The purity, spectral characteristics, and rate of nonenzymatic hydrolysis of 2,6-dichloro-4-nitrophenyl phosphate (DCNPP) were determined. Rates of DCNPP hydrolysis by prostatic acid phosphatase (PAP) and erythrocytic acid phosphatase (EAP) (both EC 3.1.3.2) were measured in the absence and in the presence of various alcohols. 1.5-Pentanediol was the most effective transphosphorylation agent for specifically enhancing the activity of PAP. 1,4-Butanediol also enhanced PAP activity but markedly inhibited EAP activity. Bovine and human serum albumin preparations also accelerated the hydrolysis of DCNPP. DCNPP can be used for the continuous or multipoint-rate assay of PAP.