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Separation of Calcium-binding Protein Derived from Enzymatic Hydrolysates of Cheese Whey Protein

  • Kim, S.B. ;
  • Shin, H.S. ;
  • Lim, J.W.
  • Received : 2003.09.01
  • Accepted : 2004.02.17
  • Published : 2004.05.01

Abstract

This study was carried out to separate the calcium-binding protein derived from enzymatic hydrolysates of cheese whey protein. CWPs (cheese whey protein) heated for 10 min at $100^{\circ}C$ were hydrolyzed by trypsin, papain W-40, protease S, neutrase 1.5 and pepsin, and then properties of hydrolysates, separation of calcium-binding protein and analysis of calcium-binding ability were investigated. The DH (degree of hydrolysis) and NPN (non protein nitrogen) of heated-CWP hydrolysates by commercial enzymes were higher in trypsin than those of other commercial enzymes. In the result of SDS-PAGE (sodium dodecyl sulphate polyacrylamide gel electrophoresis), $\beta$-LG and $\alpha$-LA in trypsin hydrolysates were almost eliminated and the molecular weight of peptides derived from trypsin hydrolysates were smaller than 7 kDa. In the RP-HPLC (reverse phase HPLC) analysis, $\alpha$-LA was mostly eliminated, but $\beta$-LG was not affected by heat treatment and the RP-HPLC patterns of trypsin hydrolysates were similar to those of SDS-PAGE. In ion exchange chromatography, trypsin hydrolysates were shown to peak from 0.25 M NaCl and 0.5 M NaCl, and calcium-binding ability is associated with the large peak, which was eluted at a 0.25 M NaCl gradient concentration. Based on the results of this experiment, heated-CWP hydrolysates by trypsin were shown to have calcium-binding ability.

Keywords

Cheese Whey Protein;Calcium-binding Protein;Enzymatic Hydrolysates

References

  1. Adamson, N. J. and E. C. Reynolds. 1996. Characterization of casein phosphopeptides prepared using alcalase: Determination of enzyme specificity. Enzyme Microb. Tech. 19:202.
  2. Bertrand-Harb, C., A. Baday, M. Dalgalarrondo, J. M. Chobert and T. Haetle. 2002. Thermal modifications of structure and codenaturation of $\alpha$-lactalbumin and $\beta$-lactoglobulin induce changes of solubility and susceptibility to proteases. Nahrung. 46:283.
  3. Bradford, M. M. 1976. A rapid and sensitive methods for the quantitation of microgram quantities of protein utilizing the principle of protein-dye binding. Anal. Biochem. 72:248.
  4. Chobert, J. M., C. Bertrand-Harb and M. G. Nicolas. 1988. Solubility and emulsifying properties of caseins and whey proteins modified enzymatically by trypsin. J. Agr. Food Chem. 36:883.
  5. Jolles, P., S. Levy-Tolendano, A. M. Fiat, C. Soria, D. Gillessen, A. Thomaidis, F. W. Dunne and J. P. Chen. 1993. Analogy between fibrinogen and casein: effect of an undecapeptide isolated from $\kappa$-casein on platelet function. Eur. J. Biochem. 158:379.
  6. Kilara, A. 1985. Enzyme-modified protein food ingredients. Biochem. 20:149.
  7. Laemmli, U. K. 1970. Cleavage of structural proteins during the assembly of the head of bacteriophage T4. Nature (Lord). 227:680.
  8. Meisel, H. and E. Schlimme. 1996. Bioactive peptides derived from milk proteins: ingredients for functional food? Kieler Milchwissenschaftliche Forshungsbeirchte. 48:343.
  9. Monti, J. C. and R. Jost. 1978. Enzymatic solubilization of heatdenatured cheese whey protein. J. Dairy Sci. 61:1233.
  10. Mullally, M. M., H. Meisel and R. J. FitzGerald. 1997. Angiotensin-I-converting enzyme inhibitory activities of gastric and pancreatic proteinase digestes of whey protein. Int. Dairy J. 7:299.
  11. Oukhatar, N. A., S. Bouhallab, F. Bureau, P. Arhan, J. L. Maubois and D. L. Bougle. 2000. In vitro digestion of caseinophosphopeptide-iron complex. J. Dairy Res. 67:125.
  12. Renner, E. 1994. Dairy foods: dairy calcium, bone metabolism and prevention of osteoporosis. J. Dairy Sci. 77:3498.
  13. Takada, Y., N. Kobayashi, K. Keto, H. Matsuyama, M. Yahiro and S. Aoe. 1997. Effects of whey protein on calcium and bone metabolism in ovariectomized rats. J. Nutr. Sci. Vitaminol (Tokyo). 43:199.
  14. Buttriss, J. 1990. The role of calcium in a balanced diet. J. Soc. Dairy Tech. 48:1.
  15. Lowry, O. H., N. J. Rosebrough, A. L. Farr and R. J. Randall. 1951. Protein measurement with the folin phenol reagent. J. Biol. Chem. 193:265.
  16. Rose, D., D. T. Davies and M. Yaguchi. 1969. Quantitative determination of the major components of casein mixture by column chromatography on DEAE-cellulose. J. Dairy Sci. 52:8.
  17. Noyelle, K. and H. van Deal. 2002. Kinetics of conformational changes induced by the binding of various metal ions to bovine $\alpha$-lactalbumin. J. Inorganic Biochem. 88:69.
  18. Fox, P. F. 1992. Advanced Dairy Chemistry-1: Proteins. Elservier Applied Science, London, New York.
  19. Jost, R. and J. C. Monti. 1977. Partial enzymatic hydrolysis of whey protein by trypsin. J. Dairy Sci. 60:1387.
  20. McIntosh, G. H., P. J. Royle, R. K. Le Leu, G. O. Regester, M. A. Johnson, R. L. Grinsted, R. S. Kenward and G. W. Snithers. 1998. Whey protein as functional food ingredient. Int. Dairy J. 8:425.
  21. Anon. 1998. Dairy components increasingly find uses in functional foods. The cheese reporter, April. 10:11.
  22. FitzGerald, R. J. and H. Meisel. 1999. Lactokinins: whey proteinderived ACE inhibitory peptides. Nahrung. 43:165.
  23. Friedlander, E. J. and A. W. Norman. 1980. Purification of chick intestinal calcium-binding protein. Methods in Enzymology 67:504.
  24. Ragno, V., P. G. Giampietro, G. Bruno and L. Businco. 1993. Allergenicity of milk protein hydrolysate formulae in children with cow milk allergy. Eur. J. Pediatr. 152:760. https://doi.org/10.1007/BF01953996
  25. Adler-Nissen, J. 1986. Enzymic hydrolysis of food proteins. Elservier Applied Science Publishers, New York, USA.
  26. Drewnowski, A. 1992. Food preference and the opioid peptide system. Trends Food Sci. Tech. 3:97.
  27. Reynolds, E. C. 1997. Anticariogenic casein phosphopeptides. Ministerial Review of Dental Services in Victoria, Australia.
  28. Gallaher, D. and M. Schmidl. 1998. Bioactive and nutraceutical entities found in whey. Paper presented at Institute of Food Technologists, Annual Meeting, June Atlanta, Georgia, USA.
  29. Kosikowski, F. 1982. Cheese and fermented milk foods. 2nd edn. Edward Brothers Inc., Michigan, USA. pp. 228-260.
  30. Veprintsev, D. B., M. Narayan, S. E. Permyakov, V. N. Uversky, C. L. Brooks, A. M. Cherskaya, E. A. Permyakov and L. J. Berliner. 1999. Fine tuning the N-terminus of a calcium binding protein: $\alpha$-lactalbumin. Proteins 37:65.
  31. De la Fuente, M. A., Y. Hemar, M. Tamehara, P. A. Munro and H. Singh. 2002. Process-induced changes in whey proteins during the manufacture of whey protein concentrates. Int. Dairy J. 12:361.
  32. Schmidt, D. G. and B. W. van Markwijk. 1993. Enzymatic hydrolysis of whey proteins. Influence of heat treatment of $\alpha$-LA and $\beta$-LG on their proteolysis by pepsin and papain. Neth. Milk Dairy J. 47:15.
  33. Guo, M. R., P. F. Fox and A. Flynn. 1995. Susceptibility of $\beta$-Lactoglobulin and Sodium Caseinate to Proteolysis by Pepsin and Trypsin. J. Dairy Sci. 78:2336.
  34. Marshall, W. E. 1994. In Functional Foods, Designer Foods, Pharmafoods, Nutraceuticals. (Ed. I. Goldberg). Chapmann and Hall, New York. pp. 242-260.
  35. Otte, J., M. Zakora, K. B. Qvist, C. E. Olsen and V. Barkholt. 1997. Hydrolysis of bovine $\beta$-lactoglobulin by various protease and identification of selected peptides. Int. Dairy J. 7:835.
  36. Miller, D. D. 1989. Calcium in the diet, food sources, recommanded intakes and nutritional bioavailability. Adv. Food Nutr. Res. 33:103.

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