FOURIER-TRANSFORM ION-CYCLOTRON RESONANCE MASS-SPECTROMETRIC EVIDENCEFOR THE FORMATION OF ALPHA-CHLOROENETHIOLATES AND THIOKETENES FROM CHLOROALKENE-DERIVED, CYTOTOXIC 4-THIAALKANOATES

The cytotoxicity of chloroalkene-derived cysteine S-conjugates is thou ght to be associated with the formation of alpha-chloroenethiolates an d thioketenes as reactive intermediates. Recent studies indicate that the formation of 1,2-dichloroethenethiolate, which may give rise to ch lorothioketene, is a key step in the bioactivation of 5,6-dichloro-4-t hia-5-hexenoic acid (Fitzsimmons et al. (1995) Biochemistry 34, 4276-4 286). We report here the use of Fourier-transform ion cyclotron resona nce mass spectrometry to provide the first direct evidence for the for mation of alpha-chloroenethiolate and thioketene species from a cytoto xic 4-thiaalkanoate. The bioactivation of 5,6-dichloro-4-thia-5-hexeno ic acid involves conversion to the corresponding CoA thioester 5,6-dic hloro-4-thia-5-hexenoyl-CoA and subsequent processing by the fatty aci d beta-oxidation pathway. It has been proposed that the bioactivation of 5,6-dichloro-4-thia-5-hexenoyl-CoA involves lo ss of 1,2-dichloroet henethiolate, followed by loss of chloride to form chlorothioketene. 1 ,2-Dichloroethenethiolate and related alpha-chloroalkenethiolates have not been observed directly in aqueous solution. Fourier-transform ion cyclotron resonance mass spectrometric experiments show that S-propyl 5,6-dichloro-4-thia-5-hexenethioate reacts in the gas phase with base (hydroxide ion) to release 1,2-dichloroethenethiolate, which is obser ved directly in the mass spectrum of the products of the gas-phase rea ction. Furthermore, the elimination of chloride from 1,2-dichloroethen ethiolate on collision-induced decomposition is facile and provides ev idence for chlorothioketene formation. Preliminary evidence for the fo rmation of 1,2-dichloroethenethiolate and chlorothioketene from S-(1,2 -dichlorovinyl)-N-acetyl-L-cysteine methyl ester was also obtained. Th ese observations support the intermediacy of alpha-chloroenethiolates and chlorothioketenes in the bioactivation of cytotoxic, chloroalkene- derived 4-thiaalkanoates and cysteine S-conjugates and demonstrate the utility of Fourier-transform ion cyclotron mass spectrometry in study ing the formation of reactive intermediates.