Premdas P D, Bowers R J, Forkert P G
Department of Anatomy and Cell Biology, Queen's University, Kingston, Ontario, Canada.
J Pharmacol Exp Ther. 2000 Jun;293(3):1112-20.
Diallyl sulfone (DASO(2)) inhibits hepatic CYP2E1. In this investigation, we have tested the hypothesis that an epoxide formed from DASO(2) is responsible for inactivation of hepatic CYP2E1 in mice. An epoxide of DASO(2) (1,2-epoxypropyl-3, 3'-sulfonyl-1'-propene; DASO(3)) was synthesized and conjugated to glutathione (GSH) to produce the conjugates S-(1R,S-[[ 1-hydroxymethyl-2,3'-sulfonyl]-1'-propenyl]ethyl)glutathione (diastereomers) and S-(1-[[2R,S-hydroxypropyl]-3, 3'-sulfonyl]-1'-propenyl)glutathione (diastereomers). Their identities were confirmed by (1)H NMR analysis, and these were used as analytical standards. HPLC analysis revealed a major peak for the GSH conjugates that eluted at 20.5 min. This peak was detected in liver microsomal incubations performed with DASO(2) in the presence of NADPH. A similar peak also was detected in incubations of CYP2E1-expressed lymphoblastoid microsomes, NADPH and DASO(2). The generation of the epoxide-derived GSH conjugates in the microsomal incubations was concentration-dependent, and reached saturation at 0. 75 to 1.0 mM DASO(2). Formation of the conjugates was also time-dependent and peaked at 2.0 h after DASO(2). Levels of DASO(3) formed from DASO(2), as estimated by production of a 4-(p-nitrobenzyl)pyridine derivative, were maximal at 1 mM DASO(2) at 30 min. CYP2E1-dependent p-nitrophenol hydroxylase activity was decreased in microsomes incubated with DASO(2), with alterations that were proportional to the concentration of DASO(2) (0.25-1.0 mM) used. Dose-dependent decreases in hydroxylase activity also were found in microsomes from mice treated in vivo with DASO(2) (25-200 mg/kg). These DASO(2)-induced decreases corresponded with reduced amounts of immunodetectable CYP2E1. Levels of spectrally detectable P450 and heme were both diminished by DASO(2). These results supported the contention that an epoxide formed from DASO(2) mediates the inactivation of hepatic CYP2E1.
二烯丙基砜（DASO(2)）可抑制肝脏中的细胞色素P450 2E1（CYP2E1）。在本研究中，我们验证了一个假说，即由DASO(2)形成的环氧化物是导致小鼠肝脏中CYP2E1失活的原因。合成了DASO(2)的环氧化物（1,2-环氧丙基-3,3'-磺酰基-1'-丙烯；DASO(3)），并使其与谷胱甘肽（GSH）结合，生成结合物S-(1R,S-[[1-羟甲基-2,3'-磺酰基]-1'-丙烯基]乙基)谷胱甘肽（非对映异构体）和S-(1-[[2R,S-羟丙基]-3,3'-磺酰基]-1'-丙烯基)谷胱甘肽（非对映异构体）。通过核磁共振氢谱（¹H NMR）分析确认了它们的结构，并将其用作分析标准品。高效液相色谱（HPLC）分析显示，谷胱甘肽结合物的一个主要峰在20.5分钟洗脱。在用DASO(2)和烟酰胺腺嘌呤二核苷酸磷酸（NADPH）进行的肝微粒体孵育中检测到了这个峰。在表达CYP2E1的淋巴母细胞微粒体、NADPH和DASO(2)的孵育中也检测到了类似的峰。微粒体孵育中环氧化物衍生的谷胱甘肽结合物的生成呈浓度依赖性，在0.75至1.0 mM DASO(2)时达到饱和。结合物的形成也是时间依赖性的，在加入DASO(2)后2.0小时达到峰值。通过4-(对硝基苄基)吡啶衍生物的生成来估算，由DASO(2)形成的DASO(3)的水平在30分钟时，1 mM DASO(2)时最高。与DASO(2)（0.25 - 1.0 mM）孵育的微粒体中，CYP2E1依赖性对硝基苯酚羟化酶活性降低，其变化与所用DASO(2)的浓度成比例。在用DASO(2)（25 - 200 mg/kg）体内处理的小鼠的微粒体中也发现了羟化酶活性的剂量依赖性降低。这些由DASO(2)诱导的降低与免疫检测到的CYP2E1量的减少相对应。DASO(2)使光谱可检测的细胞色素P450和血红素水平均降低。这些结果支持了由DASO(2)形成的环氧化物介导肝脏CYP2E1失活的观点。
