The inhibition kinetic parameters of pentachlorophenol (PCP) on bovine intestinal mucosa alkaline phosphatase (ALP) activity were studied by spectrophotometry. The results showed that PCP inhibited ALP activity in a concentration dependent manner, and the 50% inhibitory concentration (IC50) was estimated to be 4.20 mM. The apparent Michaelis–Menten constant (Km) and apparent maximum reaction rate (Vmax) was found to be decreased in the presence of PCP. Lineweaver–Burk plots indicated that the nature of the inhibition was of an uncompetitive type. Kinetic analysis indicated that the value of the inhibition constant (Ki) was 5.67 mM.