Preparation and utilization of a reagent for the isolation and purification of low-molecular-mass thiols

Problems inherent in the isolation of thiols from natural sources, such as oxidation, undesirable addition reactions, and low concentration of thiol species in cell-free extracts, can be circumvented by reversible derivatization to a less labile form which can be concentrated selectively. These objectives are realized by converting thiols to heterodisulfides in which the thiol partner is an apolar thiol with strong affinity for hydrophobic stationary phases. When reacted with 2-S-(2(')-thiopyridyl)-6-hydroxynaphthyldisulfide at pH<5, where most thiol species are relatively stable to atmospheric oxidation, mixed disulfides with 2-mercapto-6-hydroxynaphthalene as the apolar partner are obtained in good yield and can be concentrated onto a hydrophobic stationary phase. Such heterodisulfides exhibit excellent chromatographic properties when separated on reversed-phase media and the derivatization reaction can, therefore, be conveniently monitored. Following their isolation as the heterodisulfides the thiol species of interest are recovered by reduction and facile separation from the apolar 2-mercapto-6-hydroxynaphthalene partner.