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Conventional liquid chromatography/triple quadrupole mass spectrometry based metabolite identification and semi-quantitative estimation approach in the investigation of in vitro dabigatran etexilate metabolism

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Abstract

Dabigatran etexilate (DABE) is an oral prodrug that is rapidly converted by esterases to dabigatran (DAB), a direct inhibitor of thrombin. To elucidate the esterase-mediated metabolic pathway of DABE, a high-performance liquid chromatography/mass spectrometry based metabolite identification and semi-quantitative estimation approach was developed. To overcome the poor full-scan sensitivity of conventional triple quadrupole mass spectrometry, precursor–product ion pairs were predicted to search for the potential in vitro metabolites. The detected metabolites were confirmed by the product ion scan. A dilution method was introduced to evaluate the matrix effects on tentatively identified metabolites without chemical standards. Quantitative information on detected metabolites was obtained using “metabolite standards” generated from incubation samples that contain a high concentration of metabolite in combination with a correction factor for mass spectrometry response. Two in vitro metabolites of DABE (M1 and M2) were identified, and quantified by the semi-quantitative estimation approach. It is noteworthy that CES1 converts DABE to M1 while CES2 mediates the conversion of DABE to M2. M1 and M2 were further metabolized to DAB by CES2 and CES1, respectively. The approach presented here provides a solution to a bioanalytical need for fast identification and semi-quantitative estimation of CES metabolites in preclinical samples.

The scheme of the semi-quantitative estimation approach

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Acknowledgements

This study was financially supported by grant R15GM096074 from the National Institute of General Medical Sciences.

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Authors and Affiliations

  1. College of Pharmacy, Department of Clinical Pharmacy, University of Tennessee Health Science Center, 881 Madison Ave., Room 358, Memphis, TN, 38163, USA

    Zhe-Yi Hu, Vanessa L. Herring & S. Casey Laizure

  2. College of Pharmacy, Department of Clinical Pharmacy, University of Tennessee Health Science Center, 881 Madison Ave., Room 346, Memphis, TN, 38163, USA

    Robert B. Parker

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  1. Zhe-Yi Hu
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  2. Robert B. Parker
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  3. Vanessa L. Herring
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  4. S. Casey Laizure
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Correspondence to S. Casey Laizure.

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Hu, ZY., Parker, R.B., Herring, V.L. et al. Conventional liquid chromatography/triple quadrupole mass spectrometry based metabolite identification and semi-quantitative estimation approach in the investigation of in vitro dabigatran etexilate metabolism. Anal Bioanal Chem 405, 1695–1704 (2013). https://doi.org/10.1007/s00216-012-6576-4

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  • DOI: https://doi.org/10.1007/s00216-012-6576-4

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