Reactions of benzene oxide with thiols including glutathione.
S-Phenylmercapturic acid is a minor metabolite of benzene used as a biomarker for human benzene exposures. The reaction of intracellular glutathione with benzene oxide-oxepin, the initial metabolite of benzene, is presumed to give 1-(S-glutathionyl)-cyclohexa-3,5-dien-2-ol, which undergoes dehydration to S-phenylglutathione, the precursor of S-phenylmercapturic acid. To validate the proposed route to S-phenylglutathione, reactions of benzene oxide-oxepin with glutathione and other sulfur nucleophiles have been studied. The reaction of benzene oxide with an excess of aqueous sodium sulfide, followed by acetylation, gave bis-(6-trans-5-acetoxycyclohexa-1,3-dienyl)sulfide, the structure of which was proved by X-ray crystallography. Reactions of benzene oxide-oxepin in a 95:5 (v/v) mixture of phosphate buffer in D2O with (CD3)2SO were monitored by 1H NMR spectroscopy. In the absence of glutathione, the half-life of benzene oxide-oxepin was ca. 34 min at 25 degrees C and pD 7.0. The half-life was not affected in the range of 2-15 mM glutathione in the presence and absence of a commercial sample of human glutathione S-transferase (at pH 7.0, 8.0, 8.5, or 10.0). The adduct 1-(S-glutathionyl)-cyclohexa-3,5-diene-2-ol was identified in these reaction mixtures, especially at higher pH, by mass spectrometry and by its acid-catalyzed decomposition to S-phenylglutathione. Incubation of benzene oxide with N-acetyl-L-cysteine at 37 degrees C and pH 10.0 and subsequent mass spectrometric analysis of the mixture showed formation of pre-S-phenylmercapturic acid and the dehydration product, S-phenylmercapturic acid. The data validate the premise that benzene oxide-oxepin can be captured by glutathione to give (1R,2R)- and/or (1S,2S)-1-(S-glutathionyl)-cyclohexa-3,5-dien-2-ol, which dehydrate to S-phenylglutathione. The capture is a relatively inefficient process at pH 7 that is accelerated at higher pH. These studies account for the observation that the metabolism of benzene is dominated by the formation of phenol. The pathway leading to S-phenylmercapturic acid is necessarily minor on account of the low efficiency of benzene oxide capture by glutathione at pH 7 vs spontaneous rearrangement to phenol.
Henderson
Alistair P
AP
School of Natural Sciences--Chemistry, Bedson Building, University of Newcastle upon Tyne, Newcastle upon Tyne, NEI 7RU, United Kingdom. b.t.golding@ncl.ac.uk
Barnes
Martine L
ML
Bleasdale
Christine
C
Cameron
Richard
R
Clegg
William
W
Heath
Sarah L
SL
Lindstrom
Andrew B
AB
Rappaport
Stephen M
SM
Waidyanatha
Suramya
S
Watson
William P
WP
Golding
Bernard T
BT
D003510:Cyclohexanes / Q000138:chemical synthesis* / Q000737:chemistry; D005978:Glutathione / Q000737:chemistry*; D006801:Humans; D008958:Models, Molecular; D008968:Molecular Conformation; D013438:Sulfhydryl Compounds / Q000138:chemical synthesis* / Q000737:chemistry
D003510:Cyclohexanes; D013438:Sulfhydryl Compounds; D005978:Glutathione; C023372:benzene oxide