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

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.

Authors+Show Affiliations

New Use Agriculture & Natural Plant Products Program, Department of Plant Biology, School of Environmental and Biological Sciences, Rutgers University, New Brunswick, NJ 08901, USA.New Use Agriculture & Natural Plant Products Program, Department of Plant Biology, School of Environmental and Biological Sciences, Rutgers University, New Brunswick, NJ 08901, USA.New Use Agriculture & Natural Plant Products Program, Department of Plant Biology, School of Environmental and Biological Sciences, Rutgers University, New Brunswick, NJ 08901, USA; Department of Medicinal Chemistry, Ernst Mario School of Pharmacy, Rutgers University, Piscataway, NJ, USA.Department of Neurology, Icahn School of Medicine at Mount Sinai, New York, NY 10029, USA; Geriatric Research, Education & Clinical Center, James J. Peters Veterans Affairs Medical Center, Bronx, NY 10468, USA.Department of Neurology, Icahn School of Medicine at Mount Sinai, New York, NY 10029, USA; Geriatric Research, Education & Clinical Center, James J. Peters Veterans Affairs Medical Center, Bronx, NY 10468, USA.Department of Genetics and Genomic Sciences, Icahn School of Medicine at Mount Sinai, New York, NY 10029, USA.Department of Genetics and Genomic Sciences, Icahn School of Medicine at Mount Sinai, New York, NY 10029, USA.New Use Agriculture & Natural Plant Products Program, Department of Plant Biology, School of Environmental and Biological Sciences, Rutgers University, New Brunswick, NJ 08901, USA; Department of Medicinal Chemistry, Ernst Mario School of Pharmacy, Rutgers University, Piscataway, NJ, USA.New Use Agriculture & Natural Plant Products Program, Department of Plant Biology, School of Environmental and Biological Sciences, Rutgers University, New Brunswick, NJ 08901, USA; Department of Medicinal Chemistry, Ernst Mario School of Pharmacy, Rutgers University, Piscataway, NJ, USA. Electronic address: qlwu@sebs.rutgers.edu.

Pub Type(s)

Journal Article

Language

eng

PubMed ID

30241072

Citation

Zhao, Danyue, et al. "Development and Validation of an Ultra-high Performance Liquid Chromatography/triple Quadrupole Mass Spectrometry Method for Analyzing Microbial-derived Grape Polyphenol Metabolites." Journal of Chromatography. B, Analytical Technologies in the Biomedical and Life Sciences, vol. 1099, 2018, pp. 34-45.
Zhao D, Yuan B, Carry E, et al. Development and validation of an ultra-high performance liquid chromatography/triple quadrupole mass spectrometry method for analyzing microbial-derived grape polyphenol metabolites. J Chromatogr B Analyt Technol Biomed Life Sci. 2018;1099:34-45.
Zhao, D., Yuan, B., Carry, E., Pasinetti, G. M., Ho, L., Faith, J., Mogno, I., Simon, J., & Wu, Q. (2018). Development and validation of an ultra-high performance liquid chromatography/triple quadrupole mass spectrometry method for analyzing microbial-derived grape polyphenol metabolites. Journal of Chromatography. B, Analytical Technologies in the Biomedical and Life Sciences, 1099, 34-45. https://doi.org/10.1016/j.jchromb.2018.09.014
Zhao D, et al. Development and Validation of an Ultra-high Performance Liquid Chromatography/triple Quadrupole Mass Spectrometry Method for Analyzing Microbial-derived Grape Polyphenol Metabolites. J Chromatogr B Analyt Technol Biomed Life Sci. 2018 Nov 1;1099:34-45. PubMed PMID: 30241072.
* Article titles in AMA citation format should be in sentence-case
TY - JOUR T1 - Development and validation of an ultra-high performance liquid chromatography/triple quadrupole mass spectrometry method for analyzing microbial-derived grape polyphenol metabolites. AU - Zhao,Danyue, AU - Yuan,Bo, AU - Carry,Eileen, AU - Pasinetti,Giulio M, AU - Ho,Lap, AU - Faith,Jeremiah, AU - Mogno,Ilaria, AU - Simon,Jim, AU - Wu,Qingli, Y1 - 2018/09/12/ PY - 2018/03/04/received PY - 2018/09/05/revised PY - 2018/09/11/accepted PY - 2018/9/22/pubmed PY - 2018/12/12/medline PY - 2018/9/22/entrez KW - Grape polyphenols KW - Gut microbiota KW - In vitro fermentation KW - Method validation KW - Phenolic acid metabolites KW - UHPLC-QqQ-MS/MS SP - 34 EP - 45 JF - Journal of chromatography. B, Analytical technologies in the biomedical and life sciences JO - J Chromatogr B Analyt Technol Biomed Life Sci VL - 1099 N2 - 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. SN - 1873-376X UR - https://www.unboundmedicine.com/medline/citation/30241072/Development_and_validation_of_an_ultra_high_performance_liquid_chromatography/triple_quadrupole_mass_spectrometry_method_for_analyzing_microbial_derived_grape_polyphenol_metabolites_ L2 - https://linkinghub.elsevier.com/retrieve/pii/S1570-0232(18)30377-5 DB - PRIME DP - Unbound Medicine ER -