Preparative separation, structural analysis and evaluation of dentin stiffening activity of grape seed oligomeric proanthocyanidins

A strong and stable dentin-collagen matrix is a prerequisite for building long lasting dental restorations. From previous studies, a 15-fold increase in the dentin stiffening activity was observed on treatment with crude grape seed extract (GSE). Dentin stiffness assay is performed on demineralized dentin beams using a Shimadzu EZ-graph universal testing machine that evaluates mechanical strength of the dentin 1hr post treatment with sample. A partition of the GSE enriched in PAC oligomers from n = 2 to n = 4 (GSE-MP) was produced using a 3 phase solvent system. GSE-MP was found to be more active than the crude extract. A novel centrifugal partition chromatography (CPC) method using 2 step elution and 1 step extrusion has been developed to separate the complex mixture of grape seed oligomeric PACs. GSE-MP also contains galloylated oligomeric PACS (GOPACs). We have previously observed that galloylation increases the dentin stiffness activity in case of monomeric PACs. Thus isolation of GOPACs from GSE-MP using CPC followed by preparative reverse phase HPLC (RP-HPLC) is targeted in order to perform structural analyses as well as dentin bio-modification studies.

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