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Epicatechin, procyanidins, and phenolic microbial metabolites after cocoa intake in humans and rats

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Abstract

Proanthocyanidins, flavonoids exhibiting cardiovascular protection, constitute a major fraction of the flavonoid ingested in the human diet. Although they are poorly absorbed, they are metabolized by the intestinal microbiota into various phenolic acids. An analytical method, based on an optimized 96-well plate solid-phase extraction (SPE) procedure and liquid chromatography tandem mass spectrometry (SPE-LC-MS/MS) for the analysis of 19 phenolic microbial metabolites and monomeric and dimeric flavanols in urine samples, was developed and validated. Human urine samples were obtained before and after ingestion of an acute consumption of 40 g of soluble cocoa powder and rat urines before and after the prolonged administration (2 weeks) of different diets composed of natural cocoa powder. The mean recovery of analytes using the new SPE-LC-MS/MS method ranged from 87% to 109%. Accuracy ranged from 87.5% to 113.8%, and precision met acceptance criteria (<15% relative standard deviation). Procyanidin B2 has been detected and quantified for the first time in human and rat urine after cocoa consumption. Changes in human and rat urinary levels of microbial phenolic acids and flavanols were in the range of 0.001–59.43 nmol/mg creatinine and of 0.004–181.56 nmol/mg creatinine, respectively. Major advantages of the method developed include reduction of laboratory work in the sample preparation step by the use of 96-well SPE plates and the sensitive measurement of a large number of metabolites in a very short run time, which makes it ideal for use in epidemiological studies.

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Acknowledgments

This research was supported by national grants, CICYT’s (AGL: 2004-08378-C02-01/02, 2005-002823, and 2006-14228-C03-02) and Grupo Consolider-Ingenio 2010 Fun-C-Food (CSD2007-063) and CIBER 06/03 Fisiopatologia de la Obesidad y la Nutrición, Instituto de Salud Carlos III, Spain also contributed. MU-S and NK thank FPI and FPU fellowship programs, respectively, and MM of the Juan de la Cierva post-doctoral program, all from the MEC. We are also grateful to the President of Lactalis Group for his support. We also thank the technical support and recommendations of Isidre Casals and Olga Jauregui from the Scientific and Technical Services of University of Barcelona and Marta Burrull from Waters Company.

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Authors and Affiliations

  1. Nutrition and Food Science Department, XaRTA. INSA. Faculty of Pharmacy, University of Barcelona, 08028, Barcelona, Spain

    Mireia Urpi-Sarda, Nasiruddin Khan, Rosa M. Lamuela-Raventos, Joan Permanyer & Cristina Andres-Lacueva

  2. Department of Internal Medicine, Hospital Clínic. Institut d’Investigació Biomèdica August Pi i Sunyer (IDIBAPS), University of Barcelona, 08036, Barcelona, Spain

    Maria Monagas & Emilio Sacanella

  3. Laboratory of Nutrition and Bromatology, School of Pharmacy, University of Salamanca, 37007, Salamanca, Spain

    Celestino Santos-Buelga

  4. Department of Physiology, Faculty of Pharmacy, University of Barcelona, Av. Joan XXIII s/n, 08028, Barcelona, Spain

    Margarida Castell

Authors
  1. Mireia Urpi-Sarda
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  2. Maria Monagas
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  3. Nasiruddin Khan
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  4. Rosa M. Lamuela-Raventos
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  5. Celestino Santos-Buelga
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  6. Emilio Sacanella
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  7. Margarida Castell
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  8. Joan Permanyer
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  9. Cristina Andres-Lacueva
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Corresponding author

Correspondence to Cristina Andres-Lacueva.

Additional information

a INGENIO-CONSOLIDER Program, Fun-c-food CSD2007-063

b CIBER 06003 Physiopathology of Obesity and Nutrition (CIBEROBN), and RETICS RD06/0045/0003, Instituto de Salud Carlos III

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Urpi-Sarda, M., Monagas, M., Khan, N. et al. Epicatechin, procyanidins, and phenolic microbial metabolites after cocoa intake in humans and rats. Anal Bioanal Chem 394, 1545–1556 (2009). https://doi.org/10.1007/s00216-009-2676-1

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  • DOI: https://doi.org/10.1007/s00216-009-2676-1

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