Department of Chemistry and Chemical Biology, McMaster University, Hamilton, Canada.
Anal Chem. 2010 Apr 1;82(7):2959-68. doi: 10.1021/ac9029746.
Despite several decades of active research, the success of large-scale clinical trials involving antioxidants remains equivocal given the complex biological interactions of reactive oxygen/nitrogen species in human health. Herein, we outline a differential metabolomics strategy by capillary electrophoresis-electrospray ionization-mass spectrometry (CE-ESI-MS) to assess the efficacy of nutritional intervention to attenuate oxidative stress induced by strenuous exercise. A healthy volunteer was recruited to perform a submaximal prolonged ergometer cycling trial until volitional exhaustion with frequent blood collection over a 6 h time interval, which included pre-, during, and postexercise periods while at rest. A follow-up study was subsequently performed by the same subject after high-dose oral intake of N-acetyl-L-cysteine (NAC) prior to performing the same exercise protocol under standardized conditions. Time-dependent changes in global metabolism of filtered red blood cell lysates by CE-ESI-MS were measured to reveal a significant attenuation of cellular oxidation associated with high-dose oral NAC intake relative to a control. Untargeted metabolite profiling allowed for the identification and quantification of several putative early- and late-stage biomarkers that reflected oxidative stress inhibition due to nutritional intervention, including oxidized glutathione (GSSG), reduced glutathione (GSH), 3-methylhistidine (3-MeHis), L-carnitine (C0), O-acetyl-L-carnitine (C2), and creatine (Cre). Our work demonstrates the proof-of-principle that NAC pretreatment is effective at dampening acute episodes of oxidative stress by reversible perturbations in global metabolism that can provide deeper insight into the mechanisms of thiol-specific protein inhibition relevant to its successful translation as a prophylaxis in clinical medicine.
尽管已经进行了几十年的积极研究，但由于活性氧/氮物种在人类健康中的复杂生物学相互作用，涉及抗氧化剂的大规模临床试验的成功仍然存在争议。在此，我们概述了一种通过毛细管电泳-电喷雾电离-质谱（CE-ESI-MS）进行差异代谢组学的策略，以评估营养干预减轻剧烈运动引起的氧化应激的功效。招募了一名健康志愿者进行亚最大强度的长时间踏车试验，直至自愿力竭，在 6 小时的时间间隔内频繁采集血液，包括运动前、运动中和运动后休息期间。随后，同一名志愿者在高剂量口服 N-乙酰-L-半胱氨酸（NAC）后进行了后续研究，然后在标准化条件下进行相同的运动方案。通过 CE-ESI-MS 测量过滤后的红细胞裂解物的全局代谢的时间依赖性变化，以揭示与高剂量口服 NAC 摄入相关的细胞氧化的显著衰减。非靶向代谢物分析允许鉴定和定量几种假定的早期和晚期生物标志物，这些标志物反映了由于营养干预而抑制氧化应激，包括氧化型谷胱甘肽（GSSG）、还原型谷胱甘肽（GSH）、3-甲基组氨酸（3-MeHis）、L-肉碱（C0）、O-乙酰-L-肉碱（C2）和肌酸（Cre）。我们的工作证明了 NAC 预处理通过可逆扰动全局代谢有效地抑制急性氧化应激发作的原理，这可以更深入地了解与成功转化为临床医学预防剂相关的硫醇特异性蛋白抑制的机制。
