Method of measuring activity of amino acid acylase

The paper describes a method to follow acylase activity. The method is based on spectrophotometry of the amino acid released, using o-phthalic aldehyde and mercaptoethanol. The major advantage of the method are its high sensitivity and speed. With the aid of the method kinetic parameters of hydrolysis of N-acetyl-L-methionine and N-acetyl-D,L-methionine catalyzed by pig kidney acylase were determined. Michaelis constants at pH 7.5 were estimated to be 5 +/- 1 and 10 +/- 2, respectively; this being in consistency with the data in the literature. It was shown that N-acetyl-D-methionine, acetate ion and methionine at the concentrations tested (0.01-0.05 M) did not inhibit acylase from a pig kidney.