โ Scribed by Etsushiro Doi; Daisuke Shibata; Teruyoshi Matoba
No coin nor oath required. For personal study only.
Four calorimetric procedures suitable for the determination of peptidase activity on peptides having a free (Y-or e-amino group are described. Two of the methods (A and B) are modifications of the conventional ninhydrin method described by S. Moore and W. H. Stein (( 1948) J. Viol. Chem. 176, 367-388; (1954) Ibid. 211, 907-913); the heating time is shortened to 5 min at 100ยฐC and the pH of the buffer in the reagent is lowered to 4.0. Method A differs from method B in buffer concentration. The other two methods (C and D) are modifications of the Cd-ninhydrin method described by A. P. Tsarichenko (( 1966) Nauch. Tr. Krasnodar. Gos. Pedagog. Inst. 70, 86-88, as cited in Chem. Abs. 67, 79479~); the water content in the reagent is reduced to l/20 of the original reagent and the sample is heated for 5 min at 84ยฐC. Method C differs from method D in the ratio of sample to reagent. In contrast to the free amino acids which are sufficiently colored, various peptides and amino acid derivatives except for the glycylpeptides give only a faint color with these methods. These four methods are not only useful for the determination of peptidase activity on peptides (e.g., Leu-Gly and tertbutyloxycarbonyl-glycyl-lysyl-leucine), but are also useful for the determinations of amidase activity on amino acid amides (e.g., Leu-NH1) and esterase activity on amino acid esters (e.g., tyrosine ethyl ester).
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