Abstract: OBJECTIVE To study the effects of Neiyixiao on hepatic P450 enzymatic (CYP1A2, CYP2E1, CYP3A) activity in vitro by cocktail probe drugs. METHODS The amount of P450 enzyme were determined with Omura. Protein level was determined by ultraviolet spectrophotometer and bovine serum albumin was the standard substance. An HPLC method was used for the determination of caffeine, chlorzoxazone and nifedipine in the incubation system and the internal standard was antipyrine. The inhibitory effects of Neiyixiao on rat enzyme activities of CYP1A2, CYP3A and CYP2E1 were evaluated by cocktail probe drugs. RESULTS For the CYP1A2, half-life of the control group and administration group were (19.80±4.26)min and (16.63±2.66)min, respectively, but no significant difference. For the CYP2E1, half-life of the control group and administration group were (26.87±5.25)min and (18.35±8.55)min (P<0.05), respectively. For the CYP3A, half-life of the control group and administration group were (19.17±4.52)min and (29.50±7.43)min (P<0.05), respectively. CONCLUSION Administration "Neiyixiao" by master single test, the results showed that "Neiyixiao" could interact with some drugs metabolized by the above three enzymes.