Development and validation of an ultra-high performance liquid chromatography/triple quadrupole mass spectrometry method for analyzing microbial-derived grape polyphenol metabolites

Danyue Zhao, Bo Yuan, Eileen Carry, Giulio M. Pasinetti, Lap Ho, Jeremiah Faith, Ilaria Mogno, Jim Simon, Qingli Wu

Research output: Journal article publicationJournal articleAcademic researchpeer-review

8 Citations (Scopus)

Abstract

Accumulating evidence indicates that the health impact of dietary phenolic compounds, including the principal grape-derived polyphenols, (+)‑catechin and (−)‑epicatechin, is exerted by not only the parent compounds but also their phenolic metabolites generated by the gut microbiota. In this work, a new high-throughput, sensitive and reproducible analytical method was developed employing ultra-high performance liquid chromatography coupled with triple quadrupole tandem mass spectrometry (UHPLC-QqQ-MS/MS) for the simultaneous analysis of 16 microbial-generated phenolic acid metabolites (PAMs) along with their precursors, catechin and epicatechin. Following optimizing the solvent system, LC conditions and MS parameters, method validation was carried out to evaluate the sensitivity, selectivity, accuracy and precision of the proposed method, and to ensure promising recovery of all analytes extracted from the matrix prior to bioanalysis. Results showed that the optimized analytical method allowed successful confirmation and quantitation of all analytes under dynamic multiple reaction monitoring mode using trans‑cinnamic acid‑d7 as an internal standard (I.S.). Excellent sensitivity and linearity were obtained for all analytes, with lower limits of detection (LLODs) and lower limits of quantification (LLOQs) in the ranges of 0.225–2.053 ng/mL and 0.698–8.116 ng/mL, respectively. By examining blank matrix spiked with standard mixture at different concentration levels, promising recoveries at two spiking levels (low level, 91.2–115%; high level 90.2–121%), and excellent precision (RSD < 10%) were obtained. This method was then successfully applied to an in vitro study where catechin/epicatechin-enriched broth samples were anaerobically fermented with gut microbes procured from healthy human donors. All sources of bacteria employed showed remarkable activity in metabolizing grape polyphenols and distinct variations in the production of PAMs. The successful application of this method in the in vitro fermentation assays demonstrates its suitability for high-throughput analysis of polyphenol metabolites, particularly catechin/epicatechin-derived PAMs, in biological studies.

Original languageEnglish
Pages (from-to)34-45
Number of pages12
JournalJournal of Chromatography B: Analytical Technologies in the Biomedical and Life Sciences
Volume1099
DOIs
Publication statusPublished - 1 Nov 2018
Externally publishedYes

Keywords

  • Grape polyphenols
  • Gut microbiota
  • In vitro fermentation
  • Method validation
  • Phenolic acid metabolites
  • UHPLC-QqQ-MS/MS

ASJC Scopus subject areas

  • Analytical Chemistry
  • Biochemistry
  • Clinical Biochemistry
  • Cell Biology

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