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Alkylation of human serum albumin by sulfur mustard in vitro and in vivo : Mass spectrometric analysis of a cysteine adduct as a sensitive biomarker of exposure

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Author: Noort, D. · Hulst, A.G. · Jong, L.P.A. de · Benschop, H.P.
Type:article
Date:1999
Source:Chemical Research in Toxicology, 8, 12, 715-721
Identifier: 235150
doi: doi:10.1021/tx9900369
Keywords: Carbon 14 · Cysteine 52-90-4 · Mustard gas 505-60-2 · Pronase · Serum albumin · Alkylation · Binding kinetics · Human · Human tissue · In vitro study · Iran · Mass spectrometry · Quantitative assay · Alkylating Agents · Alkylation · Amino acid sequence · Biological markers · Chemical warfare · Chemical warfare agents · High Pressure Liquid Chromatography · Hydrolysis · Molecular sequence data · Protein binding · Serum albumin · Trypsin · Trypsin, EC 3.4.21.4

Abstract

To develop a mass spectrometric assay for the detection of sulfur mustard adducts with human serum albumin, the following steps were performed: quantitation of the binding of the agent to the protein by using [14C] sulfur mustard and analysis of acidic and tryptic digests of albumin from blood after exposure to sulfur mustard for identification of alkylation sites in the protein. The T5 fragment containing an alkylated cysteine could be detected in the tryptic digest with micro-LC/tandem MS analysis. Attempts to decrease the detection limit for in vitro exposure of human blood by analysis of the alkylated T5 fragment were not successful. After Pronase treatment of albumin, S-[2-[(hydroxyethyl)thio]ethyl]Cys-Pro-Phe was analyzed by means of micro-LC/tandem MS, allowing a detection limit for in vitro exposure of human blood of 10 nM, which is 1 order of magnitude lower than that obtained by means of modified Edman degradation. The analytical procedure could be successfully applied to the analysis of albumin samples from Iranian victims of the Iran-Iraq war.