Slow denaturation of wine proteins is thought to lead to protein aggregation, flocculation into a hazy suspension and formation of precipitates. The majority of wine proteins responsible for haze are grape‐derived, have low isoelectric points and molecular weight. They are grape pathogenesis‐related (PR) proteins that are expressed throughout the ripening period post véraison, and are highly resistant to low pH and enzymatic or non‐enzymatic proteolysis. Protein levels in un‐fined white wine differ by variety and range up to 300 mg/L. Infection with some common grapevine pathogens or skin contact, such as occurs during transport of mechanically harvested fruit, results in enhanced concentrations of some PR proteins in juice and wine. Oenological control of protein instability is achieved through adsorption of wine proteins onto bentonite. The adsorption of proteins onto bentonite occurs within several minutes, suggesting that a continuous contacting process could be developed. The addition of proteolytic enzyme during short term heat exposure, to induce PR protein denaturation, showed promise as an alternative to bentonite fining. The addition of haze‐protective factors, yeast mannoproteins, to wines results in decreased particle size of haze, probably by competition with wine proteins for other non‐proteinaceous wine components required for the formation of large insoluble aggregations of protein. Other wine components likely to influence haze formation are ethanol concentration, pH, metal ions and phenolic compounds.

Protein haze formation in white wine is dependent on the presence of both wine protein and other unknown wine components, termed factor(s) X. The ability to reconstitute protein haze upon heating artificial model wine solutions (500 mg/L thaumatin, 12% ethanol, 4 g/L tartaric acid) to which candidate components were added was employed to identify factor(s) X. No protein haze was formed in the absence of additives. The individual or combined addition of caffeic acid, caftaric acid, epicatechin, epigallocatechin-O-gallate, gallic acid, or ferulic acid at typical white wine concentrations did not generate protein haze. However, PVPP fining of commercial wines resulted in a reduction in protein haze, suggesting that phenolic compounds may play a modulating role in haze formation. To elucidate the nature of the unknown factor(s) wine was fractionated and fractions were back-added to model wine and tested for their essentiality. Wine fractions were generated by ultrafiltration, reverse-phase chromatography, and mixed-mode anion-exchange and reverse-phase chromatography. The only purified fraction containing the essential component(s) was free of phenolic compounds, and analysis by mass spectrometry identified sulfate anion as the dominant component. Reconstitution with KHSO4 using either commercially available thaumatin or wine proteins confirmed the role of sulfate in wine protein haze formation. The two main wine proteins, thaumatin-like protein and chitinase, differed in their haze response in model wines containing sulfate. Other common wine anions, acetate, chloride, citrate, phosphate, and tartrate, and wine cations, Fe(2+/3+) and Cu(+/2+), when added at typical white wine concentrations were not found to be essential for protein haze formation.

Thaumatin-like proteins and chitinases, which are pathogenesis-related (PR) proteins, were the major soluble protein components of grapes from five cultivars of Vitis vinifera. This dominance of PR proteins was apparent at berry softening (véraison) and then throughout berry development for the Muscat of Alexandria, Sultana, and Shiraz cultivars and in the berries of the Sauvignon Blanc and Pinot Noir cultivars examined at commercial maturity. The M(r) of the major thaumatin-like protein from Muscat of Alexandria grapes was 21 272, and those of the three major chitinases from this cultivar, ChitB, ChitC, and ChitD, were 25 588, 25 410, and 25 457, respectively. The vines in the study were irrigated and showed no obvious signs of disease. Shiraz vines that had not been irrigated throughout the season were clearly water stressed, but had levels of PR proteins in the berry similar to vines that had been fully irrigated. It appears that the production of PR proteins that cause protein instability in wines by grapes may be little influenced by environmental conditions.

Based on work undertaken over 30 years ago, most Australian winemakers use a heat test (80ºC for six hours) to check white wines for protein stability. Some winemakers consider that this test is too severe, resulting in wines being over-fined with bentonite. This paper presents the results of a study comparing the predictive ability of this test, assessed either by eye, by nephelometry or by optical density, with that of two alternative assays (80ºC for 2 hours and the Bentotest), through the use of storage trials of 8 wines designed to mimic commercial conditions of transport and storage of wine. One of the 8 unfined wines failed to develop haze even under the more severe storage conditions of this study (35ºC for a month or fluctuating between 20ºC and 35ºC for 8 days) and only 6 of the 8 unfined wines developed haze under 'best practice' storage conditions (13ºC-17ºC for 16 months). All the wines fined with bentonite at dosage rates determined by the three predictive tests used in the main part of this work remained bright in bottle after storage trials at 13ºC-17ºC for 16 months, 35ºC for 1 month or fluctuating between 20ºC and 35ºC for 8 days. Therefore the least severe stability test, heating at 80ºC for 2 hours, which generally indicated lower dosage rates in this study, accurately predicted short to medium term stability for these wines under these storage conditions. Abbreviations au spectrophotometric absorbance units; HRWSL Hickinbotham Roseworthy Wine Science Laboratory;NIR near infrared spectroscopy; NTU nephelometric turbidity units; PR pathogenesis related; ROTE roll on tamper evident screw cap; UV-VIS ultraviolet-visible