Hill B A, Kleiner H E, Ryan E A, Dulik D M, Monks T J, Lau S S
Division of Pharmacology and Toxicology, College of Pharmacy, University of Texas, Austin 78712.
Chem Res Toxicol. 1993 Jul-Aug;6(4):459-69. doi: 10.1021/tx00034a012.
Chemical reaction of 1,4-benzoquinone with GSH gives rise to several multisubstituted hydroquinone (HQ)-GSH conjugates, each of which causes renal proximal tubular necrosis when administered to male Sprague-Dawley rats. In addition, HQ has recently been reported to be nephrocarcinogenic following long-term exposure in male rats. Since neither the mechanism nor the extent of HQ oxidation and thioether formation in vivo is known, we have assessed both the qualitative and quantitative significance of HQ-thioether formation in vivo and in vitro. HQ (1.8 mmol/kg, ip) was administered to AT-125-pretreated male Sprague-Dawley rats, and bile and urine samples were analyzed with a HPLC-coulometric electrode array system (CEAS) and by liquid chromatography (LC)/continuous-flow fast atom bombardment (CF-FAB) mass spectroscopy. Five S-conjugates of hydroquinone were identified in bile, and one S-conjugate was identified in urine. The major biliary S-conjugate identified was 2-glutathion-S-ylhydroquinone [2-(GSyl)HQ] (18.9 +/- 2.7 mumol). Additional biliary metabolites were 2,5-diglutathion-S-ylhydroquinone [2,5-(diGSyl)HQ] (2.2 +/- 0.6 mumol), 2,6-diglutathion-S-ylhydroquinone [2,6-(diGSyl)HQ] (0.7 +/- 0.3 mumol),2,3,5-triglutathion-S-ylhydroquinone [2,3,5-(triGSyl)HQ] (1.2 +/- 0.1 mumol), and 2-(cystein-S-ylglycyl)hydroquinone. 2-(N-Acetylcystein-S-yl)HQ was the only urinary thioether metabolite (11.4 +/- 3.6 mumol) identified. The quantity of S-conjugates excreted in urine and bile within 4 h of HQ administration [34.3 +/- 4.5 mumol (4.3 +/- 1.1% of dose)] appears sufficient to propose a role for such metabolites in HQ-mediated nephrotoxicity and nephrocarcinogenicity. Rat liver microsomes catalyzed the NADPH-dependent oxidation of HQ (300 microM), in the presence of GSH, to form 2-(GSyl)HQ,2,5-(diGSyl)-HQ, and 2,6-(diGSyl)HQ. A fraction of the microsomal oxidation of HQ appears to be catalyzed by cytochrome(s) P450, although the exact amount remains unclear. 2-(GSyl)HQ,2,5-(diGSyl)-HQ, and 2,6-(diGSyl)HQ (300 microM) also underwent NADPH-dependent oxidation and GSH conjugation in liver microsomes. The extent of the nonenzymatic oxidation of HQ and its GSH conjugates correlated, approximately, with their half-wave oxidation potentials.
1,4 - 苯醌与谷胱甘肽（GSH）的化学反应会产生多种多取代对苯二酚（HQ）-GSH共轭物，将这些共轭物给予雄性斯普拉格 - 道利大鼠时，每一种都会导致肾近端小管坏死。此外，最近有报道称，长期暴露于HQ的雄性大鼠会发生肾致癌作用。由于体内HQ氧化和硫醚形成的机制及程度均未知，我们评估了体内和体外HQ - 硫醚形成的定性和定量意义。将HQ（1.8 mmol/kg，腹腔注射）给予经AT - 125预处理的雄性斯普拉格 - 道利大鼠，然后用高效液相色谱 - 库仑电极阵列系统（CEAS）以及液相色谱（LC）/连续流快原子轰击（CF - FAB）质谱法分析胆汁和尿液样本。在胆汁中鉴定出了五种对苯二酚的S - 共轭物，在尿液中鉴定出了一种S - 共轭物。鉴定出的主要胆汁S - 共轭物是2 - 谷胱甘肽 - S - 基对苯二酚[2 - (GSyl)HQ]（18.9±2.7 μmol）。其他胆汁代谢产物有2,5 - 二谷胱甘肽 - S - 基对苯二酚[2,5 - (diGSyl)HQ]（2.2±0.6 μmol）、2,6 - 二谷胱甘肽 - S - 基对苯二酚[2,6 - (diGSyl)HQ]（0.7±0.3 μmol）、2,3,5 - 三谷胱甘肽 - S - 基对苯二酚[2,3,5 - (triGSyl)HQ]（1.2±,0.1 μmol）以及2 - (半胱氨酸 - S - 基甘氨酸)对苯二酚。2 - (N - 乙酰半胱氨酸 - S - 基)HQ是唯一鉴定出的尿液硫醚代谢产物（11.4±3.6 μmol）。给予HQ后4小时内，尿液和胆汁中排泄的S - 共轭物量[34.3±4.5 μmol（占剂量的4.3±1.1%）]似乎足以表明这些代谢产物在HQ介导的肾毒性和肾致癌性中起作用。在谷胱甘肽存在的情况下，大鼠肝微粒体催化了HQ（300 μM）的NADPH依赖性氧化，形成2 - (GSyl)HQ、2,5 - (diGSyl) - HQ和2,6 - (diGSyl)HQ。虽然确切数量尚不清楚，但HQ的微粒体氧化似乎有一部分是由细胞色素P450催化的。2 - (GSyl)HQ、2,5 - (diGSyl) - HQ和2,6 - (diGSyl)HQ（300 μM）在肝微粒体中也经历了NADPH依赖性氧化和谷胱甘肽共轭作用。HQ及其谷胱甘肽共轭物的非酶促氧化程度与其半波氧化电位大致相关。
