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Human UDP-glucuronosyltransferase UGT1A4 forms tertiary N-glucuronides predominately with the energetically less favored tautomer of substituted 1H-indazole (benzpyrazole)

Fredenhagen, Andreas and Eggimann, Fabian and Kittelmann, Matthias and Lochmann, Thomas and Kuehnoel, Juergen (2017) Human UDP-glucuronosyltransferase UGT1A4 forms tertiary N-glucuronides predominately with the energetically less favored tautomer of substituted 1H-indazole (benzpyrazole). Journal of analytical science and technology., 8 (10). p. 10. ISSN 2093-3371; 2093-3134

Abstract

The in vitro glucuronidation of a substituted 1H-indazole (benzpyrazole), a transient receptor potential ankyrin 1 antagonist, was investigated. Depending on the mammalian liver used for production two peaks with different UV spectra and slightly different MS/MS spectra were found in the LC-MS/MS analysis. An optimized HPLC method gave a baseline separation of the two glucuronides. Preparation and isolation allowed to assign their structure by 1H-, 13C- and 15N- NMR and showed that both nitrogens of this structure were accessible to glucuronation with marked species difference. For example, the rodent mice produced both glucuronides in almost equal amounts, while rat liver formed mainly the 2- glucuronide indicating a different specificity of their UDP-glucuronosyltransferases. The glucuronide produced by human liver homogenate and by the majority of liver preparations from 11 animal species was attached at the energetically less favorable 2-position. Among 13 recombinant human UDP-glucuronosyltransferases tested only the isoenzyme 1A4 showed significant formation of both glucuronides. Again, the energetically less favorable N-glucuronide was formed in 33-fold excess. An MS/MS fragmentation mechanism was proposed for a fragment loss of 134 Da that differentiated the spectra of the two glucuronides.

Item Type: Article
Date Deposited: 13 Jun 2017 00:45
Last Modified: 13 Jun 2017 00:45
URI: https://oak.novartis.com/id/eprint/23802

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