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Evaluation of bovine rennets in terms of absolute concentrations of chymosin and pepsin A

Published online by Cambridge University Press:  01 June 2009

Patrice Martin ,
Jean-Claude Collin ,
Pascaline Garnot ,
Bruno Ribadeau Dumas  and
Germain Mocquot
Patrice Martin
Affiliation:
Institut National de la Recherche Agronomique, CNRZ
Jean-Claude Collin
Affiliation:
Institut National de la Recherche Agronomique, CNRZ
Pascaline Garnot
Affiliation:
Institut National de la Recherche Agronomique, CNRZ
Bruno Ribadeau Dumas
Affiliation:
Institut National de la Recherche Agronomique, CNRZ
Germain Mocquot
Affiliation:
Institut National de la Recherche Agronomique, CNRZ
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Summary

A method for determining chymosin and bovine pepsin A in commercial extracts of bovine veils, based on the use of the synthetic hexapeptide (Leu-Ser-Phe(NO2)-Nle-Ala-Leu-OMe) as reference substrate, is reported. Chymosin and bovine pepsin A were separated chromatographically from extracts and assayed for clotting activity on a reconstituted skim-milk standardized with reference chymosin and bovine pepsin A, themselves standardized in relation to the hexapeptide. The effect of pH on the absorbance difference between the hexapeptide and the Leu-Ser-Phe(NO2) tripeptide resulting from its hydrolysis was studied. It was found that the ‘optimal’ pH for determining the activities of the reference enzyme solutions was 4·7.

Six chymosin and 3 bovine pepsin A preparations were assayed on the hexapeptide to define the relation between the proteolytic activity and the amount of active enzyme. At pH 4·7 and 30 °C 1 mg chymosin and 1 mg bovine pepsin A hydrolysed 100 and 2700 μm-peptide/s respectively. The clotting activity of these preparations was assayed on a reconstituted skim-milk to standardize it and thus define the relation between the clotting time and the amount of active enzyme. Chymosin had a specific clotting activity twice that of bovine pepsin A. At equal clotting activities, bovine pepsin A was 55 times more active than chymosin on the hexapeptide at pH 4·7.

Type
Original Articles
Information
Journal of Dairy Research , Volume 48 , Issue 3 , October 1981 , pp. 447 - 456
Copyright
Copyright © Proprietors of Journal of Dairy Research 1981

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