Metabolic transformation of cyclopiazonic acid in liver microsomes from different species based on UPLC-Q/TOF-MS

To investigate the metabolic transformation of cyclopiazonic acid (CPA) in the liver of different species and to supplement accurate risk assessment information, the metabolism of CPA in liver microsomes from four animals and humans was studied using the ultra–high–performance liquid chromatography–quadrupole/time–of–flight method. The results showed that a total of four metabolites were obtained, and dehydrogenation, hydroxylation, methylation, and glucuronidation were identified as the main metabolic pathways of CPA. Rat liver microsomes exhibited the highest metabolic capacity for CPA, with dehydrogenated (C20H18N2O3) and glucuronic acid-conjugated (C26H28N2O10) metabolites identified in all liver microsomes except chicken, indicating significant species metabolic differences. Moreover, C20H18N2O3 was only detected in the incubation system with cytochromes P450 3A4 (CYP3A4). The hydroxylated (C20H20N2O4) and methylated (C21H22N2O3) metabolites were detected in all incubation systems except for the CYP2C9, with CYP3A4 demonstrating the strongest metabolic capacity. The “cocktail” probe drug method showed that CPA exhibited a moderate inhibitory effect on the CYP3A4 (IC50 value = 8.658μM), indicating that the substrate had a negative effect on enzyme activity. Our results provide new insights to understand the biotransformation profile of CPA in animals and humans. Environmental Implication Cyclopiazonic acid (CPA) is a secondary metabolite produced by certain species of aspergilli, with wide distribution in the environment. The co-occurrence of CPA with aflatoxin and ochratoxin has brought increased attention to CPA as an emerging mycotoxin. However, the metabolic process and mechanism in vivo of CPA remain unclear. Here, we have identified four metabolites of CPA for the first time, characterized the species metabolic differences of CPA and its interactions with four typical P450 enzymes. Our study revealed the metabolic transformation of CPA using a liver microsomal model, establishing a theoretical foundation for the risk assessment of CPA exposure.