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A Sensitive and Specific Assay for Glutathione with Potential Application to Glutathione Disulphide, Using High-performance Liquid Chromatography–tandem Mass Spectrometry by Ross L Norris; Geoffrey K Eaglesham; Glen R Shaw; Maree J Smith; Robyn K Chiswell; Alan A Seawright; Michael R Moore is a scholarly article available to read on EtoBox.

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We have utilised the combination of sensitivity and specificity afforded by coupling high-performance liquid chromatography (HPLC) to a tandem mass spectrometer (MS-MS) to produce an assay which is suitable for assaying glutathione (GSH) concentrations in liver tissue. The sensitivity suggests it may also be suitable for extrahepatic tissues. The method has been validated for GSH using mouse liver samples and also allows the assay of GSSG. The stability of GSH under conditions relevant to the assay has been determined. A 20-ml amount of a diluted methanol extract of tissue is injected with detection limits of 0.2 pmol for GSH and 2 pmol for GSSG. The HPLC uses an Altima C (15034.6 mm, 5 mm) column at 18 358C. Chromatography utilises a linear gradient from 0 to 10% methanol in 0.1% formic acid over 5 min, with a final isocratic stage holding at 10% methanol for 5 min. Total flow rate is 0.8 ml / min. The transition from the M1H ion (308.1 m /z for GSH, and 613.3 m /z for GSSG) to the 162.0 m /z (GSH) and 355.3 m /z (GSSG) fragments are monitored.

Author

Ross L Norris; Geoffrey K Eaglesham; Glen R Shaw; Maree J Smith; Robyn K Chiswell; Alan A Seawright; Michael R Moore

Publisher

Elsevier Science; Elsevier ; Elsevier BV (ISSN 0378-4347)

Published

2001

Language

EN