Major benznidazole metabolites in patients treated for Chagas disease: Mass spectrometry-based identification, structural analysis and detoxification pathways

Benznidazole is the drug of choice for the treatment of Chagas disease, but its metabolism in humans is unclear. Here, we identified and characterized the major benznidazole metabolites and their biosynthetic mechanisms in humans by analyzing the ionic profiles of urine samples from patients and untreated donors through reversed -phase UHPLC-ESI-QTOF-MS and UHPLC-ESI-QqLIT-MS. A strategy for simultaneous detection and fragmenta-tion of characteristic positive and negative ions was employed using information-dependent acquisitions (IDA). Selected precursor ions, neutral losses, and MS3 experiments complemented the study. A total of six phase-I and ten phase-II metabolites were identified and structurally characterized in urine of benznidazole-treated patients. Based on creatinine-corrected ion intensities, nitroreduction to amino-benznidazole (M1) and its subsequent N-glucuronidation to M5 were the main metabolic pathways, followed by imidazole-ring cleavage, oxidations, and cysteine conjugations. This extensive exploration of benznidazole metabolites revealed potentially toxic struc-tures in the form of glucuronides and glutathione derivatives, which may be associated with recurrent treatment adverse events; this possibility warrants further exploration in future clinical trials. Incorporation of this knowledge of the benznidazole metabolic profile into clinical pharmacology trials could lead to improved treatments, facilitate the study of possible drug-drug interactions, and even mitigation of adverse drug reactions.

Automated summary

In this study, the major benznidazole metabolites and their biosynthetic mechanisms in humans were identified and characterized through advanced analytical techniques. A total of sixteen metabolites were identified in the urine of benznidazole-treated patients, revealing the main metabolic pathways and potentially toxic structures. Incorporating this knowledge into clinical pharmacology trials could lead to improved treatments and better understanding of adverse drug reactions.