The oxidized (GSSG) and reduced (GSH) forms of glutathione were quantified simultaneously in rat liver tissues by ion-pairing reverse phase high pressure liquid chromatography coupled to a coulometric electrochemical detector (HPLC-EC). Other biological thiols namely cysteine, cystathionine, homocysteine and methionine were shown to be well resolved from the glutathiones. Standard curves for GSH and GSSG were linear over the range of glutathione concentrations found in biological tissues with a correlation coefficient (r) > 0.999. Rat liver tissue content of GSH (16.96 ± 4.29, n = 5 and GSSG (0.467 ± 0.188 n = 5) found in this study are similar to results reported by other investigators. This method is also applicable to determine glutathione levels in rat bile samples. The advantages and disadvantages of employing coulometric over amperometric, as well as precautions in establishing HPLC-EC for the detection of oxidized and reduced glutathiones is discussed.
- Coulometric electrochemical detector