Fractionation of cocoa procyanidins according to the degree of polymerization by Centrifugal Partition Chromatography

Procyanidins are present in a wide variety of foods such as cocoa, apples, and many berries. There is a growing interest in determining their pharmacological properties and significance as dietary antioxidants. We have designed a novel approach to the preparative fractionation of cocoa procyanidins according to degree of polymerization (DP) by applying Centrifugal Partition Chromatography (FCPC Kromaton). In an experiment optimized for the best separation of DP families between 5 and 10, we employed a bi-phasic solvent system, consisting of ethyl acetate – ethanol – water (6:1:5) in ascending-mode. The procyanidin oligomers (DP2 to DP10) eluted with the mobile phase in an order of increasing DP. The separation of bands between DP5 and DP10 was impressive in a context of the best preparative approaches reported so far. The fractionation was monitored by a NP HPLC analysis on a polyvinyl alcohol (PVA) column and by a C18 RP column and by MALDI-TOF. The RP HPLC analysis revealed that the dominant peaks in each DP band were the linear epicatechin (4β-8) oligomers.The FCPC fractionation appears as an efficient approach to produce higher procyanidin oligomers in gram quantities required for evaluation of their biological properties.