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.