Effect of Regular Organic Solvents on Cytochrome P450-Mediated Metabolic Activities in Rat Liver Microsomes: Fig. 1.

Abstract

ABSTRACT:The effects of regular organic solvents on the metabolic activities of various human cytochromes P450 (P450s) have been reported. However, very little is known about their influence on metabolic activities mediated by P450s in the rat liver microsomes (RLM). The purpose of this study was to investigate the effects of organic solvents such as methanol, acetonitrile, dimethyl sulfoxide (DMSO), acetone, and ethanol on CYP1A, CYP2C, CYP2D, CYP2E, and CYP3A-mediated metabolism using RLM. The results showed that the activities of most rat P450 enzymes appeared to be organic solvent-dependent, and the metabolism of the tested probes were remarkably reduced when the concentration of organic solvents was up to 5% v/v, whereas most organic solvents demonstrated no significant interference when the concentration was below 1%, with the exception of DMSO. In addition, organic solvents exhibited different inhibitory effects, for example, CYP2D and CYP2E showed a significant reduction of activities at lower concentrations of organic solvents. Hence, this phenomenon should be taken into consideration when designing in vitro metabolism studies of new chemical entities. Therefore, we recommend acetonitrile as the most suitable solvent for RLM incubations, and the content of organic solvent should be kept lower than 1% v/v. IntroductionCytochromes P450 (P450s) are the principal enzymes for the oxidation of drugs, environmental pollutants, and a large number of xenobiotics, which present in many mammalian organs such as lungs, kidneys, intestines, and livers. Organic solvents are routinely used to dissolve substrates or other chemicals in the microsomal incubations for drug metabolism, P450 inhibition, and induction studies. For example, ethoxyresorufin and pentoxyresorufin are both hydrophobic substrates as the biomarkers for CYP1A1 and CYP2B1, dimethyl sulfoxide (DMSO), or ethanol/bovine serum albumin (BSA) were added to increase the solubility in rat liver microsomes (RLM) incubation system (Rutten et al., 1992). Various studies have reported the effects of common organic solvents on the in vitro P450-mediated metabolic activities in human liver microsomes (HLM), human hepatocytes, or cDNA-expressed human microsomes (Cotreau-Bibbo et al., 1996;Draper et al., 1997;Chauret et al., 1998;Hickman et al., 1998;Busby et al., 1999;Coller et al., 1999;Palamanda et al., 2000). RLM were quite commonly used during in vitro studies (Yao et al., 2008;Bae et al., 2009;Pelkonen et al., 2009;Deroussent et al., 2010); however, fewer systemic studies about the effects of organic solvents on the activities of rat P450 enzymes have been reported, and those tested in rats were limited to an old system in which the S9 and older marker substrates were used (Kawalek and Andrews, 1980). A previous study (Martignoni et al., 2006) indicated that the species-specific P450 enzymes of CYP1A, CYP2C, CYP2D, and CYP3A showed appreciable interspecies differences in terms of catalytic activity between rats and humans. Hence, the sensitivity of the P450 en...

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