Chemical Proteomics-Based Reactive Metabolites and Protein Adducts: The New Perspective on the Metabolism of Food Furanocoumarins

Furanocoumarins are widely found in various foods and exhibit a range of biological activities. However, due to the structural characteristics of their furan ring, they are susceptible to being metabolized by cytochrome P450 enzymes, thereby forming reactive metabolites (RMs), leading to a dual effect: firstly, the inhibition of CYP enzyme activity, which may result in food-drug interactions affecting drug absorption; and secondly, the formation of covalent bonds between RMs and proteins, potentially disrupting protein function and causing toxicity. Consequently, understanding the metabolic metabolism of furanocoumarins in biological systems is essential for ensuring food safety. This study employed chemical proteomics-based RMs and protein adducts analytical approach to investigate mechanisms of psoralen (PRN), a representative simple-structured furanocoumarin. Initially, dose-dependent hepatotoxicity was confirmed in mice treated with excessive PRN exposure. Subsequently, an LC-MS strategy was developed to identify PRN RM-protein adducts both <em>in vitro</em> (liver microsomes) and <em>in vivo</em> (mouse liver). The abundance of PRN RM-protein adducts was positively correlated with the severity of liver injury. Further analysis identified CYP2C6, CYP3A1, CYP2E1, and CYP2A3 as the primary enzymes facilitating the formation of PRN RM-protein adducts. Finally, label-free quantitative proteomics, integrated with network pharmacology, indicated that disruption of glutathione metabolism may be a potential hepatotoxic pathway activated by PRN RM-protein adducts. This study provides new insights into the dual role of furanocoumarin-induced food-drug interactions and hepatotoxicity, establishes a methodological framework for assessing food safety, and offers critical insights for their judicious application.