Low-molecular-weight thiols play important roles in a variety of pathological processes and are closely associated with a wide range of diseases. In this study, a selective and sensitive method was developed for the simultaneous determination of all the 7 thiols occurring in the transsulfuration pathway (Cysteine (Cys), homocysteine (Hcys), glutathione (GSH), N-acetylcysteine (Nac), cysteinylglycine (CysGly), glutamylcysteine (GluCys) and cysteamine (CA)) in human serum by in-vitro stable isotope labeling - dispersive solid phase extraction - liquid chromatography - tandem mass spectrometry analysis (IL-DSPE-LC-MS/MS). In the proposed method, a pair of stable isotope-labeling reagents, BQB (ω-bromoacetonylquinolinium bromide) and BQB-D₇, were utilized to label thiols in human serum samples and thiol standards, respectively. The BQB labeled thiols which carry a positive charge were extracted and purified with C₈-SO₃H-based DSPE followed by LC-MS/MS analysis. Good linearities for 7 thiols occurring in the transsulfuration pathway were obtained with the coefficient of determination (R²) >0.9901. The limits of detection (LODs) were in the range of 0.7–6.0 nmol/L. The method was further applied to investigate the contents change of 7 thiols in human serum samples of type 2 diabetes mellitus (T2DM) patients and breast cancer (BC) patients. The results showed that the contents of these thiols occurring in the transsulfuration pathway significantly changed and were highly diseases-related. In addition, partial least squares discriminant analysis (PLS-DA) suggested excellent classification performance between patients and healthy controls. The findings indicated that these significantly changed thiols occurring in the transsulfuration pathway in T2DM patients and BC patients might serve as the indicator for the diagnosis of T2DM and BC. Taken together, the developed IL-DSPE-LC-MS/MS method provides a promising tool for the sensitive analysis of thiols from complex biological samples, which may promote the in-depth investigation of the functions of thiols.

低分子量硫醇在多种病理过程中起重要作用，并与多种疾病密切相关。在这项研究中，开发了一种选择性灵敏的方法，用于同时测定在转硫途径中出现的所有7种硫醇（半胱氨酸（Cys），高半胱氨酸（Hcys），谷胱甘肽（GSH），N-乙酰半胱氨酸（Nac），半胱氨酰甘氨酸（ CysGly），谷氨酰半胱氨酸（GluCys）和半胱胺由（CA））在人血清中在-体外稳定同位素标记-分散固相萃取-高效液相色谱-串联质谱分析（IL-DSPE-LC-MS / MS）。在提出的方法中，使用了一对稳定的同位素标记试剂BQB（ω（溴溴丙酮基喹啉鎓）和BQB- D ₇分别用于标记人血清样品和硫醇标准品中的硫醇。提取带有正电荷的BQB标记的硫醇，并用基于C ₈ -SO ₃ H的DSPE进行纯化，然后进行LC-MS / MS分析。测定系数（R ²）> 0.9901。检出限（LOD）在0.7–6.0 nmol / L的范围内。该方法进一步应用于调查2型糖尿病（T2DM）和乳腺癌（BC）患者的人血清样品中7种硫醇的含量变化。结果表明，在转硫途径中这些硫醇的含量发生了显着变化，并且与疾病高度相关。此外，偏最小二乘判别分析（PLS-DA）表明，患者和健康对照之间的分类性能极佳。这些发现表明，T2DM患者和BC患者在转硫途径中发生的这些显着改变的硫醇可能是诊断T2DM和BC的指标。在一起