A new colorimetric method for the serum enzyme assay was developed, using the color reaction of a ferrous salt complex and a primary aromatic amine. For this method new substrates containing primary aromatic amines were synthesized. The aromatic amine liberated by the enzyme reaction was measured by its color reaction with pentacyanoammineferroate treated with hydrogen peroxide. The determination was made at a wavelength of around 700 nm, at which the assay is not affected by colored substances in serum. Therefore a blank assay was not necessary. This method was applied to the assay of serum gamma-glutamyl transpeptidase, cystine aminopeptidase, and leucine aminopeptidase. The results obtained were in good agreement with those of the p-nitroanilide method also used to assay each enzyme. This new method can be used on a Technicon autoanalyzer system.