arango, Diego; Sieber, Oliver M; Augenlicht, Leonard H; Dhillon, Amardeep S; Weber, Thomas K; Mariadason, John M

2014-09-01

368

[Influence of calcium and phosphate ions on the adsorption rate of alkaline phosphatase and some of serum proteins on the hydroxyapatite].  

Science.gov (United States)

Calcium ions accelerate selective adsorption of AP on HAP and phosphate ions inhibit it. Since the biological function of AP is in the synthesis of phosphate ions from organic phosphates, enzyme activity contributes to its adsorption on mineralized tissues HAP, which can be associated with the peculiarities of the action of the enzyme adsorbed compared with dissolved. Apparently, the selective adsorption of enzymes and signaling molecules in the extracellular matrix to produce the microenvironment necessary for cell differentiation and function of tissue. Sorbate composition determines the type of cell differentiation, which, in turn, determines the properties of the synthesized matrix. Thereby circuiting occurs a feedback loop, to maintain homeostasis of tissues. These results can be used to develop a biomimetic material for medicine as well as the fabrication of multilayer coatings with desired properties by means of controlled speed by selectively depositing colloidal particles (nanoparticles). In the case of anisotropic particles the layers it is possible to obtain particles not only one type, but also with their desired orientation in each layer of particles by creating localized sorbent binding sites. PMID:25536793

2014-01-01

369

Chromogranin A as Serum Marker for Gastroenteropancreatic Neuroendocrine Tumors: A Single Center Experience and Literature Review  

Digital Repository Infrastructure Vision for European Research (DRIVER)

The aim of this study was to assess the clinical sensitivities of the tumor markers chromogranin A (CgA), urinary 5-hydroxyindoleacetic acid (5-HIAA) and alkaline phosphatase (AP) in neuroendocrine tumors (NETs) of the GastroEnteroPancreatic-(GEP-) system depending on tumor primary location and metastatic spread. In a retrospective single-center series, sensitivities were evaluated in serum samples from 110 patients with midgut (n = 62) and pancreatic (n = 48) NETs. CgA levels were analyzed b...

Auernhammer, Christoph J.; Christine Spitzweg; Burkhard Göke; Hoffmann, Johannes N.; Herrmann, Karin A.; Alexander Haug; Michael Vogeser; Axel Kuttner; Michael Lauseker; Svenja Nölting

2012-01-01

370

Serum neuron-specific enolase (S-NSE) and the prognosis in small-cell lung cancer (SCLC): a combined multivariable analysis on data from nine centres.  

Digital Repository Infrastructure Vision for European Research (DRIVER)

The influence of pretreatment serum neuron-specific enolase (S-NSE) in addition to more conventional prognostic factors on survival duration in small-cell lung cancer (SCLC) was investigated in 770 patients from nine centres in six countries. The other variables included stage of disease, performance status (PS), age, sex, serum lactate dehydrogenase (S-LDH), serum alkaline phosphatase (S-AP), and serum carcinoembryonic antigen (S-CEA). Increased values of S-NSE (> 12.5 micrograms-1 l) were o...

1996-01-01

371

Relationship between placental alkaline phosphatase activity and cord blood glucose, albumin and neonatal birth weight at term / Relación entre la actividad de la fosfatasa alcalina placentaria, glucosa y albúmina en sangre del cordón umbilical y el peso en recién nacidos a término  

Scientific Electronic Library Online (English)

Full Text Available SciELO Venezuela | Language: English Abstract in spanish Se ha observado que la actividad de la fosfatasa alcalina placentaria (PAP) aumenta progresivamente a medida que avanza el embarazo, posiblemente debido al incremento de su síntesis a través del tejido placentario. Por lo tanto, la presente investigación estudia la relación entre la actividad de la [...] fosfatasa alcalina placentaria y los índices bioquímicos de la nutrición fetal (glucosa sanguínea de cordón umbilical, albúmina) y crecimiento (peso neonatal). Se recolectaron muestran placentarias y sanguíneas provenientes de 105 partos y preparadas tanto para el ensayo de fosfatasa alcalina placentaria como para estimaciones de glucosa y albúmina utilizando los procedimientos estándar. Se tomaron y registraron los pesos de los recién nacidos a término. El análisis y la correlación de los datos obtenidos muestran una relación significativamente positiva entre el PAP y la glucosa en sangre del cordón umbilical (r² = 0,86, P Abstract in english It has been observed that placental alkaline phosphatase (PAP) activity progressively rises as pregnancy advances, possibly, because of its increasing synthesis by placental tissue. The present investigation therefore, examines the relationship between placental alkaline phosphatase activity and the [...] biochemical indices of foetal nutrition (cord blood glucose, albumin) and growth (neonatal birth weight). Placental and umbilical cord blood samples were collected from one hundred and five deliveries and prepared for both, placental alkaline phosphatase assay, and glucose and albumin estimations using standard procedures. The birth weights of the neonates at term were taken and recorded. Correlation analyses of the data obtained show significant positive relationships between PAP and cord blood glucose (r² = 0.86, p

Innocent, Onyesom; Adefunke Olukemi, Opajobi; Ugochukwu Enyinanya, Uzuegbu; Denis, Oriero; Joseph, Mordi; Prosper Ejiro, Awhin; Enaholo, Timothy.

2009-12-01

372

Atividade da fosfatase alcalina no lavado broncoalveolar de equinos de policiamento montado no Estado do Rio de Janeiro / Alkaline phosphatase activity in bronchoalveolar lavage of police horses in Rio de Janeiro State, Brazil  

Scientific Electronic Library Online (English)

Full Text Available SciELO Brazil | Language: Portuguese Abstract in portuguese A utilidade da determinação das atividades enzimáticas no trato respiratório posterior como ferramenta diagnóstica já foi demonstrada em várias espécies. Nesse contexto, este trabalho teve por objetivo determinar a atividade da Fosfatase Alcalina (FAL) no lavado broncoalveolar (LBA) de equinos da Po [...] lícia Militar do Estado do Rio de Janeiro, comparando animais sadios com portadores assintomáticos de doença inflamatória das vias aéreas (DIVA). Para tal, foram avaliados 28 animais adultos, machos, sem histórico de doença respiratória nos dois meses anteriores ao estudo, com os resultados dos exames físicos e laboratoriais (FAL sanguínea, hematócrito, leucograma, proteína total e fibrinogênio plasmáticos) dentro dos parâmetros fisiológicos. Os equinos foram divididos em dois grupos de acordo com o resultado da citologia broncoalveolar. A determinação da atividade da FAL foi realizada por meio de espectrofotometria a partir de alíquotas do sobrenadante do LBA preservadas em nitrogênio líquido. Para a estimativa do fluido epitelial pulmonar e da atividade da FAL neste, foi realizada a correção da diluição provocada pelo lavado. Os equinos com contagem diferencial de tipos celulares compatível com DIVA apresentaram atividade de FAL no LBA menor, quando comparados aos animais sadios, podendo essa dosagem ser utilizada como complementação do diagnóstico da DIVA. Abstract in english The use of determining the enzymatic activities in the posterior respiratory tract as a diagnostic tool has already been demonstrated in several species. In this context, this paper aims to determine the activity of alkaline phosphatase (ALP) in the bronchoalveolar lavage (BAL) of horses from the Mi [...] litary Police of the State of Rio de Janeiro, comparing healthy animals with asymptomatic carriers of an inflammatory airway disease (IAD). Twenty-eight adult male animals with no history of respiratory diseases in the last two months prior to the study were studied. Physical exam and blood laboratory test results (ALP, hematocrit, leukogram, total protein and plasma fibrinogen) were within physiological parameters. The equines were separated into two groups according to the results of the bronchoalveolar cytology. The determination of ALP was done by spectrophotometry with aliquots of the supernatant of the BAL preserved in liquid nitrogen. To estimate pulmonary epithelial lining fluid and ALP activity, correction of the dilution caused by the lavage was done. The horses with a cell type differential count compatible with IAD presented a lower ALP activity in BAL when compared to healthy animals, therefore this dosage can be used as a complement in the diagnosis of IAD.

Maria Luisa Lorêdo Abreu, Jorge; Vanessa, Viscardi; Katia Moreira, Silva; Juliana Nabuco Pereira, Otaka; Nayro Xavier de, Alencar; Rodolpho de Almeida, Torres Filho; Daniel Augusto Barroso, Lessa.

2014-01-01

373