Protein haze is an esthetic problem in white wines that can be prevented by removing grape proteins that have 13 survived the winemaking process. The haze-forming proteins are grape pathogenesis-related proteins that are highly stable during 14 winemaking, but some of them precipitate over time and with elevated temperatures. Protein removal is currently achieved by 15 bentonite addition, an inefficient process that can lead to higher costs and quality losses in winemaking. The development of 16 more efficient processes for protein removal and haze prevention requires understanding the mechanisms that are the main 17 drivers of protein instability and the impacts of various wine matrix components on haze formation. This review covers recent 18 developments in wine protein instability and removal and proposes a revised mechanism of protein haze formation. 24 by land area in the world. 1,2 Furthermore, much value is added 25 in the form of winemaking to over half the world's grapes, with 26 the production of 252 million hectoliters of wine in 2012. 2 The 27 contribution of the wine sector to the world economy in 2013 28 reached a value of U.S.$277.5 billion, 3 with a large proportion 29 of the wine exported. Thus, a substantial volume of wine is 30 subject to potentially damaging conditions during trans-31 portation and storage, such as inappropriate temperature or 32 humidity, that can cause deleterious modifications of the 33 organoleptic features of the wine. 4 f1 34Wine clarity, especially that of white wines (Figure 1), is 35 important to most consumers and is also one of the 36 characteristics that is most easily affected by inappropriate 37 shipping and storage conditions. For this reason, securing wine 38 stability prior to bottling is an essential step of the winemaking 39 process and presents a significant challenge for winemakers. A 40 stable white wine is one that is clear and free from precipitates 41 at the time of bottling, through transport and storage, to the 42 time of consumption. Hazy wine and the presence of 43 precipitates are most commonly caused by three factors: 44 microbial instability, tartrate instability, and protein heat 45 instability. 5 Microbial stability is achieved prior to bottling by 46 sulfur dioxide addition and filtration; 6 tartrate stability is 47 achieved by either cold stabilization, ion exchange resins, or 48 emr00 | ACSJCA | JCA10.0.1465/W Unicode | research.3f (R3.6.i7:4236 | 2.0 alpha 39) 2014/12/19 13:33:00 | PROD-JCAVA | rq_4578338 | 4/10/2015 14:19:11 | 11 | JCA-DEFAULT 55 lees through processing using rotary drum vacuum filtration, 56 specialized lees filtration equipment, or centrifugation 57 processes that are considered laborious and that can potentially 58 degrade wine quality. 8−10 Quality degradation and loss of wine 59 through bentonite usage has been estimated to cost the global 60 wine industry around U.S.$1 billion per year. 11 Other issues 61 and costs related to bentonite use include tank downtime for 62 bentonite treatment, ...

A thermal unfolding study of thaumatin-like protein, chitinase, and invertase isolated from Vitis vinifera Sauvignon blanc and Semillon juice was undertaken. Differential scanning calorimetry demonstrated that chitinase was a major player in heat-induced haze in unfined wines as it had a low melt temperature, and aggregation was observed. The kinetics of chitinase F1 (Sauvignon blanc) unfolding was studied using circular dichroism spectrometry. Chitinase unfolding conforms to Arrhenius behavior having an activation energy of 320 kJ/mol. This enabled a predictive model for protein stability to be generated, predicting a half-life of 9 years at 15 degrees C, 4.7 days at 30 degrees C, and 17 min at 45 degrees C. Circular dichroism studies indicate that chitinase unfolding follows three steps: an initial irreversible step from the native to an unfolded conformation, a reversible step between a collapsed and an unfolded non-native conformation, followed by irreversible aggregation associated with visible haze formation.

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Grape chitinase was found to be the primary cause of heat-induced haze formation in white wines. Chitinase was the dominant protein in a haze induced by treating Sauvignon blanc wine at 30 °C for 22 h. In artificial wines and real wines, chitinase concentration was directly correlated to the turbidity of heat-induced haze formation (50 °C for 3 h). Sulfate was confirmed to have a role in haze formation, likely by converting soluble aggregates into larger visible haze particles. Thaumatin-like protein was detected in the insoluble fraction by SDS-PAGE analysis but had no measurable impact on turbidity. Differential scanning calorimetry demonstrated that the complex mixture of molecules in wine plays a role in thermal instability of wine proteins and contributes additional complexity to the wine haze phenomenon.