Evaluation of BEH C18, BEH HILIC, and HSS T3 (C18) Column Chemistries for the UPLC-MS-MS Analysis of Glutathione, Glutathione Disulfide, and Ophthalmic Acid in Mouse Liver and Human Plasma

Abstract

Glutathione (GSH), glutathione disulfide (GSSG), and ophthalmic acid (OA) are important biological oxidative stress biomarkers to be monitored in pathological and toxicological studies. With the advent of liquid chromatography tandem mass spectrometry (LC-MS-MS) technology, sensitive and selective analysis of these biomarkers in biological samples is now being performed routinely. Due to the hydrophilic and polar natures of GSH and its endogenous derivatives, achieving good retention, resolution, and peak shape is often a chromatographic challenge. In this study, three ultra-performance (UP) LC column chemistries (namely, BEH C18, BEH HILIC, and HSS T3 [C18]) are evaluated for the UPLC-MS-MS analysis of GSH, GSSG, and OA extracted from mouse liver and human plasma samples. The chromatographic parameters evaluated are retentivity, tailing factor, MS sensitivity, and resolution of the three analytes. Based on the optimized method for each column chemistry, our results indicate that the HSS T3 (C18) column chemistry affords the best retention and separation of these analytes when operated under the ultra high-pressure chromatographic conditions.

Stress hormones promote growth of B16-F10 melanoma metastases: an interleukin 6- and glutathione-dependent mechanism

“…GSH was determined, following procedures previously described [28], by liquid chromatography-mass spectrometry using a Quattro micro triple-quadrupole mass spectrometer (Micromass, Manchester, UK) equipped with a Shimadzu LC-10ADVP pump and SCL-10AVP controller system with an SIL-10ADVP autoinjector (Shimadzu Corporation, Kyoto, Japan). Cell processing was performed according to published methodology, where rapid N-ethylmaleimide derivatization was used to prevent GSH auto-oxidation [29].…”

Glutathione and Bcl-2 targeting facilitates elimination by chemoradiotherapy of human A375 melanoma xenografts overexpressing bcl-xl, bcl-2, and mcl-1

“…GSH in tumor and non-tumor tissues was determined, following procedures previously described [16], by liquid chromatography-mass spectrometry using a Quattro micro triple-quadrupole mass spectrometer (Micromass, Manchester, UK) equipped with a Shimadzu LC-10AD VP pump and SCL-10A VP controller system with an SIL-10AD VP autoinjector (Shimadzu Corporation, Kyoto, Japan). Tissue/blood sample collection and processing were performed according to published methodology, where rapid N-ethylmaleimide derivatization is used to prevent GSH auto-oxidation [17].…”

Analysis of 76 veterinary pharmaceuticals from 13 classes including aminoglycosides in bovine muscle by hydrophilic interaction liquid chromatography–tandem mass spectrometry

“…A 6 min re-equilibration step gave a total run time of 20 min. This re-equilibration step is rather long for a UPLC column but the equilibration of the column is very significant in HILIC in order to achieve retention time reproducibility [52]. The injection volume was set at 10 L. Spray voltage was set at 4000 V, sheath gas and auxiliary gas at 25 psi and 10 a.u., respectively and capillary temperature at 300 • C. SRM mode was used and the selected transitions, collision energies and tube lenses are presented in Table S1.…”