Research and Development, USANA Health Sciences Inc, Salt Lake City, UT, USA.
Biomed Chromatogr. 2020 Sep;34(9):e4854. doi: 10.1002/bmc.4854. Epub 2020 Jun 28.
Endogenous glutathione (GSH) and glutathione disulfide (GSSG) status is highly sensitive to oxidative conditions and have broad application as a surrogate indicator of redox status in vivo. Established methods for GSH and GSSG quantification in whole blood display limited utility in human plasma, where GSH and GSSG levels are ~3-4 orders of magnitude below those observed in whole blood. This study presents simplified sample processing and analytical LC-MS/MS approaches exhibiting the sensitivity and accuracy required to measure GSH and GSSG concentrations in human plasma samples, which after 5-fold dilution to suppress matrix interferences range from 200 to 500 nm (GSH) and 5-30 nm (GSSG). The utility of the methods reported herein is demonstrated by assay performance and validation parameters which indicate good sensitivity [lower limits of quantitation of 4.99 nm (GSH) and 3.65 nm (GSSG), and high assay precision (intra-assay CVs 3.6 and 1.9%, and inter-assay CVs of 7.0 and 2.8% for GSH and GSSG, respectively). These methods also exhibited exceptional recovery of analyte-spiked plasma samples (98.0 ± 7.64% for GSH and 98.5 ± 12.7% for GSSG). Good sample stability at -80°C was evident for GSH for up to 55 weeks and GSSG for up to 46 weeks, with average CVs <15 and <10%, respectively.
内源性谷胱甘肽 (GSH) 和谷胱甘肽二硫化物 (GSSG) 的状态对氧化条件高度敏感，并且作为体内氧化还原状态的替代指标具有广泛的应用。用于全血中 GSH 和 GSSG 定量的既定方法在人类血浆中显示出有限的实用性，其中 GSH 和 GSSG 水平比全血中观察到的低 3-4 个数量级。本研究提出了简化的样品处理和分析 LC-MS/MS 方法，具有测量人类血浆样品中 GSH 和 GSSG 浓度所需的灵敏度和准确性，经过 5 倍稀释以抑制基质干扰，范围从 200 到 500nm（GSH）和 5-30nm（GSSG）。本文报道的方法的实用性通过测定性能和验证参数得到证明，这些参数表明具有良好的灵敏度[定量下限为 4.99nm（GSH）和 3.65nm（GSSG），高测定精密度（GSH 和 GSSG 的日内 CV 分别为 3.6%和 1.9%，日间 CV 分别为 7.0%和 2.8%）。这些方法还表现出分析物加标血浆样品的出色回收率（GSH 为 98.0±7.64%，GSSG 为 98.5±12.7%）。GSH 在-80°C 下的稳定性良好，长达 55 周，GSSG 长达 46 周，平均 CVs<15%和<10%，分别。
