Identification and analysis of a mercapturic acid conjugate of indole-3-methyl isothiocyanate in the urine of humans who consumed cruciferous vegetables

Pramod Upadhyaya, Adam T. Zarth, Naomi Fujioka, Vincent A. Fritz, Stephen S. Hecht

Research output: Contribution to journalArticle

Abstract

Glucobrassicin, a quantitatively significant constituent of Brassica vegetables, gives rise to indole-3-carbinol (I3C) and its dimer di-indolylmethane (DIM) when the vegetables are chewed. I3C and DIM have been extensively studied with respect to their anti-carcinogenic properties. However, the presumed intermediate isothiocyanate in their formation, indole-3-methyl isothiocyanate (IMITC), has to our knowledge never been observed, despite the fact that isothiocyanates derived from cruciferous vegetables are known to have anti-carcinogenic properties. Therefore, we investigated the formation and presence in human urine of IMITC by analyzing for its N-acetylcysteine conjugate, IMITC-NAC, in order to gain a more complete understanding of the biochemical pathways leading to formation of I3C and DIM upon consumption of vegetables rich in glucobrassicin. Standard IMITC-NAC was synthesized and its structure confirmed by NMR and MS. IMITC-NAC was identified in extracts of Brussels sprouts chopped in the presence of N-acetylcysteine. An LC-ESI–MS/MS-SRM method for analysis of IMITC-NAC, with [13C,15N]IMITC-NAC as internal standard, was developed and validated. Then, ten subjects (7 females) consumed a salad of Brussels sprouts and cabbage (containing 100–500 μmol glucobrassicin) once daily for 3 days. Urine was collected at intervals up to 24 h after vegetable consumption. Levels of IMITC-NAC in the urine of these 10 subjects ranged from 0.2 to 30.2 pmol/mL urine. These results provide the first evidence for the presumed intermediacy of IMITC in the formation of I3C and DIM in humans who consumed Brussels sprouts and cabbage as a source of glucobrassicin.

Original languageEnglish (US)
Pages (from-to)341-346
Number of pages6
JournalJournal of Chromatography B: Analytical Technologies in the Biomedical and Life Sciences
Volume1072
DOIs
StatePublished - Jan 1 2018

Keywords

  • Cruciferous vegetables
  • Glucobrassicin
  • Indole-3-carbinol
  • Indole-3-methyl isothiocayanate

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