Department of Food Science and Nutrition, University of Minnesota, St. Paul, MN 55108, USA.
Drug Metab Dispos. 2013 Jan;41(1):79-88. doi: 10.1124/dmd.112.048678. Epub 2012 Oct 3.
Rodent animal models have been widely used for studying neurologic and toxicological events associated with cocaine abuse. It is known that the mouse is more susceptible to cocaine-induced hepatotoxicity (CIH) than the rat. However, the causes behind this species-dependent sensitivity to cocaine have not been elucidated. In this study, cocaine metabolism in the mouse and rat was characterized through LC-MS-based metabolomic analysis of urine samples and were further compared through calculating the relative abundance of individual cocaine metabolites. The results showed that the levels of benzoylecgonine, a major cocaine metabolite from ester hydrolysis, were comparable in the urine from the mice and rats treated with the same dose of cocaine. However, the levels of the cocaine metabolites from oxidative metabolism, such as N-hydroxybenzoylnorecgonine and hydroxybenzoylecgonine, differed dramatically between the two species, indicating species-dependent cocaine metabolism. Subsequent structural analysis through accurate mass analysis and LC-MS/MS fragmentation revealed that N-oxidation reactions, including N-demethylation and N-hydroxylation, are preferred metabolic routes in the mouse, while extensive aryl hydroxylation reactions occur in the rat. Through stable isotope tracing and in vitro enzyme reactions, a mouse-specific α-glucoside of N-hydroxybenzoylnorecgonine and a group of aryl hydroxy glucuronides high in the rat were identified and structurally elucidated. The differences in the in vivo oxidative metabolism of cocaine between the two rodent species were confirmed by the in vitro microsomal incubations. Chemical inhibition of P450 enzymes further revealed that different P450-mediated oxidative reactions in the ecgonine and benzoic acid moieties of cocaine contribute to the species-dependent biotransformation of cocaine.
啮齿类动物模型已被广泛用于研究与可卡因滥用相关的神经和毒理学事件。众所周知，与大鼠相比，小鼠更容易受到可卡因诱导的肝毒性（C IH）的影响。然而，这种对可卡因的物种依赖性敏感性的原因尚未阐明。在这项研究中，通过基于 LC-MS 的代谢组学分析对尿液样本中的可卡因代谢进行了表征，并通过计算单个可卡因代谢物的相对丰度进一步进行了比较。结果表明，从酯水解产生的主要可卡因代谢物苯甲酰可卡因的水平在接受相同剂量可卡因治疗的小鼠和大鼠的尿液中相当。然而，来自氧化代谢的可卡因代谢物的水平，如 N-羟基苯甲酰基可卡因和羟基苯甲酰可卡因，在两种物种之间存在显著差异，表明可卡因代谢存在物种依赖性。通过精确质量分析和 LC-MS/MS 碎片分析进行的后续结构分析表明，包括 N-去甲基化和 N-羟化在内的 N-氧化反应是小鼠中的首选代谢途径，而在大鼠中则发生广泛的芳基羟化反应。通过稳定同位素示踪和体外酶反应，鉴定并阐明了小鼠特异性 N-羟基苯甲酰基可卡因的α-葡萄糖苷和大鼠中含量较高的一组芳基羟化葡萄糖醛酸。在两种啮齿动物物种之间，通过体外微粒体孵育证实了可卡因体内氧化代谢的差异。P450 酶的化学抑制进一步表明，可卡因的 ecgonine 和苯甲酸部分中不同的 P450 介导的氧化反应导致了可卡因的物种依赖性生物转化。
