Facile Purification of Milligram to Gram Quantities of Condensed Tannins According to Mean Degree of Polymerization and Flavan-3-ol Subunit Composition

Authors

• Ron H. Brown, University of Reading
• Irene Murray-Harvey, University of ReadingFollow
• Wayne E. Zeller, USDA-ARSFollow
• Laurie Reinhardt, USDA-ARSFollow
• Elisabetta Stringano, University of Reading
• An Gea, University of Reading
• Christopher Drake, University of Reading
• Honorata M. Ropiak, University of Reading
• Christos Fryganas, University of Reading
• Aina Ramsay, University of Reading
• Emily E. Hardcastle, USDA-ARS

Document Type

Article

Date of this Version

2017

Citation

J. Agric. Food Chem. 2017, 65, pp. 8072−8082, DOI: 10.1021/acs.jafc.7b03489.

Comments

U.S. government work.

Abstract

Unambiguous investigation of condensed tannin (CT) structure−activity relationships in biological systems requires well-characterized, high-purity CTs. Sephadex LH-20 and Toyopearl HW-50F resins were compared for separating CTs from acetone/water extracts, and column fractions analyzed for flavan-3-ol subunits, mean degree of polymerization (mDP), and purity. Toyopearl HW-50F generated fractions with higher mDP values and better separation of procyanidins (PC) and prodelphinidins (PD) but required a prepurification step, needed more time for large scale purifications, and gave poorer recoveries. Therefore, two gradient elution schemes were developed for CT purification on Sephadex LH-20 providing 146−2000 mg/fraction. Fractions were analyzed by thiolysis and NMR spectroscopy. In general, PC/PD ratios decreased and mDP increased during elution. 1H NMR spectroscopy served as a rapid screening tool to qualitatively determine CT enrichment and carbohydrate impurities present, guiding fractionation toward repurification or ¹H−¹³C HSQC NMR spectroscopy and thiolysis. These protocols provide options for preparing highly pure CT samples.