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Wine Protein Haze: Mechanisms of Formation and Advances in Prevention Steven C Van Sluyter, Jacqui M. McRae, Robert John Falconer, Paul A. Smith, Antony Bacic, Elizabeth J. Waters, and Matteo Marangon J. Agric. Food Chem., Just Accepted Manuscript • Publication Date (Web): 07 Apr 2015 Downloaded from http://pubs.acs.org on April 7, 2015
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Journal of Agricultural and Food Chemistry
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Wine Protein Haze: Mechanisms of Formation and Advances in Prevention
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Steven C. Van Sluytera,b,c*, Jacqui M. McRaea, Robert J. Falconerd, Paul A. Smitha, Antony
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Bacicb, Elizabeth J. Watersa,e, Matteo Marangon a,f*
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a
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Australia
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b
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University of Melbourne, Victoria 3010, Australia
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c
Department of Biological Sciences, Macquarie University, New South Wales 2109, Australia
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d
Department of Chemical & Biological Engineering, ChELSI Institute, University of Sheffield,
The Australian Wine Research Institute, P.O Box 197, Glen Osmond, South Australia 5064,
School of BioSciences and the Bio21 Molecular Sciences & Biotechnology Institute,
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Sheffield, S1 3JD, England
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e
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Australia
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f
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*Corresponding authors: E-mail:
[email protected]. Phone: +61 (0)2 9850 6316.
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Fax: +61 (0)2 9850 8245. E-mail:
[email protected]. Phone: +44 (0)1273
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890454. Fax: +44 (0) 1273 890071
Australian Grape and Wine Authority, P.O. Box 2733, Adelaide, South Australia 5000,
Plumpton College, Ditchling Road, Nr Lewes, BN7 3AE, England
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Abstract
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Protein haze is an aesthetic problem in white wines that can be prevented by removing
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grape proteins that have survived the winemaking process. The haze forming proteins are
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grape pathogenesis-related proteins that are highly stable during winemaking, but some of
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them precipitate over time and with elevated temperatures. Protein removal is currently
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achieved by bentonite addition, an inefficient process that can lead to higher costs and
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quality losses in winemaking. The development of more efficient processes for protein
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removal and haze prevention requires understanding mechanisms such as the main drivers
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of protein instability and the impacts of various wine matrix components on haze formation.
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This review covers recent developments in wine protein instability and removal and
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proposes a revised mechanism of protein haze formation.
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Keywords
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bentonite alternatives, chitinases, pathogenesis-related proteins, protease, protein
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aggregation, thaumatin-like protein, wine haze, wine heat instability, wine protein
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Introduction
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In 2012 there were 7.528 million hectares of cultivated grape vines among 92 countries,
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making grapes the largest fruit crop by land area in the world.1,2 Furthermore, much value is
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added in the form of winemaking to over half the world’s grapes, with the production of 252
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millions of hectolitres of wine in 2012.2 The contribution of the wine sector to the world
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economy in 2013 reached a value of $277.5 billion3 with a large proportion of the wine
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exported. Thus a substantial volume of wine is subject to potentially damaging conditions
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during transportation and storage, such as inappropriate temperature or humidity, that can
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cause deleterious modifications of the organoleptic features of the wine.4
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Wine clarity, especially that of white wines (Figure 1), is important to most consumers and is
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also one of the characteristics that is most easily affected by inappropriate shipping and
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storage conditions. For this reason, securing wine stability prior to bottling is an essential
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step of the winemaking process and presents a significant challenge for winemakers. A
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stable white wine is one that is clear and free from precipitates at the time of bottling,
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through transport and storage to the time of consumption. Hazy wine and the presence of
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precipitates are most commonly caused by three factors: microbial instability, tartrate
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instability, and protein heat instability.5 Microbial stability is achieved prior to bottling by
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sulfur dioxide addition and filtration;6 tartrate stability is achieved by either cold
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stabilization, ion exchange resins or electrodialysis.7
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Protein stability in commercial winemaking is almost always achieved by the addition of
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bentonite, a clay cation exchanger that binds proteins and removes them from wine
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through precipitation. Protein-bound bentonite settles loosely to the bottom of wine tanks
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as lees, which account for around 3-10% of the original wine volume.8 Wine is recovered
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from bentonite lees through processing using rotary drum vacuum filtration, specialized lees 3 ACS Paragon Plus Environment
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filtration equipment, or centrifugation - processes that are considered laborious and that
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can potentially degrade wine quality.8–10 Quality degradation and loss of wine through
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bentonite usage has been estimated to cost the global wine industry around US$ 1 billion
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per year.11 Other issues and costs related to bentonite use include tank downtime for
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bentonite treatment, occupational health risks associated with inhalation of bentonite dust
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and slip hazards induced by bentonite slurry spills, the disposal of hazardous bentonite
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waste, and bentonite interference with increasingly common membrane-based winemaking
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technologies.12 Consequently, winemakers aim to use the minimum amount of bentonite
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required for protein stability and would welcome the introduction of alternatives with fewer
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drawbacks than the current practice.
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Since the last extensive review on the topic a decade ago8, research efforts have been
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equally divided into the elucidation of protein haze-forming mechanisms, in particular the
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effects of different wine components, as well as improving bentonite efficiency and finding
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alternative stabilization strategies. Significant attention has also been paid to developing
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methods for protein purification, quantification and identification, as well as predicting wine
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haze potential (Figure 2).
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This review summarizes recent advances in the knowledge of how protein haze forms in
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wine, as well as the latest alternatives to bentonite wine protein stabilization. The findings
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of recent research and the newly-proposed mechanisms for haze will be discussed in Part I.
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New alternatives to bentonite will be discussed in Part II.
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Part I. Mechanisms of protein haze formation in white wines
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Current model of haze formation. The mechanisms associated with haze formation in wines
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are not well understood and yet is commonly cited as a two-stage process. In the first stage,
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wine proteins unfold in response to stimuli such as elevated storage temperatures. Once 4 ACS Paragon Plus Environment
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unfolded, the proteins aggregate and flocculate to form a visible haze.13 Recent
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investigations of the proteins associated with haze formation, as well as the roles of other
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wine components, have enabled the proposed model to be revised into three separate
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stages described below. The steps include protein unfolding, protein self-aggregation and
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aggregate cross-linking.
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Haze-forming proteins. The isolation and characterization of proteins from white wines has
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traditionally been a difficult task due to the presence of grape and yeast proteins as well as
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their modified versions and degradation products caused by winemaking, which produces a
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complex protein mixture.14,15 However recent advances in techniques for wine protein
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purification,16,17 as well as applications of newly-developed proteomic techniques16,18–25,
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and the release of the grape genome,26 have significantly improved research capabilities in
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the identification and quantification of grape and wine proteins.
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The most abundant classes of haze-forming proteins that occur in grape (Vitis vinifera) juice
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and white wines are chitinases and thaumatin-like proteins (TLPs).14,27–29 These proteins are
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small (
