title: Synthesis and in vitro toxicity of hydroxylamine metabolites of sulfonamides.
abstract: Among the most serious side effects of sulfonamides are hypersensitivity reactions, the pathogenesis of which has been suggested to be mediated by reactive metabolites. We have previously demonstrated dose-related covalent binding and toxicity of reactive intermediates of sulfonamides generated by a murine hepatic microsomal activating system. We hypothesized that hydroxylamine (H/A) metabolites might be likely candidates for mediating such toxicity; accordingly, we synthesized chemically the H/As of sulfadiazine and sulfamethoxazole. Synthesis was performed using 4-nitrobenzenesulfonyl chloride and either 2-aminopyrimidine or 3-amino-5-methylisoxazole, respectively, as starting materials. The resulting nitro derivatives were reduced to the corresponding H/A with hydrogen in the presence of a poisoned platinum catalyst. After synthesis and purification, toxicity of the H/As to lymphocytes of normal volunteers was evaluated using three cytotoxicity assays: 3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyl tetrazolium bromide dye conversion, trypan blue dye exclusion and propidium iodide dye exclusion. The H/As of sulfadiazine and sulfamethoxazole displayed dose-related toxicity. 1.6 mM sulfadiazine H/A produced 82% cell death, whereas 400 microM sulfamethoxazole H/A produced 62% cell death; the parent sulfonamides were not toxic to cells. The toxicity of sulfamethoxazole H/A was decreased by coincubation with glutathione or N-acetylcysteine; there was a 47% decrease in toxicity when coincubated with 100 microM glutathione, whereas there was a 55% decrease displayed when coincubation was done with 500 microM N-acetylcysteine. H/A metabolites of the sulfonamides or their nitroso derivatives, normally detoxified by conjugation to glutathione, may be the proximate toxins mediating sulfonamide hypersensitivity.
authors.affiliation: Department of Paediatrics, Faculty of Medicine, Hospital for Sick Children, Toronto, Ontario, Canada.
authors.forename: M J
authors.initials: MJ
authors.lastname: Rieder
authors.forename: J
authors.initials: J
authors.lastname: Uetrecht
authors.forename: N H
authors.initials: NH
authors.lastname: Shear
authors.forename: S P
authors.initials: SP
authors.lastname: Spielberg
mesh_terms: D002470:Cell Survival / Q000187:drug effects; D004305:Dose-Response Relationship, Drug; D005978:Glutathione / Q000494:pharmacology; D006801:Humans; D019811:Hydroxylamine; D006898:Hydroxylamines / Q000138:chemical synthesis / Q000633:toxicity*; D013449:Sulfonamides / Q000378:metabolism* / Q000633:toxicity
chemical_list: D006898:Hydroxylamines; D013449:Sulfonamides; D019811:Hydroxylamine; D005978:Glutathione