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  • Title: Measurement of biological thiols and disulfides by high-performance liquid chromatography and electrochemical detection of silver mercaptide formation.
    Author: Kuninori T, Nishiyama J.
    Journal: Anal Biochem; 1991 Aug 15; 197(1):19-24. PubMed ID: 1952064.
    Abstract:
    A rapid and sensitive method is described for the measurement of picomole levels of the biological thiols glutathione, cysteine, penicillamine, cysteamine, and ergothioneine by a combination of high-performance liquid chromatography and electrochemical detection (ECD). The compounds were separated isocratically on a reversed-phase C18 column by ion-pair chromatography with a mobile phase containing 5 mM acetic acid and 2.5 mM sodium 1-octanesulfonate. After chromatographic separation, the eluate was combined with silver nitrate dissolved in ammonium nitrate buffer at pH 10.5. A platinum disc electrode was used at -0.1 V vs Ag/AgCl to detect the amount of silver ions that had been consumed by the reaction with thiols. For measurement of disulfide, S-sulfonation with sodium sulfite or electroreduction were used to cleave the disulfide, and the thiol anions produced were detected by HPLC-ECD as for the reduced forms. The method was used to assay thiols and disulfides in biological materials.
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