Improvement of BSA Tannin Precipitation Assay by Reformulation of Resuspension Buffer

Abstract

Analysis of protein precipitable wine tannins has become more commonplace due to the simplicity of the method and the strong association between tannin content and perception of astringency. The traditional protein precipitation method for wine measures tannins and polymeric pigments using 5% w/v sodium dodecyl sulfate (SDS) and the alkaline buffer triethanolamine (TEA) pH 9.4 to dissolve the tannin-protein precipitate and support the colorimetric reaction with ferric chloride. However, this method results in a high background absorbance presumably due to the alkaline pH of the resuspension buffer, which has been shown to oxidize phenolics. Experimentation with several buffer formulations showed that pairing TEA with urea instead of SDS allowed formulation of buffers at lower pH. Urea-TEA buffers at pH 7 and 8 showed significantly lower background absorbance and drift over time as well as a significantly greater amount of tannin recovered. These trends in tannin content and iron reactive phenolics were maintained across a wide range of diluted samples (100–1200 mg/L catechin equivalents). Using a TEA-urea buffer at pH 7 or 8 may improve on previous methods of protein tannin precipitation by increasing yield of tannins from the wine and decreasing the background absorbance and drift.