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A method for determination of γ-casein and its use for investigating proteolysis in bovine milk

Published online by Cambridge University Press:  01 June 2009

Yasuo Igarashi
Yasuo Igarashi
Affiliation:
Faculty of Agriculture, Hirosaki University, Hirosaki 036, Japan
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Summary

A convenient and sensitive method for determining γ-casein content of skim milk is described. It is based on quantitative separation in which the soluble fraction obtained from skim milk (1·5 ml) in a solvent system consisting of 50% (v/v) ethanol, 0·4 M-Na thiocyanate and 0·15 M-CaCl2 was applied to a small column of DEAE-cellulose and eluted with 0·02 M-Tris-HCl buffer (pH 8·0) containing 0·03 M-NaCl and 6 M-urea. Protein in this fraction (5 ml) was measured from its absorbance at 280 nm. The method is applicable also to heated skim milk. Its availability for investigating proteolysis in milk is expected from the fact that incubation of skim milk with porcine plasmin at 37 °C for 3 h increased γ-casein content, an increase that was lowered by addition of soyabean trypsin inhibitor. Incubation of raw skim milk with 0·02% (w/v) NaN3 resulted in increase in γ-casein with time and with increasing temperature. This was accompanied by a time lag, the length of which increased with decreasing temperature. The degree of proteolysis in skim milk, expressed as the amount of increase in γ-casein after 20 h of incubation at 37 °C, showed two pH optima, one at pH 8·0 and the other at pH 7·2–7·5. These results suggested that the present method.of γ-casein determination can be used for more precise studies of the milk plasmin system in which important factors, such as plasminogen activator and inhibitors, are involved.

Type
Original Articles
Information
Journal of Dairy Research , Volume 56 , Issue 4 , August 1989 , pp. 619 - 629
Copyright
Copyright © Proprietors of Journal of Dairy Research 1989

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References

REFERENCES

Andrews, A. T. 1978 a The composition, structure and origin of proteose-peptone component 5 of bovine milk. European Journal of Biochemistry 90 59–65CrossRefGoogle ScholarPubMed
Andrews, A. T. 1978 b The composition, structure and origin of proteose-peptone component 8F of bovine milk. European Journal of Biochemistry 90 67–71CrossRefGoogle ScholarPubMed
Andrews, A. T. 1983 Proteinases in normal bovine milk and their action on caseins. Journal of Dairy Research 50 45–55CrossRefGoogle ScholarPubMed
Barry, J. G. & Donnelly, W. J. 1980 Casein compositional studies. I. The composition of casein from Friesian herd milks. Journal of Dairy Research 47 71–81CrossRefGoogle Scholar
Blakesley, R. W. & Boezi, J. A. 1977 A new staining technique for proteins in polyacrylamide gels using Coomassie brilliant blue G250. Analytical Biochemistry 82, 580–582CrossRefGoogle ScholarPubMed
Davies, D. T. & Law, A. J. R. 1977 An improved method for the quantitative fractionation of casein mixtures using ion-exchange chromatography. Journal of Dairy Research 44 213–221Google Scholar
De Rham, O. & Andrews, A. T. 1982 The roles of native milk proteinase and its zymogen during proteolysis in normal bovine milk. Journal of Dairy Research 49 577–585CrossRefGoogle ScholarPubMed
Donnelly, W. J. & Barry, J. G. 1983 Casein compositional studies. III. Changes in Irish milk for manufacturing and role of milk proteinase. Journal of Dairy Research 50 433–441CrossRefGoogle Scholar
Driessen, F. M. & Van Der Waals, C. B. 1978 Inactivation of native milk proteinase by heat treatment. Netherlands Milk and Dairy Journal 32 245–254Google Scholar
Dulley, J. R. 1972 Bovine milk protease. Journal of Dairy Research 39 1–9Google Scholar
Eigel, W. N. 1977 Formation of γ1-A2, γ2-A2 and γ3-A caseins by in vitro proteolysis of β-casein A2 with bovine plasmin. International Journal of Biochemistry 8 187–192Google Scholar
Eigel, W. N. 1981 Identification of proteose-peptone component 5 as a plasmin-derived fragment of bovine β-casein. International Journal of Biochemistry 13 1081–1086CrossRefGoogle ScholarPubMed
Groves, M. L. & Kiddy, C. A. 1968 Polymorphism of γ-casein in cow's milk. Archives of Biochemistry and Biophysics 126 188–193CrossRefGoogle ScholarPubMed
Hipp, N. J., Groves, M. L., Custer, J. H. & McMeekin, T. L. 1950 Separation of γ-casein. Journal of the American Chemical Society 72 4928–4931CrossRefGoogle Scholar
Horne, D. S. & Parker, T. G. 1981 Factors affecting the ethanol stability of bovine milk. I. Effect of serum phase components. Journal of Dairy Research 48 273–284CrossRefGoogle Scholar
Humbert, G. & Alais, C. 1979 Review of the progress of Dairy Science: The milk proteinase system. Journal of Dairy Research 46 559–571Google Scholar
Igarashi, Y. 1982 Interactions of casein components with immobilized γ-casein. Japanese Journal of Zootechnical Science 53 56–63Google Scholar
Igarashi, Y. & Saito, Z. 1972 Properties of TS-casein in aqueous alcohol solutions. Japanese Journal of Zootechnical Science 43 31–38Google Scholar
Kaminogawa, S., Mizobuchi, H. & Yamauchi, K. 1972 Comparison of bovine milk protease with plasmin. Agricultural and Biological Chemistry 36 2163–2167CrossRefGoogle Scholar
Kaminogawa, S., Yamauchi, K. & Tsugo, T. 1969 Properties of milk protease concentrated from acid-precipitated casein. Japanese Journal of Zootechnical Science 40 559–565Google Scholar
Karman, A. H. & Van Boekel, M. A. J. S. 1986 Evaluation of the Kjeldahl factor for conversion of the nitrogen content of milk and milk products to protein content. Netherlands Milk and Dairy Journal 40 315–336Google Scholar
Kitchen, B. J. 1985 Indigenous milk enzymes. In Developments in Dairy Chemistry—3. Lactose and minor constituents pp. 239–279 (Ed. Fox, P. F.). London: Elsevier Applied Science PublishersGoogle Scholar
Korycka-Dahl, M., Ribadeau Dumas, B., Chene, N. & Martal, J. 1983 Plasmin activity in milk. Journal of Dairy Science 66 704–711CrossRefGoogle Scholar
Noomen, A. 1975 Proteolytic activity of milk protease in raw and pasteurized cow's milk. Netherlands Milk and Dairy Journal 29 153–161Google Scholar
Reimerdes, E. H. 1982 Changes in the proteins of raw milk during storage. In Developments in Dairy Chemistry—1. Proteins pp. 271–288 (Ed. Fox, P. F.). London: Applied Science PublishersGoogle Scholar
Reimerdes, E. H. & Herlitz, E. 1979 The formation of γ-caseins during cooling of raw milk. Journal of Dairy Research 46 219–221Google Scholar
Reimerdes, E. H., Mrowetz, G. & Klostermeyer, H. 1975 [Milk proteases. 4. Differentiation of micelle-associated activity by means of amino acid-4-nitroanilides.] Milchwissenschaft 30 271–274Google Scholar
Richardson, B. C. 1983 The proteinases of bovine milk and the effect of pasteurization on their activity. New Zealand Journal of Dairy Science and Technology 18 233–245Google Scholar
Richardson, B. C. & Pearce, K. N. 1981 The determination of plasmin in dairy products. New Zealand Journal of Dairy Science and Technology 16 209–220Google Scholar
Rollema, H. S., Visser, S. & Poll, J. K. 1981 On the determination, purification and characterization of the alkaline proteinase from bovine milk. Netherlands Milk and Dairy Journal 35 396–399Google Scholar
Rollema, H. H., Visser, S. & Poll, J. K. 1983 Spectrophotometric assay of plasmin and plasminogen in bovine milk. Milchwissenschaft 38 214–217Google Scholar
Schaar, J. 1985 Plasmin activity and proteose-peptone content of individual milks. Journal of Dairy Research 52 369–378Google Scholar
Snoeren, T. H. M. & Van Riel, J. A. M. 1979 Milk proteinase, its isolation and action on αs2- and β-casein. Milchwissenschaft 34, 528–531Google Scholar