One unit will hydrolyse 1,0 µmole of N-benzoyl-L-tyrosine ethyl ester (BTEE) per minute at pH 7,8 and 25 °C in the presence of ionised calcium.

Inhibitors: The enzyme is inhibited by heavy metals, the natural trypsin inhibitors to various degrees, an inhibitor from potato, and organophosphorus compounds. Gel filtration of chymotrypsin removes autolysis products and other contaminants. The specificity of α-chloroketone as α-chymotrypsin inhibitor has been studied. It has been reported that phenothiazine-N-carbonyl chloride are specific for chymotrypsin inhibition. Also inhibited by AEBSF, α-1-antitrypsin, aprotinin, DFP, PMSF, TPCK and α-2-macroglobulin.