High-performance liquid chromatography-based determination of total isothiocyanate levels in human plasma: Application to studies with 2-phenethyl isothiocyanate

Leonard Liebes, C. Clifford Conaway, Howard Hochster, Sandra Mendoza, Stephen S. Hecht, James Crowell, Fung Lung Chung

Research output: Contribution to journalArticlepeer-review

78 Scopus citations

Abstract

Dietary and pharmacologic isothiocyanates (ITCs) may play a role in reducing the risk of certain cancers. The quantification of ITCs in humans is important both for epidemiological and pharmacokinetic studies. We describe a modification of an HPLC-based assay of urinary ITCs for use with human plasma. The assay utilizes the cyclocondensation reaction of 1,2-benzenedithiol with ITCs present in human plasma, followed by a twostep hexane extraction and analysis by HPLC using UV detection at 365 nm. The method shows linearity and reproducibility with human plasma over a range of 49-3003 nM phenethyl isothiocyanate (PEITC) (r2 = 0.996 ± 0.003). A similar degree of linearity was seen with two other biologically occurring conjugates of PEITC: PEITC-N-acetylcysteine (PEITC-NAC) and PEITC-glutathione (PEITC-GSH). The recovery of PEITC assessed on multiple days was 96.6 ± 1.5% and was 100% for PEITC-GSH and PEITC-NAC. The reproducibility of the assay on multiday samplings showed a mean %CV of 6.5 ± 0.3% for PEITC, 6.4 ± 4.3 for PEITC-NAC and 12.3 ± 3.9 for PEITC-GSH. In clinical studies, mean plasma ITC level of 413 ± 193 nM PEITC equivalents was determined for a non-dietary-controlled group of 23 subjects. Multiday analysis data from pharmacokinetic plasma sets of 3 subjects taking a single dose of PEITC at 40 mg showed a good CV (range: 16-21%). The applicability of the methodology to pharmacokinetic studies of PEITC in humans is demonstrated.

Original languageEnglish (US)
Pages (from-to)279-289
Number of pages11
JournalAnalytical Biochemistry
Volume291
Issue number2
DOIs
StatePublished - Apr 15 2001

Bibliographical note

Funding Information:
1 Work was supported by NCI-CN-55120, CA-166081-21, CRC-RR-00096, and CA 46535.

Keywords

  • HPLC
  • Isothiocyanates
  • PEITC
  • PEITC-GSH
  • PEITC-NAC
  • Pharmacokinetics
  • Phenethyl isothiocyanate
  • Plasma assay

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