17E-18

Protein-tannin interactions forming hazes in beer, wine and fruit juices limit shelf life and are greatly influenced by the amino acid composition of the protein and its conformation. Proline-rich proteins with an extended structure strongly interact with tannins, predominantly associating by hydrophobic stacking of the polyphenol ring against the pyrrolidine ring of proline. The proteins that most readily react to form hazes with polyphenols are referred to haze-active proteins. Heat treatment has often been used in the studies of haze-active proteins; however, this method is time-consuming and suffers from a low selectivity. A simple, fast and more selective approach is presented in this study and its feasibility as a method of identification of haze-active proteins is demonstrated. Proteins extracted from grape seeds were partially unfolded by treatment with 1% SDS and then interacted with different amounts of tannin at 4ºC, followed by gel electrophoresis. It was found that the intensity of the band at 45-kDa was decreased as tannins increased. The results from amino acid analysis of the isolated 45-kDa protein from grape seeds showed that it was higher in proline (9.46%) than the average proline content of total grape seed protein (5%). The selectivity of the developed approach was verified by proline-rich proteins (gelatin) and globular protein (bovine serum albumin) in a model system. As expected, the gelatin was removed as it reacted with the increasing added tannins, whereas BSA did not. The nonionic detergent, Triton X-100 used for protein folding, reduced the efficiency of locating haze-active proteins. Our results show that it is possible to identify haze-active proteins by modulating the accessibility of protein to tannins, and it is suggested that this new method be used by the beverage industry to design an immunoassay for the precise control of adsorbents to trouble-shoot haze problems and to quality control.