Kinetics of the conjugation of aniline mustards with glutathione and thiosulfate

The rates of the non-enzymatic conjugation of the substituted aniline mustards, melphalan, chlorambucil and p-(N,N-bis(2-chloroethyl))toluidine with glutathione and thiosulfate were determined using nuclear magnetic resonance spectroscopy. Using this method, the disappearance of drug and the formation of both the mono-thioether and bis-thioether conjugates can be monitored directly. For glutathione conjugation, the rate constants for the formation of the first and second aziridinium intermediates were similar. With thiosulfate conjugation, the rate constant for the formation of the first aziridinium intermediate is greater than the rate constant for the formation of the second aziridinium. This demonstrates that the type of nucleophile has a significant influence on the overall alkylating activity of these bifunctional mustards. The bisthioether adduct formed from the reaction between p-(N:N-bis([2-C-13]-2-chloroethyl))toluidine and glutathione and thiosulfate can be identified and scrambling of the C-13 label in the product provides strong evidence that the alkylation must occur through an aziridinium intermediate. (C) 1997 Elsevier Science ireland Ltd.