Sample records for chymosin

  1. Camel and bovine chymosin

    DEFF Research Database (Denmark)

    Langholm Jensen, Jesper; Mølgaard, Anne; Navarro Poulsen, Jens Christian;


    Bovine and camel chymosin are aspartic peptidases that are used industrially in cheese production. They cleave the Phe105-Met106 bond of the milk protein κ-casein, releasing its predominantly negatively charged C-terminus, which leads to the separation of the milk into curds and whey. Despite...... having 85% sequence identity, camel chymosin shows a 70% higher milk-clotting activity than bovine chymosin towards bovine milk. The activities, structures, thermal stabilities and glycosylation patterns of bovine and camel chymosin obtained by fermentation in Aspergillus niger have been examined...... interactions arising from variation in the surface charges and the greater malleability both in domain movements and substrate binding contribute to the better milk-clotting activity of camel chymosin towards bovine milk....

  2. Stimulation of chymosin secretion by simultaneous expression with chymosin-binding llama single-domain antibody fragments in yeast

    NARCIS (Netherlands)

    Harmsen, M.M.; Smits, C.B.; Geus, de B.


    We studied the effect of coexpression of chymosin and chymosin-binding llama single-domain antibody fragments (VHHs) on the secretion of chymosin by Saccharomyces cerevisiae cells. A VHH expression library containing chymosin-specific VHHs was obtained by immunization of a llama and coexpressed with

  3. Influence of pH on retention of camel chymosin in curd

    DEFF Research Database (Denmark)

    Børsting, Mette Winther; Qvist, Karsten B.; Ardö, Ylva Margareta


    Retained coagulant in cheese initiates casein breakdown and influences cheese structure and flavour formation. This study investigated the influence of milk pH on retention of camel chymosin in curd and compared it with bovine chymosin. Milk at five different pH levels was coagulated with same...... coagulation activity. The retention of camel chymosin in curd was rather constant at ∼20% between pH 6.65 and 6.00, while it increased almost linear from 2 to 21% for bovine chymosin. The lower pH dependence for retention of camel chymosin than of bovine chymosin may be explained by a lower negative charge...

  4. Structure and Function of Bovine and Camel Chymosin

    DEFF Research Database (Denmark)

    Jensen, Jesper Langholm

    The central step in cheese making is the separation of milk into curd and whey. This can be done enzymatically by hydrolysis of the Phe105-Met106 bond or nearby bonds in bovine κ-casein, which releases its hydrophilic C-terminal leading to coagulation of the milk. The preferred enzyme...... for this action is bovine chymosin from the calf's stomach, as it has a high activity towards the Phe105-Met106 bond, and a low activity towards other bonds in the milk proteins, as the latter can lead to loss of protein in the whey and release of peptides with a bitter taste. Chymosin was isolated from camel...... this difference through the study of the structures of bovine and camel chymosin, and preparation of catalytically inactive enzymes in complex with substrate. Their milk-clotting activities was determined using the traditional assay on skimmed milk, and a fluorescence resonance energy transfer (FRET) assay...

  5. Camel and bovine chymosin: the relationship between their structures and cheese-making properties

    Energy Technology Data Exchange (ETDEWEB)

    Langholm Jensen, Jesper [University of Copenhagen, (Denmark); Chr. Hansen A/S, Bøge Allé 10-12, DK-2970 Hørsholm (Denmark); Mølgaard, Anne; Navarro Poulsen, Jens-Christian [University of Copenhagen, (Denmark); Harboe, Marianne Kirsten [Chr. Hansen A/S, Bøge Allé 10-12, DK-2970 Hørsholm (Denmark); Simonsen, Jens Bæk [University of Copenhagen, (Denmark); Lorentzen, Andrea Maria; Hjernø, Karin [University of Southern Denmark, (Denmark); Brink, Johannes M. van den; Qvist, Karsten Bruun [Chr. Hansen A/S, Bøge Allé 10-12, DK-2970 Hørsholm (Denmark); Larsen, Sine, E-mail: [University of Copenhagen, (Denmark)


    Analysis of the crystal structures of the two milk-clotting enzymes bovine and camel chymosin has revealed that the better milk-clotting activity towards bovine milk of camel chymosin compared with bovine chymosin is related to variations in their surface charges and their substrate-binding clefts. Bovine and camel chymosin are aspartic peptidases that are used industrially in cheese production. They cleave the Phe105-Met106 bond of the milk protein κ-casein, releasing its predominantly negatively charged C-terminus, which leads to the separation of the milk into curds and whey. Despite having 85% sequence identity, camel chymosin shows a 70% higher milk-clotting activity than bovine chymosin towards bovine milk. The activities, structures, thermal stabilities and glycosylation patterns of bovine and camel chymosin obtained by fermentation in Aspergillus niger have been examined. Different variants of the enzymes were isolated by hydrophobic interaction chromatography and showed variations in their glycosylation, N-terminal sequences and activities. Glycosylation at Asn291 and the loss of the first three residues of camel chymosin significantly decreased its activity. Thermal differential scanning calorimetry revealed a slightly higher thermal stability of camel chymosin compared with bovine chymosin. The crystal structure of a doubly glycosylated variant of camel chymosin was determined at a resolution of 1.6 Å and the crystal structure of unglycosylated bovine chymosin was redetermined at a slightly higher resolution (1.8 Å) than previously determined structures. Camel and bovine chymosin share the same overall fold, except for the antiparallel central β-sheet that connects the N-terminal and C-terminal domains. In bovine chymosin the N-terminus forms one of the strands which is lacking in camel chymosin. This difference leads to an increase in the flexibility of the relative orientation of the two domains in the camel enzyme. Variations in the amino acids

  6. Isolation of milk-clotting enzyme from transgenic sheep milk and its comparison with calf chymosin. (United States)

    Mezina, M N; Lavrenova, G I; Prokof'ev, M I; Starovoitova, V V; Ermolaev, V I; Chernyh, V Y; Balandina, G N; Demidovich, S S


    Technology for preparation of chymosin from milk of transgenic sheep has been elaborated. Purification of the preparation by ion-exchange chromatography on aminosilochrom and biospecific chromatography on bacitracin-Sepharose yielded homogeneous active enzyme. Hydrolysis of protein substrates (hemoglobin, BSA, and sodium caseinate) by the transgenic sheep chymosin and stability of the enzyme at various values of pH were studied. Judging by the amino acid composition, the N-terminal sequence involving six amino acid residues, molecular mass, stability at various pH values, and the catalytic activity against the protein substrates, the transgenic sheep chymosin is identical to calf chymosin.

  7. [Occupational allergies against pepsin, chymosin and microbial rennet]. (United States)

    van Kampen, V; Lessmann, H; Brüning, T; Merget, R


    Rennet is a mixture of the proteolytic enzymes pepsin and chymosin (rennin), which is usually obtained from the fourth stomach of young ruminants. While pepsin is also used in the pharmaceutical industry, both enzymes (pepsin and chymosin) are used for the coagulation of milk protein in the manufacture of cheese. Additionally, microbial rennet, which is naturally produced by certain microorganisms, has been used as a substitute for natural rennet in the cheese production for decades. Exposure to enzyme dusts has long been known to cause occupational immediate hypersensitivities. The present paper reviews the results of an evaluation of the literature data concerning occupational airway sensitisation due to natural and microbial rennet. Cases of specific airway sensitisation caused by rennet could be shown clearly by several studies. Positive skin prick and challenge tests as well as specific IgE antibodies have been described, thus suggesting an immunological mechanism.

  8. 21 CFR 184.1685 - Rennet (animal-derived) and chymosin preparation (fermentation-derived). (United States)


    ... 21 Food and Drugs 3 2010-04-01 2009-04-01 true Rennet (animal-derived) and chymosin preparation (fermentation-derived). 184.1685 Section 184.1685 Food and Drugs FOOD AND DRUG ADMINISTRATION, DEPARTMENT OF... (animal-derived) and chymosin preparation (fermentation-derived). (a)(1) Rennet and bovine rennet...

  9. Bovine Chymosin: A Computational Study of Recognition and Binding of Bovine κ-Casein

    DEFF Research Database (Denmark)

    Palmer, David S.; Christensen, Anders Uhrenholt; Sørensen, Jesper


    Bovine chymosin is an aspartic protease that selectively cleaves the milk protein κ-casein. The enzyme is widely used to promote milk clotting in cheese manufacturing. We have developed models of residues 97-112 of bovine κ-casein complexed with bovine chymosin, using ligand docking, conformational...

  10. Effects of Adding Chymosin to Milk on Calcium Homeostasis

    DEFF Research Database (Denmark)

    Møller, Ulla Kristine; Jensen, Lars Thorbjørn; Mosekilde, Leif


    Calcium intake and absorption is important for bone health. In a randomized double-blind cross-over trial, we investigated effects of adding chymosin to milk on the intestinal calcium absorption as measured by renal calcium excretion and indices of calcium homeostasis. The primary outcome...... of the study was 24-h renal calcium excretion that is considered a proxy measure of the amount of calcium absorbed from the intestine. We studied 125 healthy men and women, aged 34 (25-45) years on two separate days. On each day, a light breakfast was served together with 500 ml of semi-skimmed milk to which...

  11. Development of a Radioimmunoassay for Bovine Chymosin B

    Directory of Open Access Journals (Sweden)

    Borceux, JP.


    Full Text Available The present study was conducted to develop and validate a specific radioimmunoassay system for measurement of bovine chymosin B (bChyB concentrations in plasma samples. Bovine ChyB was used for immunization of rabbits and as standard and tracer. Chymosin B concentrations were measured in plasma samples from two groups of calves (Group 1: calves sampled from birth to 24 hours; Group 2: calves sampled from Day 1 to 21 after birth and from one cow during the peri-partum period. Detection limit of the assay was 9.0 ng/ml. Recovery was higher than 89.3%. Repeatability and reproducibility ranged from 1.52% to 5.23% and from 1.52% to 12.57% respectively. No cross-reaction was found with pepsinogen A from bovine, porcine or human origins. In Group 1, bChyB concentrations increased from 47.3 ± 45.1 ng/ml (5 min after birth to 325.5 ± 161.2 ng/ml (12 hours after birth, then no significant change was observed till 24 hours after birth (293.0 ± 161.5 ng/ml. In Group 2, concentrations decreased from Day 1 (455.3 ± 191.1 ng/ml to Day 21 (117.9 ± 85.1 ng/ml. In adult cow, mean concentration was 136.0 ± 32.3 ng/ml. In conclusion, bChyB is able to cross the stomach basal membrane and to reach the blood circulation at detectable levels in both young calves and adult cows.

  12. Factors influencing chymosin-induced gelation of milk from individual dairy cows

    DEFF Research Database (Denmark)

    Gustavsson, F.; Glantz, M; Buitenhuis, Albert Johannes


    Optimisation of cheese yield is crucial for cheese production; a previous study showed large variations in chymosin-induced coagulation in milk from the second most common Swedish dairy breed, Swedish Red. In the present study, the effect of gross composition, protein composition, total and ionic...... was, theoretically, four times larger than the amount of calcium that is normally added in cheese processing......Optimisation of cheese yield is crucial for cheese production; a previous study showed large variations in chymosin-induced coagulation in milk from the second most common Swedish dairy breed, Swedish Red. In the present study, the effect of gross composition, protein composition, total and ionic...

  13. Development and evaluation of a monoclonal antibody-based inhibition ELISA for the quantification of chymosin in solution. (United States)

    Rolet-Répécaud, O; Arnould, C; Dupont, D; Gavoye, S; Beuvier, E; Achilleos, C


    Chymosin is the major enzyme of natural rennet, traditionally used in cheese making for its high milk-clotting activity. For technical reasons, an accurate characterization of rennet should include its total clotting activity and also its enzymatic composition. Monoclonal antibodies specific to chymosin were obtained from mice immunized with purified bovine chymosin, and an inhibition enzyme-linked immunosorbent assay (ELISA) was developed for the quantification of chymosin in solution. No cross-reactivity was observed with other milk-clotting enzymes commonly used in cheese making. The limit of detection and limit of quantification were 125 and 400 ng/mL, respectively. The values of precision within and among runs were 7.23 and 7.39%, respectively, and satisfying recovery, from 92 to 119%, was found for spiked samples. The inhibition ELISA was successfully applied to commercial rennets, and the results were consistent with those obtained using the standard chromatographic method (IDF 110: A, 1987).

  14. Dependence of chymosin and pepsin partition coefficient with phase volume and polymer pausidispersity in polyethyleneglycol-phosphate aqueous two-phase system. (United States)

    Spelzini, Darío; Picó, Guillemo; Farruggia, Beatriz


    The influence of the phase volume ratio and polymer pausidispersity on chymosin and pepsin partition in polyethylenglycol-phosphate aqueous two-phase systems was studied. Both proteins showed a high affinity for the polyethylenglycol rich phase with a partition coefficient from 20 to 100 for chymosin and from 20 to 180 for pepsin, when the polyethyleneglycol molecular mass in the system varied between 1450 and 8000. The partition coefficient of chymosin was not affected by the volume phase ratio, while the pepsin coefficient showed a significant decrease in its partition coefficient with the increase in the top/bottom phase volume ratio.

  15. Recombinant chymosin used for exact and complete removal of a prochymosin derived fusion tag releasing intact native target protein

    DEFF Research Database (Denmark)

    Justesen, Sune; Lamberth, Kasper; Nielsen, Lise-Lotte B;


    characteristics for the exact removal of fusion tags. It is readily available in highly purified recombinant versions approved by the FDA for preparation of food for human consumption. We suggest that one should consider extending the use of chymosin to the preparation of pharmaceutical proteins....

  16. Cloning, expression and optimized production in a bioreactor of bovine chymosin B in Pichia (Komagataella) pastoris under AOX1 promoter. (United States)

    Noseda, Diego Gabriel; Recúpero, Matías Nicolás; Blasco, Martín; Ortiz, Gastón Ezequiel; Galvagno, Miguel Angel


    The codon sequence optimized bovine prochymosin B gene was cloned under the control of the alcohol oxidase 1 promoter (AOX1) in the vector pPIC9K and integrated into the genome of the methylotrophic yeast Pichia (Komagataella) pastoris (P. pastoris) strain GS115. A transformant clone that showed resistance to over 4 mg G418/ml and displayed the highest milk-clotting activity was selected. Cell growth and recombinant bovine chymosin production were optimized in flask cultures during methanol induction phase achieving the highest coagulant activity with low pH values, a temperature of 25°C and with the addition of sorbitol and ascorbic acid at the beginning of this period. The scaling up of the fermentation process to lab-scale stirred bioreactor using optimized conditions, allowed to reach 240 g DCW/L of biomass level and 96 IMCU/ml of milk-clotting activity. The enzyme activity corresponded to 53 mg/L of recombinant bovine chymosin production after 120 h of methanol induction. Western blot analysis of the culture supernatant showed that recombinant chymosin did not suffer degradation during the protein production phase. By a procedure that included high performance gel filtration chromatography and 3 kDa fast ultrafiltration, the recombinant bovine chymosin was purified and concentrated from fermentation cultures, generating a specific activity of 800 IMCU/Total Abs(280 nm) and a total activity recovery of 56%. This study indicated that P. pastoris is a suitable expression system for bioreactor based fed-batch fermentation process for the efficient production of recombinant bovine chymosin under methanol-inducible AOX1 promoter.

  17. Solvent Binding Analysis and Computational Alanine Scanning of the Bovine Chymosin-Bovine κ-Casein Complex Using Molecular Integral Equation Theory. (United States)

    Palmer, David S; Sørensen, Jesper; Schiøtt, Birgit; Fedorov, Maxim V


    We demonstrate that the relative binding thermodynamics of single-point mutants of a model protein-peptide complex (the bovine chymosin-bovine κ-casein complex) can be calculated accurately and efficiently using molecular integral equation theory. The results are shown to be in good overall agreement with those obtained using implicit continuum solvation models. Unlike the implicit continuum models, however, molecular integral equation theory provides useful information about the distribution of solvent density. We find that experimentally observed water-binding sites on the surface of bovine chymosin can be identified quickly and accurately from the density distribution functions computed by molecular integral equation theory. The bovine chymosin-bovine κ-casein complex is of industrial interest because bovine chymosin is widely used to cleave bovine κ-casein and to initiate milk clotting in the manufacturing of processed dairy products. The results are interpreted in light of the recent discovery that camel chymosin is a more efficient clotting agent than bovine chymosin for bovine milk.

  18. Proteolytic Activity in Reduced-Fat Cheddar Cheese Made with Lactic Acid Bacteria and Camel Chymosin

    DEFF Research Database (Denmark)

    Børsting, Mette Winther

    be the need of an extended ripening period to reach a similar cheese structure as in cheeses produced with BC. The aim of this project was to compensate for the lower proteolytic activity in cheese produced with CC compared to BC. Selection of dairy lactic acid bacteria (LAB) for cheese production with high...... for their ability to influence proteolysis and structure during cheese ripening. In an attempt to improve the screening methods and contribute to the development of a new classification system of Latcococcus lactic strains, the peptide profile formed by selected strains after growth in milk was analyzed...... mediated an increase in the total amount of amino acids as well as a shorter structure. A model system, used to study the retention of chymosin in a curd, showed that the retention of CC was less dependent on pH compared to BC, and the retention of CC was higher than BC in the pH interval 6...

  19. Chymosin-induced hydrolysis of caseins: Influence of degree of phosphorylation of alpha-s1-casein and genetic variants of beta-casein

    NARCIS (Netherlands)

    Bijl, E.; Valenberg, van H.J.F.; Sikkes, S.; Jumelet, S.; Sala, G.; Olieman, K.; Hooijdonk, van A.C.M.; Huppertz, T.


    The objective of this study was to investigate the impact of natural variations in aS1-casein and b-casein composition of milk on chymosin-induced hydrolysis of these caseins in milk gels and in sodium caseinate solutions. At 50% casein degradation, 15% more of aS1-casein with eight phosphate groups

  20. Use of grape pomaces to produce biomass of aKomagataella pastoris strain expressing a bovine chymosin activity. (United States)

    Kingston, Diego; Novelli, Guido F; Cerrutti, Patricia; Recupero, Matias N; Blasco, Martin; Galvagno, Miguel A


    The use of agroindustrial wastes not only decreases bioprocesses and disposal costs but also contributes to the upgrading of the residues. An active recombinant methanol-inducible bovine chymosin has been expressed in our laboratory in the yeastKomagataella pastoris, and grape pomace extracts (GRE) were proposed as a convenient C-energy source for the biomass production of the genetically engineered strain. Carbon and nitrogen sources, growth factors, and initial pH conditions were selected by classical methodology; thereafter, growth conditions optimization was performed using statistical designed experiments (DoEs). In the presence of (in g·L(-1)) 67.0 monosaccharides (glucose and fructose) from GRE, 5.0 (NH4)2SO4, and 10.0 sugar cane molasses (CMz), a yield of 20.0 g·L(-1) cell dry weight (CDW) was obtained aerobically after 60 h incubation at 28°C and pH 4.0. Applying a fed-batch strategy with methanol:sorbitol as the enzyme inducers, a chymosin production of 8.53 International Milk Clotting Units (IMCU) per mg protein was obtained in the supernatant. The results presented show that through a statistical design, a simple, cheap, and easy to prepare culture medium could be developed using two agroindustrial derivatives (GRE and CMz) to obtain a higher value added product.

  1. Rearch advancement of the strain breeding which can produce chymosin in China%我国凝乳酶产生菌育种的研究进展

    Institute of Scientific and Technical Information of China (English)

    张超垒; 王成忠; 张志国; 张玲梅


    In terms of natural screening,mutation breeding and the breeding of gene engineering, this paper reviewed the rearch advancement of the strain breeding which can produce chymosin in China.%综述了目前我国在凝乳酶产生菌在自然选育、诱变育种和基因工程育种这3方面的研究进展情况.

  2. 重组毕赤酵母产凝乳酶发酵条件研究%Fermentation conditions of chymosin by recombinant Pichia pastoris

    Institute of Scientific and Technical Information of China (English)

    张健; 张莉; 李玉秋; 杨贞耐


    The optimal conditions for producing chymosin in 5L fermenter by recombinant Pichia pastoris Gsl 15/pPICZaA-Prochy were determined in order to offer technical bases for pilot scale production. After induction for 84h, the highest enzyme activity (152SU/ml) was achieved with batch culture, in which, lOg/L sorbitol was added at the beginning of induction phase, methanol concentration was kept at 0.1%, 10g/l peptone was added every 12h, pH value was kept at 3.0 and the concentration of oleic acid was 0.2%. The purification of chymosin was analyzed by sodium dodecyl sul-fete benzene agarose gel electrophoresis (SDS-PAGE). Under the concentration of salt ions at 2mol/L and flow rate 2.5mL/min, the recovery rate of chymosin was 82.33% and the enzyme sample was concentrated by 20 folds.%以重组毕赤酵母GS115/pPICZaA-Prochy为凝乳酶生产菌种,研究其在5L发酵罐水平的最佳产酶条件,为中试生产提供依据.采用批量发酵,甲醇浓度0.1%,诱导阶段每12h添加10g/L蛋白胨,发酵液pH值为3.0,油酸浓度0.2%,诱导初期批量加入10g/L山梨醇共基培养,诱导84h可获得152SU/mL的酶活力.采用十二烷基磺酸钠-苯琼脂糖凝胶电泳分析,在发酵液(去菌体后)样品盐离子浓度2mol/L,流速2.5mL/min,酶回收率达到82%,浓缩倍数20倍.

  3. Rapid quantification of casein in skim milk using Fourier transform infrared spectroscopy, enzymatic perturbation, and multiway partial least squares: Monitoring chymosin at work

    DEFF Research Database (Denmark)

    Baum, Andreas; Hansen, P. W.; Nørgaard, Lars


    least squares regression. Because we observed casein-specific kinetic changes, the retrieved models were independent of the chemical background matrix and were therefore robust against possible covariance effects. We tested the robustness of the models by spiking the milk solutions with whey, calcium......In this study, we introduce enzymatic perturbation combined with Fourier transform infrared (FTIR) spectroscopy as a concept for quantifying casein in subcritical heated skim milk using chemometric multiway analysis. Chymosin is a protease that cleaves specifically caseins. As a result...... of hydrolysis, all casein proteins clot to form a creamy precipitate, and whey proteins remain in the supernatant. We monitored the cheese-clotting reaction in real time using FTIR and analyzed the resulting evolution profiles to establish calibration models using parallel factor analysis and multiway partial...

  4. 米黑毛霉凝乳酶制备切达干酪的工艺参数优化%Optimization of Preparation Process for Cheddarc Cheese using Chymosin from Mucor miehei

    Institute of Scientific and Technical Information of China (English)

    马江; 曹磊; 张忠明; 关明玲; 刘昕; 师希雄; 姚拓; 张卫兵


    为确定米黑毛霉凝乳酶制作切达干酪的最佳工艺条件,以感官评分和出品率为响应值,在单因素试验的基础上,采用响应面法对主要工艺参数进行了优化.试验得到的米黑毛霉凝乳酶制作切达干酪的最佳工艺参数为:酶添加量为2900.38SU/L、凝乳pH为6.2、CaCl2添加量为0.04%、凝乳温度为34℃、发酵剂添加量为0.14%;在此条件下,干酪感官评分为(95.2±0.34),试验结果与预测值接近,证明模型拟合程度较好.%In order to determine the optimal processing parameters for hard cheese using chymosin from Mucor miehe, the response surface methodology (RSM) was used to optimize the main technological parameters, using sensory evaluation and yield as response value. The optimal processing parameters was obtained as follows:the addi-tion of chymosin was 2900.38SU/L, the curd pH was 6.2, the addition of CaCl2 was 0.04%, the curd temperature was 34℃,the addition of starter culture was 0.14%. Under these conditions, the sensory score of cheese was (95.2±0.34). The model turned out to be with good fitting degree.

  5. 7~35日龄羔羊胃肠胃蛋白酶、凝乳酶、乳糖酶活性与其 mRNA 相对表达量和胃肠发育的关系研究%A Study on Relationships between the Activities of Repsin,Chymosin, Lactase and Their mRNA Relative Expressional Volumes and Development of the Stomach and Intestines of 7-35 Day Old Lambs

    Institute of Scientific and Technical Information of China (English)

    牛越峰; 雒秋江; 陈勇; 李凤鸣; 张慧玲


    powder substitute (MPS)(n =28)or fish meal substitute (FMS)(n =28)respectively,and from 21 day old,8 of lamb fed with MPS or FMS were turned to be fed with starter ration respectively. Each group of lamb at 7,14,21,28 and 35 day old was sacrificed to take the abomasum and duodenum tis-sue,measuring the activities of the pepsin,chymosin and lactase and the corresponding relative expression of mRNA,for studying the development of 3 digestive enzymes of lamb fed with MPS,FMS and starter ra-tion respectively.The result showed that in abomasum or duodenum tissue of newborn lamb the activities of the chymosin and lactase were decreased with the age of lamb by 1.3 U/g (r 2 = 0.92,P 0.05)respectively,but the pepsin activity was weakly related with the age.The relationships be-tween the activities of the pepsin,chymosin and lactase and thier corresponding relative expression of mR-NA was poor,the mRNA expressions of the pepsin and chymosin were fluctuated by high-low-high with the age,however the activity of lactose was not significantly changed with the age of lamb.When FMS was fed,the activities of pepsin,and chymosinwere decreased,the weight of abomasum was (36.0 ±3.0)g at 7 day old of lamb,and was (38.3±5.4)g at 35 day old,suggesting a growth retardation.Turning to starter ration at 21 days of lamb fed priviously with MPS did not significantly affect the activities of the pepsin, chymosin and lactase.But the abomasum and duodenum and daily weight gains of the lambs reduced.Fur-thermore,the activities of pepsin,chymosin and lactase in the milk substitutes,which were fed priviously to the lambs,tended to declining which affected obviously abomasum and duodenum and daily weight gains of the lambs.It is concluded that the pepsin activity is weakly related with the age of lamb,but the activities of chymosin and lactase were decreased with the age;the milk substitutes may affect the latter digestion and growth of lamb;the digestive obstruction on FMS of lamb may mainly be because

  6. Cheese from Ultrafiltered Milk : whey proteins and chymosin activity

    NARCIS (Netherlands)

    Buijsse, C.


    The manufacture of (semi-)hard cheese from ultrafiltered milk (UF-cheese) enables the partial incorporation of whey proteins in the cheese, thereby increasing its yield. The transfer of whey proteins in curd from (UF-)milk was studied in relation to the degree of ultrafiltration of the milk and the

  7. The adsorption of lysozyme and chymosin onto emulsion droplets and their association with caseins

    NARCIS (Netherlands)

    Roos, de A.


    The proteolytic action of proteases present in cheese plays a major role in the ripening of cheese. These proteases originate from the rennet, the starter cultures and from the milk itself. The proteolysis in cheese results in the degradation of the casein proteins into smaller peptides and free ami

  8. 凝乳酶的研究进展%Rearch advancement of Chymosin

    Institute of Scientific and Technical Information of China (English)

    朱仁俊; 石振兴; 甘伯中; 胡永金



  9. 凝乳酶水解酪蛋白生产酪蛋白糖巨肽工艺条件的研究%Study on technology conditions of chymosin hydrolyzing casein to produce casein glycomacropeptide

    Institute of Scientific and Technical Information of China (English)

    张秀媛; 袁永俊


    酪蛋白糖巨肽(casein glycomacropeptide,简称CGMP)是K-酪蛋白经酶解后生成的一类舍有糖链的多肽,具有许多功能特性.本文研究了凝乳酶水解酪蛋白生产CGMP的工艺条件,通过正交实验得到最佳工艺条件是底物浓度10mg/mL、酶的添加量为0.6%、酶解时间为2.5h.唾液酸回收率为80.7%,蛋白质回收率为3.03%,糖基化程度(唾液酸/蛋白质)为77.6μg/mg.

  10. Impact of selected coagulants and starters on primary proteolysis and amino acid release related to bitterness and structure of reduced-fat Cheddar cheese

    DEFF Research Database (Denmark)

    Børsting, Mette Winther; Qvist, Karsten Bruun; Rasmussen, Morten;


    Despite substantial research, it is still difficult to produce high quality reduced-fat Cheddar cheese. The objective of this study was to investigate how two coagulants, bovine chymosin (BC) and camel chymosin (CC) having different proteolytic activities and two starter cultures, an O-culture (O......) and a thermophilic strain of Lactobacillus plus O-culture (OLb) having different abilities to release amino acids, contribute to the structure and flavour development in reduced-fat Cheddar cheese. Cheeses manufactured using the four combinations of coagulants and cultures were analysed during a 28-week ripening...... cheeses with BC had a higher amount of peptides released by chymosin, e. g. the bitter peptide beta-casein (f193-209) or by starter proteases from the chymosin-produced peptide alpha(s1)-CN (f1-23). BC cheeses were also judged to be softer by the sensory panel. Cheeses containing the OLb-culture had...

  11. Characterization of enzymatically induced aggregation of casein micelles in natural concentration by in situ static light scattering and ultra low shear viscosimetry

    DEFF Research Database (Denmark)

    Lehner, D.; Worning, Peder; G, Fritz


    The aggregation of casein micelles in undiluted skim milk after the addition of chymosin was studied bystatic light scattering and ultra low shear viscometry. The static light scattering measurements were madewith two different sample thicknesses, 72 and 16 mum. The scattering data were analyzed...

  12. Research progress of the calf rennet substitutes for cheese-making%干酪用牛凝乳酶替代品的研究进展

    Institute of Scientific and Technical Information of China (English)

    普燕; 张富春


    牛凝乳酶具有高凝乳活性和低非特异蛋白水解活性而广泛用于干酪制备.随着干酪市场的逐年增长,牛凝乳酶出现供不应求,其替代品应运而生,包括其他动物凝乳酶、植物凝乳酶、微生物凝乳酶、重组凝乳酶.在这些替代品中,重组牛凝乳酶显示出与天然牛凝乳酶相似的酶学特性,并且纯度更高,制作的干酪品质更好.目前已经有重组牛凝乳酶上市出售,成功替代了天然牛凝乳酶,但关于提高重组牛凝乳酶表达量与催化活性的研究远未止步.文中主要综述了凝乳酶结构、牛凝乳酶的替代品和重组凝乳酶的研究进展,详细综述了一些提高重组牛凝乳酶的表达方法,为外源蛋白在宿主中的高水平表达提供了有效参考.%Calf rennet is widely used in the process of cheese-making due to its high milk-clotting activity and low nonspecific proteolytic activity.As the cheese market increases in recent years,calf rennet supply falls short of demand and its substitutes are studied extensively,such as animal rennet,plant coagulant,microbial coagulant and recombinant chymosin.Among these substitutes,the recombinant bovine chymosin shows high similar enzymic properties to the natural calf rennet,and possesses high purity,thus the quality of the cheese made with it is better.Now the recombinant bovine chymosins have been listed for sale and the recombinant bovine chymosin have replaced the natural calf rennet successfully.But the studies on the improvement of expression quantity and catalytic activity of recombinant bovine chymosin never stop.In this paper,the research progress on the structure of chymosin,the calf rennet substitutes and the recombinant chymosin,and particularly the methods used to improve the expression quantity of the recombinant bovine chymosin is reviewed.These methods can offer certain reference for expression of foreign proteins at a high level in hosts.

  13. Effects of gelation temperature on Mozzarella-type curd made from buffalo and cows' milk. 1: rheology and microstructure. (United States)

    Hussain, Imtiaz; Grandison, Alistair S; Bell, Alan E


    The rheology and microstructure of Mozzarella-type curds made from buffalo and cows' milk were measured at gelation temperatures of 28, 34 and 39 °C after chymosin addition. The maximum curd strength (G') was obtained at a gelation temperature of 34 °C in both types of bovine milk. The viscoelasticity (tan δ) of both curds was increased with increasing gelation temperature. The rennet coagulation time was reduced with increase of gelation temperature in both types of milk. Frequency sweep data (0.1-10 Hz was recorded 90 min after chymosin addition, and both milk samples showed characteristics of weak viscoelastic gel systems. When both milk samples were subjected to shear stress to break the curd system at constant shear rate, 95 min after chymosin addition, the maximum yield stress was obtained at the gelation temperatures of 34 °C and 28 °C in buffalo and cows' curd respectively. The cryo-SEM and CLSM techniques were used to observe the microstructure of Mozzarella-type curd. The porosity was measured using image J software. The cryo-SEM and CLSM micrographs showed that minimum porosity was observed at the gelation temperature of 34 °C in both types of milk. Buffalo curd showed minimum porosity at similar gelation temperature when compared to cows' curd. This may be due to higher protein concentration in buffalo milk.

  14. Caseinomacropeptide self-association is dependent on whether the peptide is free or restricted in kappa-casein

    DEFF Research Database (Denmark)

    Mikkelsen, T.L.; Frokiaer, H.; Topp, C.


    dodecyl sulfate-PAGE, N-terminal sequencing, and mass spectrometry. The hydrophobicity of CMP was studied by means of 1-anilino-naphthalene-8-sulfonic acid binding experiments. Four CMP products prepared by different methods were studied: CMP produced by enzymatic (chymosin or pepsin) hydrolysis of kappa......-casein (CN), and 2 commercial CMP products. Both commercial products and CMP-resulting from chymosin-hydrolysis of kappa-CN (at pH 6.6) had elution volumes with a MW corresponding to 35 kDA at pH 8.0 and 3.4. Caseinomacropeptide prepared from pepsin-hydrolysis of kappa-CN (at pH 2.5) eluted as multiple peaks...... with apparent MW of 35, 18, and 9 kDa, again independently of pH. Hydrolysis of kappa-CN with chymosin or pepsin at different pH values (pH 2.5, 3.4, and 6.6) produced differently sized aggregates of CMP, largely depending on the pH of the hydrolysis. These results indicate that, whereas CMP molecules...

  15. Casein phosphopeptides drastically increase the secretion of extracellular proteins in Aspergillus awamori. Proteomics studies reveal changes in the secretory pathway

    Directory of Open Access Journals (Sweden)

    Kosalková Katarina


    Full Text Available Abstract Background The secretion of heterologous animal proteins in filamentous fungi is usually limited by bottlenecks in the vesicle-mediated secretory pathway. Results Using the secretion of bovine chymosin in Aspergillus awamori as a model, we found a drastic increase (40 to 80-fold in cells grown with casein or casein phosphopeptides (CPPs. CPPs are rich in phosphoserine, but phosphoserine itself did not increase the secretion of chymosin. The stimulatory effect is reduced about 50% using partially dephosphorylated casein and is not exerted by casamino acids. The phosphopeptides effect was not exerted at transcriptional level, but instead, it was clearly observed on the secretion of chymosin by immunodetection analysis. Proteomics studies revealed very interesting metabolic changes in response to phosphopeptides supplementation. The oxidative metabolism was reduced, since enzymes involved in fermentative processes were overrepresented. An oxygen-binding hemoglobin-like protein was overrepresented in the proteome following phosphopeptides addition. Most interestingly, the intracellular pre-protein enzymes, including pre-prochymosin, were depleted (most of them are underrepresented in the intracellular proteome after the addition of CPPs, whereas the extracellular mature form of several of these secretable proteins and cell-wall biosynthetic enzymes was greatly overrepresented in the secretome of phosphopeptides-supplemented cells. Another important 'moonlighting' protein (glyceraldehyde-3-phosphate dehydrogenase, which has been described to have vesicle fusogenic and cytoskeleton formation modulating activities, was clearly overrepresented in phosphopeptides-supplemented cells. Conclusions In summary, CPPs cause the reprogramming of cellular metabolism, which leads to massive secretion of extracellular proteins.

  16. Influence of colloidal calcium phosphate level on the microstructure and rheological properties of rennet-induced skim milk gels

    DEFF Research Database (Denmark)

    Koutina, Glykeria; Knudsen, Jes Christian; Andersen, Ulf


    lactose, to obtain varying levels of micellar calcium and phosphorus but constant value of pH, serum and free calcium, and serum phosphorus. Bovine chymosin was added to the skim milk samples after dialysis and microstructural and rheological properties during gel formation were recorded at 30°C. Samples......Colloidal calcium phosphate is an essential part of casein micelles and being responsible for their stability. Different mineralization of casein micelles was obtained by acidification of skim milk to pH 6.5, 6.0 or 5.5, followed by a dialysis method, using simulated milk ultrafiltrate without...

  17. Partial calcium depletion during membrane filtration affects gelation of reconstituted milk protein concentrates. (United States)

    Eshpari, H; Jimenez-Flores, R; Tong, P S; Corredig, M


    Milk protein concentrate powders (MPC) with improved rehydration properties are often manufactured using processing steps, such as acidification and high-pressure processing, and with addition of other ingredients, such as sodium chloride, during their production. These steps are known to increase the amount of serum caseins or modify the mineral equilibrium, hence improving solubility of the retentates. The processing functionality of the micelles may be affected. The aim of this study was to investigate the effects of partial acidification by adding glucono-δ-lactone (GDL) to skim milk during membrane filtration on the structural changes of the casein micelles by observing their chymosin-induced coagulation behavior, as such coagulation is affected by both the supramolecular structure of the caseins and calcium equilibrium. Milk protein concentrates were prepared by preacidification with GDL to pH 6 using ultrafiltration (UF) and diafiltration (DF) followed by spray-drying. Reconstituted UF and DF samples (3.2% protein) treated with GDL showed significantly increased amounts of soluble calcium and nonsedimentable caseins compared with their respective controls, as measured by ion chromatography and sodium dodecyl sulfate-PAGE electrophoresis, respectively. The primary phase of chymosin-induced gelation was not significantly different between treatments as measured by the amount of caseino-macropeptide released. The rheological properties of the reconstituted MPC powders were determined immediately after addition of chymosin, both before and after dialysis against skim milk, to ensure similar serum composition for all samples. Reconstituted samples before dialysis showed no gelation (defined as tan δ=1), and after re-equilibration only control UF and DF samples showed gelation. The gelation properties of reconstituted MPC powders were negatively affected by the presence of soluble casein, and positively affected by the amount of both soluble and insoluble

  18. Genetic variation and posttranslational modification of bovine κ-casein: effects on caseino-macropeptide release during renneting. (United States)

    Jensen, Hanne B; Pedersen, Katrine S; Johansen, Lene B; Poulsen, Nina A; Bakman, Mette; Chatterton, Dereck E W; Larsen, Lotte B


    Chymosin-induced cleavage of κ-casein (κ-CN) occurs during the first enzymatic phase in milk coagulation during cheese manufacturing, where the hydrophilic C-terminal peptide of κ-CN, named caseino-macropeptide (CMP), is released into the whey. The CMP peptide is known for its rather heterogeneous composition with respect to both genetic variation and multiple posttranslational modifications, including phosphorylation and O-linked glycosylation. An approach of liquid chromatography coupled with mass spectrometry was used to investigate (1) the overall protein profile and (2) the release of various forms of CMP after addition of chymosin to individual cow milk samples from 2 breeds, Danish Jersey (DJ) and Danish Holstein-Friesian (DH). The cows were selected to represent distinct homo- and heterozygous types of the κ-CN genetic variants A, B, and E (i.e., genotypes AA, BB, AB, EE, and AE). Initially, investigation of the protein profile showed milk with κ-CN BB exhibited the highest relative content of κ-CN, whereas AE milk exhibited the lowest, and after 40min of renneting >90% of intact κ-CN was hydrolyzed by chymosin in milk representing all κ-CN genotype. By in-depth analysis of the CMP chromatographic profile, multiple CMP isoforms with 1 to 3 O-linked glycans (1-3 G) and 1 to 3 phosphate groups (1-3 P) were identified, as well as nonmodified CMP isoforms. The number of identified CMP isoforms varied to some extent between breeds (21CMP isoforms identified in DJ, 26CMP isoforms in DH) and between κ-CN genetic variants (CMP variant A being the most heterogeneous compared with CMP B and E), as well as between individual samples within each breed. The predominant forms of glycans attached to CMP were found to be the acidic tetrasaccharide {N-acetyl-neuraminic acid α(2-3)galactose β(1-3)[N-acetyl-neuraminic acid α(2-6)]N-acetyl galactose} or trisaccharides {N-acetyl-neuraminic acid α(2-3)galactose β(1-3)N-acetyl galactose and galactose β(1-3)[N

  19. Properties of buffalo Mozzarella cheese as affected by type of coagulante

    Directory of Open Access Journals (Sweden)

    Nawal S. Ahmed


    Full Text Available   Background. Mozzarella is one of several pasta filata or stretched curd cheeses that originated in Italy. The name pasta filata refers to a unique plasticizing and texturing treatments of the fresh curd in hot water that imparts to the finished cheese its characteristic fibrous structure and melting properties. Mozzarella cheese made from standardized buffalo milk with 3 and 1.5% fat. The effect of coagulant types (calf rennet, chymosin and Mucor miehei rennet on the cheese properties was carried out. Material and methods. Fresh raw buffalo milk and starter cultures of Streptococcus salvarius ssp. thermophilus and Lactobacillus delbrueckii ssp. bulgaricus were used. The coagulants were calf rennet powder (HA-LA, microbial rennet powder (formase ISOTL from Mucor miehei and chymosin derived by fermentation (CDF. Milk, curd, whey,kneading water and cheese were analysed. The slab gel electrophoresis patterns of Mozzarella cheese were also applied. Statistical analyses were also applied on the obtained data. Results. Recovery of DM of both curd and cheese decreased in case of using Mucor miehei rennet while the recovery of TP and fat content in both curd and cheese and their loss to whey and kneading water were nearly the same. Soluble nitrogen and soluble tyrosine and tryptophan contents elevated with increasing the storage period. Increasing rate of the soluble nitrogen in case of using Mucor miehei rennet was higher compared to that made with the other types of coagulant. TVFA content increased with advancing the storage period, also increased with increasing the fat content of the original milk fat used. No effect can be seen due to the coagulant types. The meltability increased with storage period progress. While the effect of the type of coagulant enzyme hade neglect effect on meltability fat leakage and oiling off. Mozzarella cheese made with Mucor miehei rennet obtained the highest firmness compared with those made using calf rennet, or

  20. Investigation on Enzyme Clotting Technology of Soft Cheese%软质干酪的酶凝乳工艺研究

    Institute of Scientific and Technical Information of China (English)

    吕斯思; 陈有亮; 金鸟君


    In order to study the best cheese production process of substitute clotting enzymes, L9(34)and L16(43) designs of experiments on calf rennet ,chymopapain and Mucor miehei chymosin combinations were applied to determine the best technological parameters on cheese production. The results show that the optimum parameters of single enzymes are:calf rennet 0.0125%,temperature 32 ℃,clotting time 4Smin;chymopapain 0.01%,temperature 33 ℃,clotting time 35 min; Mucor miehei chymosin 0.0035%,temperature 32 ℃,clotting time 40 min. The best technological parameters of enzyme combinations are: calf rennet 0.004%, chymopapain 0.006%,Mucor miehei chymosin 0.0023%,temperature 32 ℃,clot-ting time 30 min,which can effectively optimize cheese production process.%为了探求软质干酪的最佳酶凝乳工艺,采用L9(34)和L16(45)正交试验,研究了皱胃酶、木瓜酶、米黑毛霉酶三种单酶的最佳工艺以及复配酶最佳工艺,确定了皱胃酶添加量0.0125%,凝乳温度32℃,时间45min;木瓜酶添加量0.01%,凝乳温度33℃,时间35min;米黑毛霉酶添加量0.0035%,凝乳温度32C,时间40min;复配酶最佳工艺参数为皱胃酶用量0.004%,木瓜酶用量0.006%,米黑毛霉酶用量0.0023%,温度32℃,凝乳时间30min.结果表明,单酶复配具有协同作用,能够有效优化干酪酶凝乳工艺.

  1. Prospect and development of Xinjiang curd-cheese%新疆奶酪现状及前景探讨

    Institute of Scientific and Technical Information of China (English)

    古丽奴儿·吐拉西; 普燕; 张富春


    分别对新疆传统奶酪的种类、生产方法、酶凝奶酪和酸凝奶酪的凝乳机理、凝乳酶特性及其在新疆奶酪中的应用现状等问题进行了详细介绍,进而探讨了新疆奶酪产业的发展前景.%Coagulation is the fundamental process in cheese making. Cheese prepared by different methods have different flavors. In this paper , the classification and production of Xinjiang traditional cheese are introduced in detail, the mechanism of curd-cheese and acid-cheese ,characteristic and application of chymosin in Xinjiang cheese are discussed as well in this paper, which intend to describe the prospect for the Xinjiang curd-cheese.

  2. Influence of lamb rennet paste on the lipolytic and sensory profile of Murcia al Vino cheese. (United States)

    Ferrandini, E; Castillo, M; de Renobales, M; Virto, M D; Garrido, M D; Rovira, S; López, M B


    The influence of lamb rennet paste (71.1% chymosin, 177 international milk-clotting units/mL, 4.57U/g of lipase activity) during the ripening of Murcia al Vino goat cheese was studied. The aim of this study was to improve the knowledge of the effect of lamb rennet paste on the lipolytic patterns in this type of cheese by reference to the evolution of total and free fatty acids. A sensory analysis was carried out to compare cheeses made with commercial and paste rennet. The rennet paste showed higher lipolytic activity, enhancing the production of short-chain free fatty acids. In addition, the cheese produced with lamb rennet paste had a slightly more bitter and piquant taste, making it an attractive commercial alternative that can be used to develop new varieties of goat cheese.

  3. Influence of diet and of lamb slaughtering age on the coagulating properties of rennet paste

    Directory of Open Access Journals (Sweden)

    M. Albenzio


    Full Text Available Rennet composition varies along with several factors such as source (animal species, herbage, microbial and genetic, physical state (liquid, powder and paste, enzymatic composition (chymosin/ pepsin ratio, lipolitic enzymes. Main factors influencing rennet paste are diet (milk feeding, weaning and slaughtering age. Rennet paste composition influences cheese-making and ripening processes in cheese. Little is known about the effects of probiotic addition to milk substitute on the microbial status and enzyme profile of rennet, and on ripening characteristics of cheese. The aim of the study was to evaluate the effects of rearing systems and of lamb slaughtering age on microbial profile and time of coagulation of rennet pastes used for Pecorino Foggiano cheese-making.

  4. Systematic study of the employment of enzymatic catalysts on the formation of carbon-heteroatoms

    Directory of Open Access Journals (Sweden)

    Paula Vanessa S. Rizzo


    Full Text Available The classic thio-Michael reaction is the one that involves the addition of a sulfur atom like the nucleophile of the reaction to an electrophile α,β -insaturated. There are many reports in the literature which describe the need of the electrophile activation using, for instance, Lewis acid. However, there is a slope that aims this activation using biocatalysts such as enzymes. In this sense, this study aimed to analyze some enzymes on the thio-Michael reaction. Some biocatalysts were tested such as the enzymes like Lipase, Lipozye and Chymosine (Figure 1. The reactions results indicated that the application of these biocatalysts proportioned a shorter reaction time from one day to four hours with high yield levels.

  5. Mucor miehei's microbial rennin production characteristics in a fed-batch proccess

    Directory of Open Access Journals (Sweden)

    C. P. Sánchez Henao


    Full Text Available The Mucor miehei zygomycete produces an acid protease (EC: resembling calf rennet chymosin characteristics. It has been suggested that low glucose concentration levels could be why enzyme synthesis, co-mes to an end even though enzyme production is still great (Escobar and Barnett, 1993, 1995. To overcome this possible limitation, a two stage research process was designed; the relationship between protease production and sugar consumption was studied initially to determine the periods of time when enzyme production is still high and glucose concentration close to zero. The following stage concentrated on developing a glucose fed-batch process during the afore mentioned time periods to observe any increase or decrease in enzyme production. During the batch studies, it was found that maximum enzyme activity (EA was 165 UC/ml for an average glucose consumption rate of 0.1813 g/1 h. Based on the previous.

  6. Proteinases from buckwheat (Fagopyrum esculentum moench seeds: Purification and properties of the 47 kDa enzyme

    Directory of Open Access Journals (Sweden)

    Timotijević Gordana S.


    Full Text Available Aspartic proteinases from buckwheat seeds are analyzed. Three forms of 47 kDa, 40 kDa and 28 kDa, were purified from mature buckwheat seeds, while two forms of 47 kDa and 28 kDa were detected in developing buckwheat seeds using pepstatin A affinity chromatography. A form of 47 kDa was selectively precipitated from other forms by ammonium sulfate precipitation. This enzyme resembles the chymosin-like pattern of proteolytic activity, as it was shown using BSA and k-casein as substrates, clarifying its ability for milk-clotting. The 47 kDa aspartic proteinase form is localized in the membrane fraction. .

  7. Proteolysis of Livanjski cheese during ripening

    Directory of Open Access Journals (Sweden)

    Samir KALIT


    Full Text Available Livanjski cheese belongs to the group of hard cheeses which is traditionally produced in Livno (Bosnia and Herzegovina. Proteolytic changes during the ripening of Livanjski cheese have not been investigated extensively. The aim of this paper was to determine its proteolytic changes during the different stages of ripening. Five Livanjski cheeses (from raw cow’s or a mixture of sheep’s and cow’s milk were observed during the ripening to evaluate its typical proteolytic profile. An electophoretic profile of Livanjski cheese was determined by Urea-polyacrylamide gel electrophoresis (urea-PAGE and a densitometric evaluation of the urea-PAGE gels was performed using a densitometer. The water-soluble nitrogen fraction in the total nitrogen (WSN %TN and the 12%-TCA-soluble nitrogen fraction in the total nitrogen (TCA-SN %TN of the cheese were determined using the Kjeldahl method. Degradation of αs1-casein by chymosin caused a significant decrease (P < 0.05 of relative content of this protein in Livanjski cheese at the sixth week point of ripening. Due to the activity of chymosin on αs1-casein, αs1-I-casein and αs1-II-casein developed, which caused a significant increase (P < 0.05 of Index alpha. The relative ratio of β-casein significantly decreased (P < 0.05 during ripening leading to a significant accumulation (P < 0.05 of degraded product (sum γ1-casein, γ2-casein and γ3-casein. These proteolytic changes caused a significant increase (P < 0.05 of Index betta. Accumulation of medium, small peptides and amino acids caused a significant (P < 0.05 increase of the relative content of WSN %TN and TCA-SN %TN. In general, proteolysis of Livanjski cheese during ripening was moderate probably due to the low moisture content and low water activity, although it was produced from raw milk. Taking into account that the ratio β-casein : αs1-casein at the end of ripening was 1.46, it could be concluded that degradation of αs1-casein could be the

  8. Glycomacropeptide Attenuates Inflammation, Pruritus, and Th2 Response Associated with Atopic Dermatitis Induced by 2,4-Dinitrochlorobenzene in Rat (United States)

    Muñoz, Fabiola Carolina; Cervantes-García, Daniel; Jiménez, Mariela; Ventura-Juárez, Javier


    Atopic dermatitis (AD) is one of the most common skin diseases, whose incidence is increasing in industrialized countries. The epicutaneous application of a hapten, such as 2,4-dinitrochlorobenzene (DNCB), evokes an experimental murine AD-like reaction. Glycomacropeptide (GMP) is a dairy bioactive peptide derived from hydrolysis of κ-casein by chymosin action. It has anti-inflammatory, prebiotic, and immunomodulatory effects. The present study was aimed to investigate the effect of GMP administration on DNCB-induced AD in rats. The severity of inflammatory process, pruritus, production of cytokines, and total immunoglobulin E (IgE) content were measured, and the histopathological features were analyzed. GMP reduced the intensity of inflammatory process and edema of DNCB-induced dermatitis, with a significant decrease in eosinophils recruitment and mast cells hyperplasia. In addition GMP suppressed the serum levels of total IgE and IL-4, IL-5, and IL-13 expression in AD-lesions. Besides, the levels of IL-10 were significantly increased. Remarkably, GMP administration before AD-induction abolished pruritus in dermatitis-like reactions in the rats. Taken together, these results indicate that GMP has an inhibitory effect on AD by downregulating Th2 dominant immune response, suggesting GMP as a potential effective alternative therapy for the prevention and management of AD.

  9. Physiological and Biochemical Characteristics of Lactation in Half-lactating Yak

    Institute of Scientific and Technical Information of China (English)

    ZHENG Yu-cai; ZHONG Guang-hui; WANG Yong; PENG Xian-wen; MAO Yong-jiang; ZOU Si-xiang; CHEN Wei-hua; CHEN Jie


    Yaks of two lactating types, the half-lactating yak and the total lactating yak, were investigated in their milk compositions, milk coagulation properties, and contents of three kinds of hormones, glucose,and protein in plasma. The half-lactating Maiwa yak and Jiulong yak contained significantly higher contents of protein, fat, and activities of alkaline phosphatase and γ-glutamyl transpeptidase in milk than that of the corresponding total-lactating yak breeds, with reduced milk yield and similar lactose level and relative percentages of main milk protein components. The half-lactatin yak resembled yaks in a late stage of lactation in their biochemical composition of milk; however, significant differences were also observed, which indicated that the former was in a special stage of lactation quite different from dairy cows. Milk of the total-lactating or halflactating yaks could be coagulated normally by adding chymosin, with a similar coagulation time. No significant difference was observed between prolactin and progesterone concentrations in plasma of the half-lactating yaks and total-lactating yaks; however, half-lactating yaks had significantly lower level of oestradiol-17 β in plasma than total-lactating yaks.

  10. Biosynthesis of some Mannich bases

    Directory of Open Access Journals (Sweden)

    Bruna S. D. R. Aranha


    Full Text Available The Mannich reaction is extremely useful to form molecules that contain nitrogen groups. Currently, there is a great interest turned to this class of compounds due to its biological properties. The recent literature reports that the utilization of enzymes can be an adequate methodology on this type of reaction. According to the exposed above and with the intention of synthesizing the -aminoketones, some catalysts like Chymosin and also some microorganisms such as Saccharomyces cerevisiae, (sourdough bread and Lactobacilos (ATCC 90827LA10N19.10-E.U.A were applied. These reactions were carried out by using water as solvent at room temperature excelling for the Green Chemistry principles. Figure 1 shows the reaction scheme involving 1 eq.mol of aniline, 1,3 eq.mol of formaldehyde, 10 ml of water and 0,25g of the catalyst during four days. The reaction furnished satisfactory yields which are in accordance with the literature data.

  11. 水产品下脚料高值化利用技术研究现状%High-value Utilization Technology of Aquatic Waste

    Institute of Scientific and Technical Information of China (English)



    The deep development and utilization of aquatic waste is summarized briefly, these aquatic waste can be used in food natural fungicide, chymosin, thioguanine, chondroitin sulfate, dennatan sulfate, oligomeric peptides, small peptides oral solution, CMC, meaty flavor, fish enzyme, glyeopruteins and taurine, which provided a reference for further development and utilization of fish waste resources.%简单综述了水产品下脚料深度开发利用的研究情况,主要可应用在食品天然防霉剂、凝乳酶、硫鸟嘌呤(6 -TG)、硫酸软骨素、硫酸皮肤素、寡聚肽、小分子肽口服液、高吸收性钙剂(CMC)、热反应型肉香调味基料、鱼酶、糖蛋白、牛磺酸中,为进一步开发利用鱼下脚料资源提供参考.

  12. Recent patents on microbial proteases for the dairy industry. (United States)

    Feijoo-Siota, Lucía; Blasco, Lucía; Rodríguez-Rama, José Luis; Barros-Velázquez, Jorge; Miguel, Trinidad de; Sánchez-Pérez, Angeles; Villa, Tomás G


    This paper reviews the general characteristics of exo and endopeptidases of microbial origin currently used in the milk industry. It also includes recent patents developed either to potentiate the enzymatic activity or to improve the resulting milk derivatives. The main application of these proteases is in the cheese-making industry. Although this industry preferentially uses animal rennets, and in particular genetically engineered chymosins, it also utilizes milk coagulants of microbial origin. Enzymes derived from Rhizomucor miehei, Rhizomucor pusillus and Cryphonectria parasitica are currently used to replace the conventional milk-clotting enzymes. In addition, the dairy industry uses microbial endo and exoproteases for relatively new applications, such as debittering and flavor generation in cheese, accelerated cheese ripening, manufacture of protein hydrolysates with improved functional properties, and production of enzyme-modified cheeses. Lactic acid bacteria play an essential role in these processes, hence these bacteria and the proteases they produce are currently being investigated by the dairy industry and are the subject of many of their patent applications.

  13. A sialic acid assay in isolation and purification of bovine k-casein glycomacropeptide: a review. (United States)

    Nakano, Takuo; Ozimek, Lech


    Sialic acid is a carbohydrate moiety of k-casein glycomacropeptide (GMP), which is a 64 amino acid residue C-terminal sialylated phosphorylated glycopeptide released from k-casein by the action of chymosin during cheese making. GMP lacks aromatic amino acids including phenylalanine, tyrosine, and tryptophan. Because of its unique amino acid composition and various biological activities, GMP is thought to be a potential ingredient for dietetic foods (e.g., a food for PKU patients) and pharmaceuticals. Thus, increased attention has been given to the development of techniques to purify GMP. In this review, techniques of GMP purification described in patents and scientific research papers were introduced. A sialic acid assay is the important method to track GMP isolation and purification processes, for which the thiobarbituric acid reaction with 1-propanol as a chromophore extracting solvent is an inexpensive, practical and specific technique. Sephacryl S-200 gel filtration chromatography, cellulose acetate electrophoresis, and sodium dodecyl sulfate polyacrylamide gel electrophoresis are the major techniques to identify sialic acid specific to GMP. Sephacryl S-200 chromatography and cellulose acetate electrophoresis are also used to detect GMP sialic acid in whey pearmeate and whey added commercial margarine samples. Future research includes development of an economical industrial scale method to produce high purity GMP.

  14. Chemical and functional properties of glycomacropeptide (GMP) and its role in the detection of cheese whey adulteration in milk: a review. (United States)

    Neelima; Sharma, Rajan; Rajput, Yudhishthir Singh; Mann, Bimlesh


    Glycomacropeptide (GMP) is a C-terminal part (f 106-169) of kappa-casein which is released in whey during cheese making by the action of chymosin. GMP being a biologically active component has gained much attention in the past decade. It also has unique chemical and functional properties. Many of the biological properties have been ascribed to the carbohydrate moieties attached to the peptide. The unique set of amino acids in GMP makes it a sought-after ingredient with nutraceutical properties. Besides its biological activity, GMP has several interesting techno-functional properties such as wide pH range solubility, emulsifying properties as well as foaming abilities which are shown to be promising for applications in food and nutrition industry. These properties of GMP have given new dimension for the profitable utilization of cheese whey to the dairy industry. A number of protocols for isolation of GMP from cheese whey have been reported. Moreover, its role in detection of sweet/rennet whey adulteration in milk and milk products has also attracted attention of various researchers, and many GMP-specific analytical methods have been proposed. This review discusses the chemico-functional properties of GMP and its role in the detection methods for checking cheese or sweet whey adulteration in milk. Recent concepts used in the isolation of GMP from cheese whey have also been discussed.

  15. Major technological advances and trends in cheese. (United States)

    Johnson, M E; Lucey, J A


    Over the last 25 yr, cheese production in the United States has more than doubled with most of the increase due to production in the western states. Processing large volumes of milk into cheese has necessitated changes in vat size and design, reliance on computer software, and milk standardization, including use of membrane concentration of milk either at the cheese plant or on the farm. There has been increased interest in specialty cheeses including cheese made from sheep, goat, and organic milks. In addition, membrane processing of whey into various value-added components has become routine. Changes in cheese manufacturing protocols have resulted in a reduction of the manufacturing time and the necessity for consistent and reliable starter activity. Major advances in the genetics of microorganisms have not only resulted in widespread use of fermentation-produced chymosin but also in starter bacteria with improved resistance to bacteriophage infection. Genomics and proteomics have increased the likelihood of the development of nonstarter adjuncts with specific enzymatic activity. Indeed, the use of adjunct microorganisms to produce cheese with a unique flavor profile or to produce cheese with more consistent or better quality flavor has gained almost universal acceptance.

  16. Detection and confirmation of milk adulteration with cheese whey using proteomic-like sample preparation and liquid chromatography-electrospray-tandem mass spectrometry analysis. (United States)

    Campos Motta, T M; Hoff, R B; Barreto, F; Andrade, R B S; Lorenzini, D M; Meneghini, L Z; Pizzolato, T M


    Caseinomacropeptide (CMP) is a peptide released by chymosin in cheese production, remaining in whey. Thus, CMP can be used as a biomarker to fluid milk adulteration through whey addition. Commonly, CMP is analyzed by reversed phase (RP-HPLC) or size-exclusion chromatography (SEC). However, some psychrotropic microorganisms - specially Pseudomonas fluorescens - when present in storaged milk, can produce, by enzymatic pathway, a CMP-like peptide generally called pseudo-CMP. These two peptides differ from each other only by one amino acid. RP-HPLC and SEC methods are unable to distinguish these two peptides, which demand development of a confirmatory method with high selectivity. Considering the several degrees of glycosilation and phosphorylation sites in CMP, allied with possible genetic variation (CMP A and CMP B), analytical methods able to differentiate these peptides are extremely complex. In the present work, we developed a proteomic-like technique for separation and characterization of these peptides, using liquid chromatography coupled to mass spectrometry with electrospray ionization able to differentiate and subsequently quantify CMP and pseudo-CMP in milk samples in order to identify adulteration or contamination of these products. The method shows satisfactory precision (<11%) with a detection limit of 1.0 µg mL(-1) and quantification limit of 5.0 µg mL(-1). Specificity, matrix effects and applicability to real samples analysis were also performed and discussed.

  17. Interfacial and foaming interactions between casein glycomacropeptide (CMP) and propylene glycol alginate. (United States)

    Martinez, María J; Pizones Ruiz-Henestrosa, Víctor M; Carrera Sánchez, Cecilio; Rodríguez Patino, Juan M; Pilosof, Ana M R


    Proteins and polysaccharides are widely used in food formulation. While most of the proteins are surface active, only few polysaccharides can adsorb at the air-water interface; this is the case of propylene glycol alginates (PGA). It is known that casein glycomacropeptide (CMP), a bioactive polypeptide derived from κ-casein by the action of chymosin, presents a great foaming capacity but provides unstable foams. So, the objective of this work was to analyze the impact of mixing CMP and a commercial variety of PGA, Kelcoloid O (KO), on the interfacial and foaming properties at pH 7.0. It was determined the surface pressure isotherm, the dynamics of adsorption and the foaming properties for CMP, KO and the mixed system CMP-KO. CMP dominated the surface pressure of CMP-KO mixed system. The presence of KO synergistically improved the viscoelastic properties of surface film. The foaming capacity of CMP was altered by KO. KO foam presented a higher stability than CMP foam and it controlled the stability against drainage and the initial collapse in the mixed foam.

  18. The Role Of Milk Peptide As Antimicrobial Agent In Supporting Health Status

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    Eni Kusumaningtyas


    Full Text Available Antimicrobial peptide is commonly present in all species as a component of their innate immune defense against infection. Antimicrobial peptides derived from milk such as isracidin, casocidin, casecidin and other fragments with variety of amino acid sequence are released upon enzymatic hydrolysis from milk protein К-casein, α-casein, β-casein, α-lactalbumin and β- lactoglobulin. These peptides were produced by the activity of digestive or microbial protease such as trypsin, pepsin, chymosin or alcalase. The mode of action of these peptides is by interaction of their positive with negative charge of target cell membrane leading to disruption of membrane associated with physiological event such as cell division or translocation of peptide across the membrane to interact with cytoplasmic target. Modification of charged or nonpolar aliphatic residues within peptides can enhance or reduce the activities of the peptides against a number of microbial strains and it seems to be strain dependent. Several peptides act not only as an antimicrobial but also as an angiotensin-converting enzyme inhibitor, antioxidant, immunomodulator, antiinflamation, food and feed preservative. Although the commercial production of these peptides is still limited due to lack of suitable large-scale technologies, fast development of some methods for peptide production will hopefully increase the possibility for mass production.

  19. Potential of quixaba (Sideroxylon obtusifolium latex as a milk-clotting agent

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    Anna Carolina da Silva


    Full Text Available There are several obstacles to the use of chymosin in cheese production. Consequently, plant proteases have been studied as possible rennet substitutes, but most of these enzymes are unsuitable for the manufacture of cheese. The aim of this study was to evaluate the potential of latex from Sideroxylon obtusifolium as a source of milk-clotting proteases and to partially characterize the enzyme. The enzyme extract showed high protease and coagulant activities, with an optimal pH of 8.0 and temperature of 55 °C. The enzyme was stable in wide ranges of temperature and pH. Its activity was not affected by any metal ions tested; but was inhibited by phenylmethanesulfonyl fluoride and pepstatin. For the coagulant activity, the optimal concentration of CaCl2 was 10 µmol L- 1. Polyacrylamide gel electrophoresis showed four bands, with molecular weights between 17 and 64 kDa. These results indicate that the enzyme can be applied to the cheese industry.

  20. Affinity chromatography matrices for depletion and purification of casein glycomacropeptide from bovine whey. (United States)

    Baieli, María F; Urtasun, Nicolás; Martinez, María J; Hirsch, Daniela B; Pilosof, Ana M R; Miranda, María V; Cascone, Osvaldo; Wolman, Federico J


    Casein glycomacropeptide (CMP) is a 64- amino acid peptide found in cheese whey, which is released after κ-casein specific cleavage by chymosin. CMP lacks aromatic amino acids, a characteristic that makes it usable as a nutritional supplement for people with phenylketonuria. CMP consists of two nonglycosylated isoforms (aCMP A and aCMP B) and its different glycosylated forms (gCMP A and gCMP B). The most predominant carbohydrate of gCMP is N-acetylneuraminic acid (sialic acid). Here, we developed a CMP purification process based on the affinity of sialic acid for wheat germ agglutinin (WGA). After formation of chitosan beads and adsorption of WGA, the agglutinin was covalently attached with glutaraldehyde. Two matrices with different WGA density were assayed for CMP adsorption. Maximum adsorption capacities were calculated according to the Langmuir model from adsorption isotherms developed at pH 7.0, being 137.0 mg/g for the matrix with the best performance. In CMP reduction from whey, maximum removal percentage was 79% (specifically 33.7% of gCMP A and B, 75.8% of aCMP A, and 93.9% of aCMP B). The CMP was recovered as an aggregate with an overall yield of 64%. Therefore, the matrices developed are promising for CMP purification from cheese whey. © 2016 American Institute of Chemical Engineers Biotechnol. Prog., 33:171-180, 2017.

  1. Separation and characterization of mares' milk alpha(s1)-, beta-, kappa-caseins, gamma-casein-like, and proteose peptone component 5-like peptides. (United States)

    Egito, A S; Miclo, L; López, C; Adam, A; Girardet, J M; Gaillard, J L


    The equine alpha(s1)- and beta-caseins (CN) were purified by chromatography on DEAE-cellulose and by reversed-phase HPLC. The alpha(s1)-, beta-, and kappa-CN were characterized either by monodimensional urea-PAGE or sodium dodecylsulfate (SDS)-PAGE or by bidimensional electrophoresis. Kappa-casein was characterized after electrophoresis by glycoprotein-specific staining. To identify alpha(s1)-CN without ambiguity, internal sequences were determined after trypsin or chymosin digestion of purified alpha(s1)-CN. These sequences, that could be estimated to correspond to 62% of the full protein, presented strong identities with regions of alpha(s1)-CN primary structures of other species. In particular, 51, 48, 43, and 40% identities were obtained with corresponding regions of sow, dromedary, cow, and human alpha(s1)-CN, respectively. On the other hand, trace amounts of equine gamma-CN-like and proteose peptone component 5-like peptides were found in the whole CN. They were identified by microsequencing and corresponded to beta-CN peptides generated by plasmin action on the whole CN. The equine alpha(s1), beta-, and kappa-CN were separated by bidimensional electrophoresis in numerous isoelectric variants with apparent isoelectric points distributed between pH 4.4 to 6.3, 4.4 to 5.9, and 3.5 to 5.5, respectively. The beta- and kappa-CN displayed a more acidic character in the mare than in the cow.

  2. Effects of gelation temperature on Mozzarella-type curd made from buffalo and cows' milk: 2. Curd yield, overall quality and casein fractions. (United States)

    Hussain, Imtiaz; Yan, Jen; Grandison, Alistair S; Bell, Alan E


    The overall quality of Mozzarella-type curds made from buffalo and cows' milks were measured at gelation temperatures of 28, 34 and 39°C, and cutting times of 45, 60, 75 and 90min after chymosin addition. The curd yield and moisture content decreased with increasing gelation temperature, while whey fat losses increased. The effect of higher gelation temperature (39°C) was more pronounced in cows' milk than buffalo milk. This results in more fat losses and lower yields in both milk samples at a gelation temperature of 39°C. The minimum losses of fat and protein in rennet whey occurred at a gelation temperature of 34°C in both milk samples. The curd yield was higher in buffalo milk as compared to cows' milk. This is due to difference in total solids (fat and protein contents) of the two types of bovine milk. The different cutting times had a small effect on the yield and overall quality of curds made from both milk types. Curd moisture and loss tangent have a strong relationship with respect to effects of gelation temperature. Two different curd drainage methods (centrifugation and Buchner funnel filtration) were used to compare the final overall quality of Mozzarella-type curds made from both milk types. The α(s1) and β casein fractions were found to be in different proportions in the two milk types. The total- and casein bound-calcium were higher in buffalo milk than cows' milk. The total protein, casein and fat were also found to be higher in buffalo milk than cows' milk.

  3. Secretory Expression, Purification and Identification of Rennet from Mucor pusillus%微小毛霉凝乳酶的分泌表达、产物纯化及鉴定

    Institute of Scientific and Technical Information of China (English)

    王景会; 李玉秋; 郑丽; 李倬琳; 高岩; 杨贞耐


    The rennet from Mucor pusillus is one of the major sources of tnicrobial rennet, but it has some defects compared with the bovine chymosin. Aspartic proteinases from fungi are usually more heat resistant . When these enzymes are used for cheese making, the residue of enzymatic activity after heat treatment (55 ℃) following milk coagulation may still exist and cause proteolysis during cheese storage, bitter taste, softness and lower yield of the cheese products. In order to develop a rennet product with suitable milk clotting properties, the gene of rennet was cloned from Mucor pusillus, and the expression vector pPIC9K/M was constructed. The plasmids of pPIC9K/M was linearized with Sac I , and transformed into Pichia pastoris GS115 competent cells. The rennet was secretory expressed in Pichia pastoris successfully , and a strong band at about 46 kD was shown by SDS - PAGE. Activity tests showed that the rennet activity in the culture supernatant was 311.8 U/mL. The purity of recombinant rennet reached 92% with a 51.89% activity recovery.%采用基因工程的方法对微小毛霉(Mucor pusillus)凝乳酶进行分子改造,获得适于乳品工业生产用的凝乳酶,克隆到了微小毛霉凝乳酶基因,构建了酵母表达质粒pPIC9K/M。线性化后电击转入毕赤酵母GS115,在甲醇诱导下进行凝乳酶的初步发酵试验,通过pH反应及酶抑制反应试验证明,重组凝乳酶获得了分泌表达。SDS-PAGE分析表明重组凝乳酶的分子量约为46kD,与理论值(45.3 kD)基本相符,培养基上清液中凝乳酶的活性为311.8 U/mL,纯化的总回收率为51.89%,纯度达到92%。

  4. A comparison of the determination of the rennet coagulation properties of bovine milk

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    Roman Pytel


    Full Text Available The aim of the work was compared of two different methods (the visual method and the nephelo turbidimetry method for determination of rennet coagulation time. It was observed the effect of heat treatment of milk; types of rennet and addition of different amount of CaCl2 into the pasteurized milk. It was used two different chymosin rennet. For the visual method was milk sample (100 mL equilibrated at 35 °C, 1 mL of rennet was added into milk and was measured the time required for the first visible flakes (visual method. For the determination rennet coagulation time by nephelo-turbidimetry was removed part of milk with rennet and placed into nephelo-turbidimetry. Milk had a titratable acidity in the range from 6.5 to 7.0 °SH, average pH of milk was 6.68. Dry matter content was in range from 12.351 to 13.142%. The average content of protein by Kjeldahl was 3.14%, fat by Gerber 4.34%, lactose by polarimetry 4.68% and calcium content 1.1%. The pasteurized milk had the worst rennet coagulation time about 32 s compared to the raw milk. The difference coagulation time between milk with addition of 20 µL CaCl2 and 40 µL CaCl2 was in range 21 s to 26 s by visual method. The difference coagulation time between milk with addition of 20 µL CaCl2 and 40 µL CaCl2 was 15 s by nephelo-turbidimetry method. There occurred statistically non-significant differences in most of the measurements, comparing the visual and the nephelo-turbidimetric method. The heat treatment, addition of CaCl2 and using of different rennet had an influence on the curd category. It was obtained, that using nephelo-turbidimetry shown objective results for measuring the rennet coagulation time contrary the subjective visual method. Further, the results obtained by nephelo-turbidimetry are accurate and determined with the lower variation.

  5. Synthesis of some derivatives of compounds β-aminoketonic through Mannich reaction by usingbiocatalysts

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    Ana Claudia S. Lima


    : lipase, Lipozyme® and chymosin. The reactions were kept under stirring at room temperature from 24 to 48 hours in those which the enzymes were usedlike catalysts. All the reactions were monitored by thin layer chromatography. The crude of the reaction was purified on a silica gel column (70-230 mesh and the obtained compounds were characterized by infrared spectroscopy, gas chromatography and nuclear magnetic resonance. All the reactions biocatalyzed by enzymes showed biocatalytic potential, but their concentration, the basicity of the amine and the reaction medium interfered in the reaction yield. The both three enzymes presented a higher affinity as for the less polar solvents (THF as for the higher polar solvents(DMSO. However it was proposed the introduction of new low cost biocatalysts and environmentally safe on Mannich reactions.

  6. 母乳糖巨肽水平检测及其与多种初生婴儿配方奶粉的比较%Detection of glycomacropeptide content in breast milk and comparison with varieties of newborn infant formula

    Institute of Scientific and Technical Information of China (English)

    瞿柳红; 黄龙光; 陶莉; 吴圣楣; 周伟


    Objective To detect the glycomacropeptide (GMP) content of human breast milk,and take it as reference of newborn infant formula milk powder to optimize the nutritional content of infant formulas.Methods Thirty primiparas who fit the following conditions were selected:healthy,no special diet habits,living stably,having adequate milk,aged between 25 to 39 years old,and term delivery,were selected.Then they were divided into 2 groups,the colostrum group and mature milk group,and each group had 15 cases.Each case had been collected of breast milk 5 mL(front milk).The gathering time of colostrum group was the 2nd day postpartum and mature milk group was the 42nd day postpartum.Hydrolyzing the breast milk by chymosin at 37 ℃,then the sialic acid(SA) content was detected by a sialic acid detection kit,and GMP content was represented by SA content.In addition,6 brands of formula milk powder were detected in the same way as breast milk after being made into standard liquid milk.The differences between groups were compared by analysis of variance (ANOVA).Results Best conditions for enzymolysis were:chymosin concentration 0.25 g/L,hydrolysis time 120 min.SA content of colostrum group was (3486.98 ± 406.70) mg/L,while mature milk group was (2687.95 ± 375.85) mg/L,as the former was significantly higher than the latter (P < 0.01),but the differences between individuals within each group were small (CVco1 =0.12,CVma ilk =0.14).The average level of SA content of various infant formulas was (1196.93 ± 608.40)mg/L,which was significantly lower than colostrum and mature milk(all P < 0.01).SA contents of various brands of formulas were various,and the difference among these brands was relatively big(CV =0.63).Conclusions The content of GMP in human colostrum was higher than that of the mature milk.The contents of the GMP in different brands of infant formula milk powder were uneven,and the measured values were quite different with human milk.However,because of the difference