Mechanisms of oxidation of acetaminophen
The metabolism of drugs and other xenobiotics normally expediates elimination and detoxification. However, in some instances inert compounds are transformed into potent alkylating or arylating agents capable of eliciting tissue lesions. The analgesic acetaminophen, which can produce liver necrosis in overdose, is one example.The pathway initially considered responsible for biotransformation of acetaminophen to a toxic metabolite involved cytochrome P-450 oxidation to N-hydroxyacetaminophen with dehydration to the electrophile N-acetyl-p-benzoquinone imine (NAPQI). N-Hydroxyacetaminophen was synthesized, and carrier trapping experiments conducted to determine its formation rate in vitro. Inclusion of an N-hydroxyacetaminophen carrier pool in liver microsomal incubations of radiolabeled acetaminophen did not decrease radiolabled protein binding. Radioactivity was not present in the re-isolated carrier pool. Thus, N-hydroxyacetaminophen apparently is not an acetaminophen metabolite in vitro.An alternative peroxidatic mechanism was examined by comparing products of acetaminophen oxidation by P-450 and horseradish peroxidase. Peroxidase incubations of radiolabeled acetaminophen resulted in binding of radiolabel to added protein. Addition of L-ascorbic acid or glutathione, inhibitors of P-450-mediated acetaminophen binding, decreased binding catalyzed by perioxidase. An acetaminophen-glutathione adduct was isolated from incubations including glutathione identical by HPLC to that formed in P-450 incubations. A free radical was generated during the peroxidase acetaminophen oxidations as detected by ESR. It was concluded that acetaminophen reactive metabolites produced by P-450 and peroxidase may be the same, but from current understanding of the enzymes the mechanisms of oxidation probably differ.The purported arylating acetaminophen metabolite, NAPQI, was synthesized, and HPLC procedures were developed for its detection as an acetaminophen oxidation product in cumene hydroperoxide-supported P-450 incubations. Metabolite partitioning provided compelling evidence for NAPQI formation in microsomes by demonstrating similarities between the acetaminophen reactive metabolite and NAPQI. NAPQI was cytotoxic in isolated hepatocytes, and portal infusion demonstrated an in vivo toxicity histologically identical to that observed with acetaminophen. NADH, NADPH, and NADPH-cytochrome P-450 reductase rapidly reduced NAPQI. Calculations indicated that up to 80% of NAPQI formed enzymatically from acetaminophen may undergo reduction back to acetaminophen. NAPQI reacts as an electrophile and as an oxidant; both properties may be important in the mediation of acetaminophen-induced toxicity.
- Medicinal chemistry