Diana Gazzola

Roles of proteins, polysaccharides, and phenolics in haze formation in white wine via reconstitution experiments

Residual proteins in finished wines can aggregate to form haze. To obtain insights into the mechanism of protein haze formation, a reconstitution approach was used to study the heat-induced aggregation behavior of purified wine proteins. A chitinase, four thaumatin-like protein (TLP) isoforms, phenolics, and polysaccharides were isolated from a Chardonnay wine. The same wine was stripped of these compounds and used as a base to reconstitute each of the proteins alone or in combination with the isolated phenolics and/or polysaccharides. After a heating and cooling cycle (70 °C for 1 h and 25 °C for 15 h), the size and concentration of the aggregates formed were measured by scanning ion occlusion sensing (SIOS), a technique to detect and quantify nanoparticles. The chitinase was the protein most prone to aggregate and the one that formed the largest particles; phenolics and polysaccharides did not have a significant impact on its aggregation behavior. TLP isoforms varied in susceptibility to haze formation and in interactions with polysaccharides and phenolics. The work establishes SIOS as a useful method for studying wine haze.

Study of combined effect of proteins and bentonite fining on the wine aroma loss.

The wine aroma loss as a consequence of treatments with bentonite is due to the occurrence of multiple interaction mechanisms. In addition to a direct effect of bentonite, the removal of aroma compounds bound to protein components adsorbed by the clay has been hypothesized but never demonstrated. We studied the effect of bentonite addition on total wine aroma compounds (extracted from Moscato wine) in a model solution in the absence and presence of total and purified (thaumatin-like proteins and chitinase) wine proteins. The results showed that in general bentonite alone has a low effect on the loss of terpenes but removed ethyl esters and fatty acids. The presence of wine proteins in the solution treated with bentonite tended to increase the loss of esters with the longest carbon chains (from ethyl octanoate to ethyl decanoate), and this was significant when the purified proteins were used. The results here reported suggest that hydrophobicity can be one of the driving forces involved in the interaction of aromas with both bentonite and proteins.

The proteins of the grape (Vitis vinifera L.) seed endosperm: Fractionation and identification of the major components

In the present study, grape (Vitis vinifera L.) seed endosperm proteins were characterized after sequential fractionation, according to a modified Osborne procedure. The salt-soluble fraction (albumins and globulins) comprised the majority (58.4%) of the total extracted protein. The protein fractions analysed by SDS-PAGE showed similar bands, indicating different solubility of the same protein components. SDS-PAGE in non-reducing and reducing conditions revealed the polypeptide composition of the protein bands. The main polypeptides, which were similar in all the grape varieties analysed, were identified by LC-MS/MS as homologous to the 11S globulin-like seed storage proteins of other plant species, while a monomeric 43 kDa protein presented high homology with the 7S globulins of legume seeds. The results provide new insights about the identity, structure and polypeptide composition of the grape seed storage proteins.

Characterization of a Grape Class IV Chitinase

A chitinase was purified from Vitis vinifera Manzoni Bianco grape juice and characterized. On the basis of proteomic analysis of tryptic peptides, a significant match identified the enzyme as a type IV grape chitinase previously found in juices of other V. vinifera varieties. The optimal pH and temperature for activity toward colloidal chitin were found to be 6 and 30 °C, respectively. The enzyme was found to hydrolyze chitin and oligomers of N-acetylglucosamine, generating N,N′-diacetylchitobiose and N-acetylglucosamine as products, but was inactive toward N,N′-diacetylchitobiose. The enzyme exhibited both endo- and exochitinase activities. Because yeast contains a small amount of chitin in the cell wall, the possibility of growth inhibition was tested. At a concentration and pH expected in ripe grapes, no inhibition of wine yeast growth by the chitinase was observed.

Spectroscopy reveals that ethyl esters interact with proteins in wine.

Impairment of wine aroma after vinification is frequently associated to bentonite treatments and this can be the result of protein removal, as recently demonstrated for ethyl esters. To evaluate the existence of an interaction between wine proteins and ethyl esters, the effects induced by these fermentative aroma compounds on the secondary structure and stability of VVTL1, a Thaumatin-like protein purified from wine, was analyzed by Synchrotron Radiation Circular Dichroism (SRCD) spectroscopy. The secondary structure of wine VVTL1 was not strongly affected by the presence of selected ethyl esters. In contrast, VVTL1 stability was slightly increased by the addition of ethyl-octanoate, -decanoate and -dodecanoate, but decreased by ethyl-hexanoate. This indicates the existence of an interaction between VVTL1 and at least some aroma compounds produced during fermentation. The data suggest that proteins removal from wine by bentonite can result in indirect removal of at least some aroma compounds associated with them.

Advantages of the KDS/BCA Assay over the Bradford Assay for Protein Quantification in White Wine and Grape Juice

The present study compared the performance of two colorimetric protein assays, the Bradford and the potassium dodecyl sulfate/bicinchoninic acid (KDS/BCA) assays, for use in wine and grape juice analysis. The Bradford assay was affected by protein type, whereas the KDS/BCA assay had lower protein-to-protein variation. Bovine serum albumin and lysozyme yielded an absorbance (562 nm) vs. protein concentration slope (dose–response curve) similar to that of wine proteins. In the Bradford assay, the presence of 12% ethanol and 200 mg/L of wine polyphenols decreased the protein absorbance by 28 and 16%, respectively, whereas in the KDS/BCA assay such interference was not significant. Among 64 white wines, the correlation between protein haze potential, determined by a heat test, and protein content was better for the KDS/BCA assay. This study confirmed the superiority of the KDS/BCA assay over the Bradford assay for quantifying protein in white grape juice and wine, and it yielded better predictive value with respect to the risk of white wine protein instability.

Grape seed extract: the first protein-based fining agent endogenous to grapes

Background and Aims There is a growing interest in finding alternative wine fining agents to replace potentially allergenic animal-derived and plant-derived proteins. In this context, the potential use of grape-derived fining agents would be beneficial as they would not introduce additional, potentially allergenic products to the finished wine. In this project, grape seed flour, a by-product of the grape oil seed industry, has been used to prepare a grape seed extract (GSE) for testing as a novel wine fining agent. Methods and Results The fining performance of GSE was compared with that of patatin, pea proteins, polyvinylpolypyrrolidone and potassium caseinate in a white and in a rose wine, and of ovalbumin and gelatin in a red wine. Reduction of turbidity, effect on wine colour, the concentration of phenolic substances, browning potential and wine sensory attributes were determined. Grape seed extract was effective in decreasing white wine turbidity when compared to potassium caseinate. In red wine, GSE removed some anthocyanin and proanthocyanins, while wine colour was only slightly affected. The greatest GSE effect was observed on the sensory properties of the treated wines, as it strongly reduced the vegetal notes in the rose wine and improved the overall taste of the red wine as a result of the reduction in both acidity and astringency. Conclusions Grape seed extract can be considered a valid allergen-free alternative to the most common wine fining agents. Significance of the Study Grape seed extract is the first effective fining agent endogenous to grapes, thus not attracting the legal restrictions concerning the presence of foreign substances.