Himmelstein M W, Carpenter S C, Hinderliter P M, Snow T A, Valentine R
E.I. du Pont de Nemours and Company, Haskell Laboratory for Toxicology and Industrial Medicine, PO Box 50, 1090 Elkton Road, Newark, DE 19711, USA.
Chem Biol Interact. 2001 Jun 1;135-136:267-84. doi: 10.1016/s0009-2797(01)00214-9.
Based on analogy with butadiene and isoprene, the metabolism of beta-chloroprene (2-chloro-1,3-butadiene, CD) to reactive intermediates is likely to be a key determinant of tumor development in laboratory rodents exposed to CD by inhalation. The purpose of this study is to identify species differences in toxic metabolite (epoxide) formation and detoxification in rodents and humans. The in-vitro metabolism of CD was studied in liver microsomes of B6C3F1 mice, Fischer/344 and Wistar rats, Syrian hamsters, and humans. Microsomal oxidation of CD in the presence of NADP(+), extraction with diethyl ether, and analysis by GC-mass selective detection (MSD) indicated that (1-chloroethenyl)oxirane (CEO) was an important metabolite of CD in the liver microsomal suspensions of all species studied. Other potential water-soluble oxidative metabolites may have been present. The oxidation of CD was inhibited by 4-methyl pyrazole, an inhibitor of CYP 2E1. CEO was sufficiently volatile at 37 degrees C for vial headspace analysis using GC-MSD single ion monitoring (m/z=39). CEO was synthesized and used to conduct partition measurements along with CD and further explore CEO metabolism in liver microsomes and cytosol. The liquid-to-air partition coefficients for CD and CEO in the microsomal suspensions were 0.7 and 58, respectively. Apparent species differences in the uptake of CEO by microsomal hydrolysis were hamster approximately human>rats>mice. Hydrolysis was inhibited by 1,1,1-trichloropropene oxide, a competitive inhibitor of epoxide hydrolase. A preliminary experiment indicated that the uptake of CEO in liver cytosol by GSH conjugation was hamster>rats approximately mice (human cytosol not yet tested). In general, the results suggest that metabolism may help explain species differences showing a greater sensitivity for CD-induced tumorigenicity in mice, for example, compared with hamsters. Additional experiments are in progress to quantify the kinetic parameters of CD oxidation and CEO metabolism by enzymatic hydrolysis and conjugation by glutathione S-transferase for in cytosol. A future goal is to use the kinetic rates to parameterize a physiologically based toxicokinetic model and relate the burden of toxic metabolite to the cancer dose-response observed in experimental animals.
基于与丁二烯和异戊二烯的类比，β-氯丁二烯（2-氯-1,3-丁二烯，CD）代谢生成反应性中间体可能是通过吸入接触CD的实验啮齿动物肿瘤发生的关键决定因素。本研究的目的是确定啮齿动物和人类在有毒代谢物（环氧化物）形成和解毒方面的种属差异。在B6C3F1小鼠、Fischer/344和Wistar大鼠、叙利亚仓鼠以及人类的肝微粒体中研究了CD的体外代谢。在NADP(+)存在的情况下，CD的微粒体氧化、用乙醚萃取以及通过气相色谱-质量选择性检测（MSD）分析表明，（1-氯乙烯基）环氧乙烷（CEO）是所有研究物种肝微粒体悬浮液中CD的一种重要代谢物。可能还存在其他潜在的水溶性氧化代谢物。CD的氧化受到CYP 2E1抑制剂4-甲基吡唑的抑制。CEO在37℃时挥发性足够高，可使用气相色谱-质谱单离子监测（m/z = 39）进行小瓶顶空分析。合成了CEO，并将其与CD一起用于进行分配测量，并进一步探索肝微粒体和细胞溶质中CEO的代谢。微粒体悬浮液中CD和CEO的液-气分配系数分别为0.7和58。微粒体水解对CEO的摄取存在明显的种属差异，仓鼠>人类>大鼠>小鼠。水解受到环氧水解酶竞争性抑制剂1,1,1-三氯环氧丙烷的抑制。一项初步实验表明，谷胱甘肽结合对肝细胞质中CEO的摄取情况为仓鼠>大鼠≈小鼠（尚未测试人类细胞质）。总体而言，结果表明代谢可能有助于解释种属差异，例如，与仓鼠相比，小鼠对CD诱导的肿瘤发生表现出更高的敏感性。正在进行额外实验以量化CD氧化和CEO代谢的动力学参数，这些代谢通过酶促水解以及谷胱甘肽S-转移酶在细胞溶质中的结合来进行。未来的目标是利用动力学速率参数化基于生理学的毒代动力学模型，并将有毒代谢物的负荷与实验动物中观察到的癌症剂量反应联系起来。
