Department of Biotechnology, Graduate School of Engineering, Osaka University, 2-1 Yamadaoka, Suita, Osaka 565-0871, Japan; Safety Research Laboratories, Dainippon Sumitomo Pharma Co. Ltd, 3-1-98 Kasugade-naka, Konohana-ku, Osaka 554-0022, Japan.
J Appl Toxicol. 2011 Aug;31(6):524-35. doi: 10.1002/jat.1591. Epub 2010 Dec 10.
Gas chromatography-mass spectrometry (GC-MS) has great advantages for analyzing organic/amino acids, which are often targets in efficacy and/or toxicity studies. Although GC-MS has been used for the detection of many metabolic disorders, applications of GC-MS-based metabolomics in pharmacology/toxicology are relatively underdeveloped. We intended to investigate applicability of a GC-MS-based metabolomics approach for toxicological evaluation, and tried to elucidate the mechanism of hydrazine-induced hepatotoxicity. Rats were administered hydrazine chloride orally (120 and 240 mg kg⁻¹), and urine, plasma and liver samples were collected at 24 or 48 h post-dosing. Conventional clinical chemistry and liver histopathology were performed, urine and plasma were analyzed by GC-MS, and metabolic profiles were assessed using chemometric techniques. Principal component analysis score plots showed clear separation of the groups, indicating dose-dependent toxicity and recovery. The mechanism of toxicity was investigated based on semi-quantification data of identified metabolites. Amino acid precursors of glutathione (cystein, glutamate and glycine) and a product of glutathione metabolism (5-oxoproline) were elevated dose-dependently, accompanied with elevation of ascorbate levels. In addition, intermediates of the TCA cycle were decreased, whereas participants of the urea cycle and other amino acids were increased. These alterations were associated with histopathological changes such as fatty degeneration and glycogen accumulation. Application of GC-MS-based metabolomics revealed that oxidative stress and GSH consumption play important roles in the etiology of hydrazine-induced hepatotoxicity, demonstrating that this approach is a useful tool in pharmacology and toxicology for screening, elucidating mode of action and biomarker discovery.
气相色谱-质谱联用（GC-MS）在分析有机/氨基酸方面具有很大的优势，这些物质通常是疗效和/或毒性研究的目标。尽管 GC-MS 已被用于检测许多代谢紊乱，但基于 GC-MS 的代谢组学在药理学/毒理学中的应用相对较少。我们旨在研究基于 GC-MS 的代谢组学方法在毒理学评价中的适用性，并试图阐明肼诱导肝毒性的机制。大鼠经口给予盐酸肼（120 和 240 mg kg⁻¹），在给药后 24 或 48 小时收集尿液、血浆和肝组织样本。进行常规临床化学和肝组织病理学检查，用 GC-MS 分析尿液和血浆，并用化学计量学技术评估代谢谱。主成分分析得分图显示，各组之间有明显的分离，表明存在剂量依赖性毒性和恢复。根据鉴定代谢物的半定量数据，研究了毒性的机制。谷胱甘肽（半胱氨酸、谷氨酸和甘氨酸）的氨基酸前体和谷胱甘肽代谢产物（5-氧脯氨酸）呈剂量依赖性升高，同时伴有抗坏血酸水平升高。此外，三羧酸循环的中间产物减少，而尿素循环和其他氨基酸的参与者增加。这些变化与脂肪变性和糖原积累等组织病理学变化有关。基于 GC-MS 的代谢组学的应用表明，氧化应激和 GSH 消耗在肼诱导肝毒性的发病机制中起重要作用，证明该方法是药理学和毒理学中一种有用的筛选、阐明作用机制和发现生物标志物的工具。
