Abstract Optimum conditions for the assay of chick intestinal phytase were studied on native and commercial “technical” and “purified” preparations. Activity was near maximum when the substrate concentration was about 0.25 mM sodium phytate (1.5 mM phytate P) in the presence of 1.0 mM MgCl2 and pH about 7.2. Under these conditions two requirements for comparative enzyme studies were fulfilled: (i) a linear relationship between length of incubation period and the measured product; (ii) a linear relationship between enzyme concentration and the measured product. The activity was inhibited by excess substrate and inorganic orthophosphate. Phytase activity was compared to β-glycerophosphatase activity in regard to stability and effect of constituents found in native preparations. When stored at +5°C., both activities gradually increased over a period of one month. Freezing and thawing resulted in losses of phytase but not β-glycerophosphatase activity. Studies performed by adding inactivated native enzymes and the ash from mucosa to the commercial “purified” preparations showed that at the concentrations present in the assay, the organic constituents inhibited phytase activity and enhanced β-glycerophosphatase activity.Ultrafiltration studies also supported the conclusion that these constituents enhance β-glycerophosphatase activity, but not phytase activity. Studies on mixed substrates, phytate with phenyl phosphate or β-glycerophosphate did not establish the specificity of the enzyme.