220 E.J. Waters and C.B. Colby
between 9 vol.% and 13 vol.% ethanol may be enough to affect the tertiary structure
and/or aggregation of proteins.
Whilst sulfate appears to be fundamental to haze formation, other wine compo-
nents such as phenolic compounds remain as candidate haze modulators. One pos-
sibility is that white wine phenolic compounds affect the particle size of denatured
aggregated proteins, possibly through crosslinking. Several researchers (Oh et al.
1980; Siebert et al. 1996b) have suggested a hydrophobic mechanism for the inter-
action between phenolic compounds and proteins, in which the protein has a fixed
number of phenolic binding sites. More of these sites are exposed when the protein
is denatured.
Protein haze in white wine thus differs in several aspects from protein haze in
beer. It is well established that beer protein haze is due to interactions between
proteins, derived from the barley storage protein hordein and rich in proline, and
hop polyphenolic compounds (Bamforth 1999; Miedl et al. 2005; Siebert 1999;
Siebert and Lynn 2003). White wine proteins are not derived from storage proteins
of grape seed nor are they as rich in proline as hordein. In addition, wine protein haze
formation cannot be eliminated by removing polyphenolic compounds by PVPP
(Pocock et al. 2006) while in beer this has been applied as a commercial strategy
(Leiper et al. 2005; Madigan et al. 2000).
6C.6 Bentonite Fining
The major winemaking process to affect the levels of proteins in wine is bentonite
fining. Bentonite, a montmorillonite clay, is used almost universally throughout the
wine industry for the prevention of wine protein haze through removal of proteins
before bottling (Blade and Boulton 1988; Ferreira et al. 2002; Høj et al. 2000).
The adsorption of wine proteins onto bentonite is principally attributed to cationic
exchange with the bentonite clay. Wine proteins are positively charged at wine pH,
and thus can be exchanged onto bentonite, which carries a net negative charge
(Blade and Boulton 1988; Ferreira et al. 2002; Høj et al. 2000).
A number of studies have indicated that different protein fractions require dis-
tinct bentonite concentrations for protein removal and consequent heat stabilisation
(Duncan 1992; Ferreira et al. 2002). Bentonite fining has been shown to remove
higher pI (5.8–8.0) and intermediate molecular weight (MW; 32–45 kDa) proteins
first (Hsu and Heatherbell 1987b). However, these represent only a small portion
of the soluble proteins. Proteins with a MW of 60–65 kDa, and with wide pI
range (4.1–8.0) were highly resistant to removal by bentonite fining (i.e. required
significant bentonite addition) and typically remained in protein-stabilised wine
(Hsu and Heatherbell 1987b). Hsu and Heatherbell (1987b) concluded that it is nec-
essary to remove lower pI (4.1–5.8), lower MW (12.6 kDa and 20–30 kDa) proteins,
which represent a major component of proteins present, to protein stabilise wines.
Contrary to these findings, a study by Dawes et al. (1994) found that there was no
bentonite selectivity based on isoelectric point, and that bentonite fining resulted
in the removal of all the different protein fractions. Further, the amount of protein