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Development, validation and application of a comprehensive stereoselective LC/MS-MS assay for bupropion and oxidative, reductive, and glucuronide metabolites in human urine.

Publication ,  Journal Article
Teitelbaum, AM; Flaker, AM; Kharasch, ED
Published in: J Chromatogr B Analyt Technol Biomed Life Sci
August 1, 2016

A stereoselective assay was developed for the quantification of bupropion and oxidative, reductive, and glucuronide metabolites (16 analytes total) in human urine. Initially, authentic glucuronide standards obtained from commercial sources were found to be incorrectly labeled with regard to stereochemistry; the correct stereochemistry was unequivocally reassigned. A trifurcated urine sample preparation and analysis procedure was employed for the stereoselective analysis of bupropion, hydroxybupropion, erythrohydrobupropion, and threohydrobupropion enantiomers, and hydroxybupropion, erythrohydrobupropion and threohydrobupropion β-d-glucuronide diastereomers in urine. Method 1 stereoselectively analyzed bupropion (R and S), and unconjugated free hydroxybupropion (R,R and S,S), erythrohydrobupropion (1R,2S and 1S,2R), and threohydrobupropion (1R,2R and 1S,2S) using chiral chromatography with an α1-acid glycoprotein column. Because no hydroxybupropion β-d-glucuronide standards were commercially available, method 2 stereoselectively analyzed total hydroxybupropion aglycones (R,R and S,S-hydroxybupropion) after urine hydrolysis by β-glucuronidase. Hydroxybupropion β-d-glucuronide (R,R and S,S) urine concentrations were calculated as the difference between total and free hydroxybupropion (R,R and S,S) concentrations. Due to incomplete β-glucuronidase hydrolysis of erythrohydrobupropion and threohydrobupropion β-d-glucuronide diastereomers, method 3 stereoselectively analyzed intact erythrohydrobupropion and threohydrobupropion β-d-glucuronide diastereomers using C18 column chromatography. All analytes were quantified by positive ion electrospray tandem mass spectrometry. The assay was fully validated over analyte-specific concentrations. Intra- and inter assay precision were within 15% for each analyte. The limits of quantification for bupropion (R and S), hydroxybupropion (R,R and S,S), threohydrobupropion (1S,2S and 1R,2R), erythrohydrobupropion (1R,2S and 1S,2R) were 10, 50, 100, and 100ng/mL, respectively. The limits of quantification for (1R,2R)-threohydrobupropion β-d-glucuronide, (1S,2S)-threohydrobupropion β-d-glucuronide, and (1R,2R)-erythrohydrobupropion β-d-glucuronide were each 50ng/mL. Due to the abundance of bupropion and metabolites in human urine, no efforts were made to optimize sensitivity. All analytes were stable following freeze thaw cycles at -80°C. This assay was applicable to clinical pharmacokinetic investigations of bupropion in patients and to in vitro metabolism of the primary bupropion metabolites to their glucuronides.

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Published In

J Chromatogr B Analyt Technol Biomed Life Sci

DOI

EISSN

1873-376X

Publication Date

August 1, 2016

Volume

1027

Start / End Page

239 / 253

Location

Netherlands

Related Subject Headings

  • Tandem Mass Spectrometry
  • Smoking Cessation
  • Oxidation-Reduction
  • Limit of Detection
  • Humans
  • Glucuronides
  • Chromatography, High Pressure Liquid
  • Bupropion
  • Antidepressive Agents, Second-Generation
  • 3401 Analytical chemistry
 

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Teitelbaum, A. M., Flaker, A. M., & Kharasch, E. D. (2016). Development, validation and application of a comprehensive stereoselective LC/MS-MS assay for bupropion and oxidative, reductive, and glucuronide metabolites in human urine. J Chromatogr B Analyt Technol Biomed Life Sci, 1027, 239–253. https://doi.org/10.1016/j.jchromb.2016.05.036
Teitelbaum, Aaron M., Alicia M. Flaker, and Evan D. Kharasch. “Development, validation and application of a comprehensive stereoselective LC/MS-MS assay for bupropion and oxidative, reductive, and glucuronide metabolites in human urine.J Chromatogr B Analyt Technol Biomed Life Sci 1027 (August 1, 2016): 239–53. https://doi.org/10.1016/j.jchromb.2016.05.036.
Teitelbaum, Aaron M., et al. “Development, validation and application of a comprehensive stereoselective LC/MS-MS assay for bupropion and oxidative, reductive, and glucuronide metabolites in human urine.J Chromatogr B Analyt Technol Biomed Life Sci, vol. 1027, Aug. 2016, pp. 239–53. Pubmed, doi:10.1016/j.jchromb.2016.05.036.
Journal cover image

Published In

J Chromatogr B Analyt Technol Biomed Life Sci

DOI

EISSN

1873-376X

Publication Date

August 1, 2016

Volume

1027

Start / End Page

239 / 253

Location

Netherlands

Related Subject Headings

  • Tandem Mass Spectrometry
  • Smoking Cessation
  • Oxidation-Reduction
  • Limit of Detection
  • Humans
  • Glucuronides
  • Chromatography, High Pressure Liquid
  • Bupropion
  • Antidepressive Agents, Second-Generation
  • 3401 Analytical chemistry