A validated HPLC-FLD method for analysis of intestinal absorption and metabolism of capsaicin and dihydrocapsaicin in the rat

Mónika Kuzma, Krisztina Fodor, Gábor Maász, Péter Avar, Gyula Mózsik, Tibor Past, Emil Fischer, Pál Perjési

Research output: Contribution to journalArticle

13 Citations (Scopus)

Abstract

A sensitive and selective reverse-phase high performance liquid chromatographic method with fluorescence detection has been developed for determination of capsaicin (8-methyl-N-vanillyl-(trans)-6-nonenamid) and dihydrocapsaicin (8-methyl-N-vanillylnonanamide) in samples generated in rat small intestine luminal perfusion experiments. The experiments were designed to study the biotransformation of capsaicinoids in the small intestine in the rat. The chromatographic separation was performed at room temperature on a ZORBAX Eclipse® XDB-C8 column using isocratic elution with a mobile phase consisting 0.05M orthophosphoric acid solution and acetonitrile (60:40, v/v; pH 3.0) with a flow rate of 1.5mL/min. Fluorescence detection was performed at excitation and emission wavelengths of 230 and 323nm, respectively. The method was evaluated for a number of validation characteristics (accuracy, repeatability and intermediate precision, limit of detection, limit of quantification and calibration range). The limit of detection (LOD) was 50ng/mL and the limit of quantification (LOQ) was 100ng/mL for both capsaicin and dihydrocapsaicin reference standards dissolved in blank perfusate. The method was successfully applied for investigation of intestinal absorption of capsaicin and dihydrocapsaicin while 30μg/mL standardized Capsicum extract - containing capsaicin and dihydrocapsaicin - was luminally perfused for a 90min period. The structure of the glucuronide metabolites of capsaicin and dihydrocapsaicin appeared in the perfusate was identified by mass spectrometry.

Original languageEnglish
Pages (from-to)59-66
Number of pages8
JournalJournal of Pharmaceutical and Biomedical Analysis
Volume103
DOIs
Publication statusPublished - Jan 5 2015

Keywords

  • Capsaicin and dihydrocapsaicin
  • Fluorescent detection
  • Glucuronide metabolites
  • High performance liquid chromatography
  • Intestinal metabolism

ASJC Scopus subject areas

  • Analytical Chemistry
  • Pharmaceutical Science
  • Drug Discovery
  • Spectroscopy
  • Clinical Biochemistry

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