Inhibitory Effect Of Schizandrin A On The Activity Of Cytochrome 3A In Rats

Inhibitory effect of Schizandrin A on the Activity of Cytochrome 3A in RatsDrug interaction is a common phenomenon which should not be neglected to optimize regimens for patients. It had been observed occasionally that the serum concentration of Tacrolimus, a drug mainly metabolized by cytochrome P450 3A, could be obviously raised by a Chinese traditional medicine -WUZHI Capsule which mainly consisted of Schizandrin A. The exact mechanism for this unexpected interaction remained unknown. To address this issue, we investigated the effect of Schizandrin A on hepatic cytochrome P450 3A (CYP3A) in SD rats by analyzing the ability of CYP3A to hydroxylate testosterone into 66-hydroxytestosterone.Part A. Establishment of the Assay for Determining CYP3A Activity in Rat Murine Hepatic MicrosomeLiver tissue was surgically obtained from adult SD rats and homogenized, and murine hepatic microsome was prepared from homogenate by differential centrifugation. The microsome protein was diluted to a final concentration of 20mg/ml. The total volume of incubation system was 500μl, including 1.0mg/ml of murine hepatic microsome, 100mM of Tris-HAc (pH 7.4), lmM of NADPH and testosterone (25, 50,100, 200,400mM). The system was first incubated for 5 minutes at 37.0°C, then NADPH was added to it and reaction was terminated by 25μl perchloric acid after 30 minutes. After centrifugating for 5min, supernatant was filtered with 0.45μm microporous filter for futher use. Hepatic microsome of each rat was established in 3 parallel tubes. High perfomance liquid chromatography (HPLC) was adopted to determine the concentration of 6β-hydroxytestosterone produced in the murine hepatic microsome incubation system. Chromatographic analysis was performed on chromatographic column of Hypersil-BDS C18 5μm 200mm×4.6mm (Dalian Elite Analytival Instruments Co.) at room temperature. Samples were eluted at a flow rate of 1ml/min from the column with mobile phase consisting of methanol, acetonitrile and water, and the eluted products were detected at wavelength 254nm.Results indicated that neither endogenous material nor metabolite in the hepatic microsome would interfere with the analysis of samples. An ideal linear standard curve...