Establish A Microbial Model With The Activity Of Drug-metabolizing Enzyme CYP2C9

According to the metabolism of four drugs : tolbutamide, gliburide, indometacin and diclofenac in mammalian and through seeing about Cunninghamella elegans, C.blakesleana , C.echinulata's , Mucor circinelloides AS 3.3421 and F17 abilities to transform tolbutamide, the initial microbial transformation models of CYP2C9 were set up. Three drugs metabolized by CYP2C9 (gliburide, diclofenac and indometacin) were used as substrates. Their metabolites and transformation yields were detected by liquid chromatography-mass spectrometry, and their major metablites were the same as in humans, hence the estabolished microbial models have extensive usability. The specific inhibitors of human cytochrome P450 2C9 were carried out to inhibit the 4'-hydroxylation of tolbutamide by C.blakesleana AS3.910, the result suggested that the correlated drug-metabolizing enzyme in Cunninghamella blakesleana AS3.910 was similar to CYP2C9 and that C.blakesleana AS3.910 can be used as microbial transformation model of CYP2C9.1. Metabolism of tolbutamide in microbial models The hypoglycemic drug tolbutamide is commonly used as a probe drug toevaluate CYP2C9 enzyme activity in terms of production of 4'-hydroxytolbutamide. In the present study, an initial screen of seven filamentous fungi was carried out to identify which was the most competent to convert tolbutamide into 4'-hydroxytolbutamide. From this screen, the fungus Cunninghamella blakesleeana AS 3.910 was selected as a suitable bioreactor. At a concentration of 1.2 mg/ml, the growing fungus transformed 95.0% of tolbutamide into 4'-hydroxytolbutamide in 96 h. With resting culture, the yield could reach 91.7% and exceeded 91.0% even when the tolbutamide concentration was increased to 4.0 mg/ml. On scale-up to 3 L buffer containing 12.0 g tolbutamide, 90% of tolbutamide was transformed into...