1 Bioactivation of sulphamethoxazole (SMX) to chemically-reactive metabolites and subsequent protein conjugation is thought to be involved in SMX hypersensitivity. We have therefore examined the cellular metabolism, disposition and conjugation of SMX and its metabolites in vitro. 2 Flow cytometry revealed binding of N-hydroxy (SMX-NHOH) and nitroso (SMX-NO) metabolites of SMX, but not of SMX itself, to the surface of viable white blood cells. Cellular haptenation by SMX-NO was reduced by exogenous glutathione (GSH). 3 SMX-NHOH and SMX-NO were rapidly reduced back to the parent compound by cysteine (CYS), GSH, human peripheral blood cells and plasma, suggesting that this is an important and ubiquitous bioinactivation mechanism. 4 Fluorescence HPLC showed that SMX-NHOH and SMX-NO depleted CYS and GSH in bu er, and to a lesser extent, in cells and plasma. 5 Neutrophil apoptosis and inhibition of neutrophil function were induced at lower concentrations of SMX-NHOH and SMX-NO than those inducing loss of membrane viability, with SMX having no e ect. Lymphocytes were signi®cantly (P50.05) more sensitive to the direct cytotoxic e ects of SMX-NO than neutrophils. 6 Partitioning of SMX-NHOH into red blood cells was signi®cantly (P50.05) lower than with the hydroxylamine of dapsone. 7 Our results suggest that the balance between oxidation of SMX to its toxic metabolites and their reduction is an important protective cellular mechanism. If an imbalance exists, haptenation of the toxic metabolites to bodily proteins including the surface of viable cells can occur, and may result in drug hypersensitivity.

A value for human liver microsomal scaling of 40 mg microsomal protein per gram liver has been established. The reason for underprediction previously reported for 52 different drug substrates was not the use of an incorrect value for the scaling factor.

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1 Cytochrome P450-mediated bioactivation of sulphamethoxazole to a hydroxylamine has been implicated in the hypersensitivity reactions associated with co-trimoxazole administration. Inhibiting the formation of the hydroxylamine may be one method of preventing the high frequency of toxicity which is observed in HIV-infected patients. Therefore, in this study, we have investigated the ability of fluconazole and ketoconazole, known cytochrome P450 inhibitors, to inhibit the formation of sulphamethoxazole hydroxylamine. 2 Ten healthy male volunteers were given co-trimoxazole (800 mg sulphamethoxazole and 160 mg trimethoprim) alone or 1 h after either fluconazole ( 150 mg) or ketoconazole (200 mg) in a randomized fashion with a washout period of at least 1 week between each phase. Urine was collected for 24 h, and sulphamethoxazole and its metabolites were quantified by electrospray LC-MS. 3 Ketoconazole had no effect on the urinary recovery of sulphamethoxazole or any of its metabolites. In contrast, fluconazole significantly (P<0.001) inhibited the formation of sulphamethoxazole hydroxylamine by 50.0±15.1%. Fluconazole also inhibited the oxidation of sulphamethoxazole to the 5-methylhydroxy and 5-methylhydroxy acetate metabolites by 69.9±15.8% and 64.0±12.0%, respectively, but had no effect on the amount of sulphamethoxazole, N 4 -acetyl sulphamethoxazole, or sulphamethoxazole N 1 -glucuronide excreted in urine. 4 The potential clinical benefit of using fluconazole to prevent hypersensitivity to co-trimoxazole in patients with AIDS needs to be assessed in a prospective study using both metabolite formation and the clinical occurrence of adverse reactions as end-points.

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