Characterization of midazolam metabolism in locusts: the role of a CYP3A4-like enzyme in the formation of 1′-OH and 4-OH midazolam

Abstract

1. The metabolism of midazolam was investigated in vivo in locusts in order to evaluate the presence of an enzyme with functionality similar to human CYP3A4/5.

2. Hydroxylated metabolites of midazolam identical to human metabolites were detected in locusts and the apparent affinities (K_m values) were in the same range as reported in humans (in locusts: 7–23 and 33–85 µM for the formation of the 1′-OH and 4-OH metabolites, respectively).

3. The formation of hydroxylated metabolites could successfully be inhibited by co-administration of ketoconazole, a known CYP3A4/5 inhibitor.

4. Besides phase I metabolites, a number of conjugated metabolites were detected using high-resolution mass spectrometry. The most abundant metabolites detected were structurally identified by ¹H NMR as two N-glucosides. NMR analysis strongly suggested that the glycosylation occurred at the two nitrogens (either one in each case) of the imidazole ring.

5. Distribution of midazolam and the glucose conjugates were successfully measured using desorption electrospray mass spectrometry imaging revealing time-dependent changes in distribution over time.

6. In conclusion, it appears that an enzyme with functionality similar to human CYP3A4/5 is present in locusts. However, it appears that conjugation with glucose is the main detoxification pathway of midazolam in locusts.

• DESI-MS imaging
• mass spectrometry
• metabolite identification
• pharmacokinetics
• S. gregaria

Declaration of interest

This work was supported by The Danish National Advanced Technology Foundation (grant 023-2011-3) and The Carlsberg Foundation. The authors declare no conflicts of interests. The authors alone are responsible for the content and writing of this article.

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Supplementary Information