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A Technical Guide for Wine Producers
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RECENT ARTICLES | WYNBOER HOME
Bentonite - more than just dirt
Paul
K.
Bowyer1
&
Virginie Moine-Ledoux2
1
Technical
Manager – Australasia, Laffort Oenologie Australia, 5 Williams
Circuit, Pooraka SA 5095, Australia
2
Product
Manager – Fining Agents, Laffort Oenologie, BP 40, 33015
Bordeaux Cedex, France
Stability
is an aspect of winemaking that all winemakers think about throughout
the year, whether it’s the stability of grape aromatics at the
crusher during vintage, the stability of colour during maturation or
micro oxygenation, or simply the stability of potassium bitartrate
(Bowyer, 2001) when Joe Public puts a bottle of white into the
fridge. Some of these stability problems are more easily dealt with
than others, such as using inert gases during winemaking or
inhibiting bitartrate precipitation through cold stabilisation (or
with greater technological finesse and efficiency by using a purified
mannoprotein such as Mannostab™
–
more
on that one in a future article). The one stability issue that we
have not yet found an alternative treatment for is protein
instability.
Proteins
The
structure of proteins governs many of their characteristics. For
starters, they can have
up to four distinct modes of structural variation (referred to as
their primary, secondary, tertiary and quaternary structures).
Proteins, like many organic molecules, are stable only under certain
conditions. They are affected by changes in pH, the polarity of the
solution they are in (Bowyer, 2003) and are also at the mercy of
exposure to heat. When some proteins are heated to levels that exceed
normal biological conditions (~37ºC) the hydrogen bonds that
help the molecule retain its secondary, tertiary and quaternary
structures are disrupted. Once these bonds are destroyed it is
practically impossible for them to regain their original form,
meaning that the protein has been irrevocably altered. We call this
process “denaturation”, and there are many common
examples of this process. Albumin, the major protein of egg white, is
soluble at room temperature but denatures and becomes insoluble when
heated, hence the term egg “white”. Crustacean shells
change colour when they are cooked, due to the denaturing of a shell
protein bound to a pigment. In the combined state the pigment appears
blue, but when the binding protein is denatured the pigment reverts
to its free molecular colouration of orange-pink.
A
protein’s primary structure (i.e. the amino acid sequence) can
have a major impact on the physical characteristics of the molecule.
For example, some amino acids have functionality that responds to a
change in pH, such as a wine during fermentation. Any change in
solution pH therefore can impact upon a protein’s primary
structure, which in turn can impact on higher protein structures and
also on the solubility of the protein. As with all weak acid and base
interactions, equilibrium can be reached (Bowyer, 2001) and an
overall or net charge on the molecule exists at any given pH. If the
charge is high, either positive or negative, the protein will be
quite soluble, since charged species dissolve in water fairly easily
(remember that wine is typically >85% water). However, if these
charges cancel each other out, the overall charge on the protein will
be zero. The solution pH at which this occurs is defined as the
protein’s isoelectric point (pI), and the protein’s
solubility will be lowest (Figure
1).
For
an excellent and recent overview on wine proteins see Waters et al
(2005). Proteins are present naturally in grapes, and are found in
the grape skin, flesh and seed (Fukui and Yokotsuka, 2003). Protein
is, in the case of both white and red wines, associated with the
formation of hazes and is typically removed before bottling. If left
in the wine the possibility of haze formation remains, which is
poorly regarded by consumers. The conditions under which a
protein-induced haze will form are known, with the most prominent
factor involved being the elevation of a wine’s temperature.
This can take place during transportation (it can get quite hot on
the back of a truck parked in the sun in South Africa for a few
hours), in the boot of a car driving away from a cellar door
(likewise) or even in that bottle you’ve got in the small wine
rack above the fridge, just above the heat exchanging element.
Most
wine proteins have an isoelectric point that lies above typical wine
pH (pH 3.0 - 3.5), thus in wine they will bear a net positive charge
(in contrast, laccase has a pI of around 2.5, meaning that in wine it
bears a negative charge, hence its lack of removal by treatment with
bentonite, and it remains quite soluble), (Ribéreau-Gayon et
al, 2006b). This is the fundamental principle on which the action of
bentonite towards wine protein is based, yet it is only half of the
equation. To understand the other half, we need to examine the
structure of bentonite in a likewise manner.
Bentonite
Bentonite is a very interesting
mineral, finding application in many diverse areas such as sand
casting, catalysis, dam sealing, intestinal evacuation, cat litter,
sandcastle building and of course, wine processing. It even plays a
role in the removal of contaminants found in wine (Ruediger et al,
2004). Bentonite is quite topical, making regular appearances in both
the scientific and industrial literature (for a recent article
focusing on efficiency of the usage of bentonite, see Colby et al,
2006). Bentonite is a hydrated aluminium silicate and a member of the
smectite class of clays. It commonly contains two forms of mineral,
kaolinite and montmorillonite, with the latter being the major form
found in the bentonite that is active in wine processing. Both
minerals have crystal structures that exist as sheets or layers.
Montmorillonite can be subdivided again into two forms, depending on
the predominant cation (positively charged ion) found in the
inter-laminar regions of the sub-crystalline structure. If this seems
a little confusing it will become clearer with a little more
explanation.
Bentonite
(both kaolinite and montmorillonite) is comprised mainly of oxides of
aluminium and silicon. The difference between kaolinite and
montmorillonite structures lies in the type of layering and the ratio
of silicon to aluminium oxides. In kaolinite the layering is quite
simple, with a layer each of silicon and aluminium oxides, as shown
in Figure
2. In
the case of kaolinite there is one very important difference between
the outer oxygen layers: the outer layer of oxygen atoms closest to
silicon is present as oxygen, whilst the oxygen atoms in the outer
layer adjacent to aluminium is present in the form of hydroxyl (OH)
groups in order to balance the charge. This means that the kaolinite
crystal lattice sheet structure is different on the top and bottom.
More importantly, the difference means that the upper surface of one
layer is able to hydrogen bond to the lower surface of the other, so
the overall structure resembles a layer cake.
The hydrogen bonding between the
layers of kaolinite imparts two important properties to the mineral.
Firstly, the layers are held quite tightly together. Secondly, as a
result of the difficulty of separating the kaolinite layers due to
the hydrogen bonding, it is not easy for water molecules to enter the
inter-laminar region. The importance of this will become clear in a
moment.
In
montmorillonite the same structural components as kaolinite are
present, with two important exceptions. In the first exception, a
second layer of silicon oxide is present, such that the aluminium
oxide layer sits in between the two silicon oxide layers. This forms
a symmetrical structure similar to a Jersey caramel lolly (Figure
3).
This means that on both upper and lower faces the lattice projects
only oxygen atoms, hence the possibility of hydrogen bonding is
removed. The second exception is that occasionally in the
montmorillonite structure aluminium is replaced with a different
metal such as iron, manganese or magnesium. This may seem like a
small change, but it is fact the crux of bentonite’s ability to
remove wine proteins.
When
aluminium ionises it forms trivalent Al3+.
Magnesium, manganese and iron tend to form divalent cations, hence
the substitution of a trivalent cation for a divalent cation
generates a deficiency in positive charge. Overall, therefore, the
lattice takes on a net negative charge.
In practical terms in montmorillonite
we have flat crystalline sheets bearing negative charges.
Electrostatically, like charges repel one another in a similar manner
to identical poles of magnets. In montmorillonite this translates
into the lattice sheets repelling one another quite fiercely, yet
bentonite does not simply explode into a fine dust of individual
lattice sheets. Why does this not happen?
In nature structures such as minerals
do not exist in totality as charged species. The charge must always
be balanced by other species. In the case of montmorillonite, this
charge excess is balanced by the presence of cations that reside in
the inter-laminar regions, between the lattice sheets. When these
cations are incorporated, the overall charge of the mineral is
balanced and the material as a whole bears no net charge. The type of
cation that fulfils this charge-balancing role impacts markedly on
the performance of the bentonite in terms of its capacity for
swelling and protein removal.
For
an excellent website to help you visualise kaolinite and
montmorillonite (listed therein as smectite) using movable
three-dimensional images structures, visit:
http://www.soils.wisc.edu/virtual_museum/displays.html. The images in
Figures 2 and 3 were generated using these models.
Sodium bentonite
In
sodium bentonite the inter-laminar region is occupied mainly by
sodium ions and water molecules. The high polarity of water molecules
makes them excellent stabilisers for cations and anions (negatively
charged ions) alike. Sodium forms monovalent cations (i.e. Na+).
The ionic radius of Na+
is
around 100 pm (1 x 10-10
m,
or 1 Å) (Aylward and Findlay, 1987), which makes Na+
reasonably
large in terms of the volume each cation displaces. In filling the
inter-laminar region of montmorillonite with a complement of water
molecules, Na+
is
acting as a sort of ionic glue, holding the lattice plates together.
It is not particularly efficient glue due to its small charge
density, a result of its monovalent charge and portly ionic radius,
so the overall structure is relatively loosely constrained.
When
sodium bentonite is added to water, the sheets begin to break apart,
since the Na+
ions
do not possess sufficient charge density to hinder the process to any
great extent. This process is expedited by using hot (higher energy)
water. Effectively, the absorption of water is very high because the
plates separate easily, meaning that the swelling capacity of sodium
bentonite is very high. This is the reason that the hydration of
sodium bentonite can take some time, requiring continual stirring,
often overnight.
The high dispersion rate of sodium
bentonite has some interesting ramifications. Firstly, the surface
area of sodium bentonite is very high. Secondly, the high dispersion
rate means that flocculation and settling are both slower, and the
formation of a relatively stable colloidal suspension can occur. This
can impact dramatically on wine or juice processing efficiency.
Thirdly, the protein removal capacity is very high, since the
activity for protein removal is governed primarily by the charge
density of the lattice and the surface area the mineral occupies: as
each increases, so does the protein binding capacity.
Calcium bentonite
The
only important difference between sodium and calcium bentonites is
the relative proportions of the two cations in the inter-laminar
region. In calcium bentonite, the divalent calcium ion predominates.
The ionic radius of Ca2+
is
99 pm (Aylward and Findlay, 1987), which is almost identical to that
of Na+.
Although the ions occupy the same volume in space the charge density
of the Ca2+
ion
is twice that of the Na+
ion.
In physical terms this impacts greatly on physiochemical
characteristics of the two minerals, since the negatively charged
montmorillonite lattice sheets are more tightly held together by the
calcium ions.
Calcium
bentonite, having a better “glue” in between the lattice
sheets in the form of calcium ions, does not disperse in water as
readily as sodium bentonite, meaning that hydration (swelling) does
not occur to the same extent. Flocculation and settling for calcium
bentonite are much more rapid than for sodium bentonite, since the
calcium ions cause faster re-association of the lattice sheets
(Figure
4).
In winemaking terms this means that calcium bentonites settle more
rapidly and to a more compact state than sodium bentonites. The
surface area presented by a suspension of calcium bentonite is not as
great as for sodium bentonite, even though the negative charge on the
lattice plates is equivalent, hence the protein removal capacity of
calcium bentonite is lower than for sodium bentonite. A graph
illustrating the differing protein removal capacities of some
bentonite samples with differing inter-laminar compositions is given
in Figure
5.
Sodium-activated calcium bentonite
Some bentonites naturally contain a
close ratio of sodium and calcium in the inter-laminar region of the
structure. It is also possible to alter the cationic balance in the
inter-laminar region of bentonite artificially through chemical
treatment. This is achieved by heating a calcium-rich bentonite in
water at approximately 80ºC with sodium carbonate. The purpose
of this process is to enrich the naturally calcium-dominant bentonite
with sodium, thereby increasing its protein removal capacity. The
results of this process of activation can vary depending on the type
(origin) of the bentonite used.
For
an example of a scientific evaluation of this process, see Gougeon et
al (2003). In this study the authors examined two calcium bentonites
before and after sodium “activation” to elucidate the
relative amounts of protein adsorption and the extent of Ca2+/Na+
exchange.
The findings of the study indicated that cation exchange occurs
mainly only at the edges of the inter-laminar region, which
constitutes a relatively low proportion of the total exchangeable
cation content. Activation did lead to an increase in the amount of
protein adsorption, although this was not quantified.
Bentonite mode of action –
ion exchange
Now
that we have an understanding of the structures of both proteins and
bentonite, we can consider more closely what actually happens when
the two are combined in wine. Recall that most wine proteins possess
a positive charge in wine, and that the lattice sheets of bentonite
have negative charges. When they are combined in wine, the proteins
are electrostatically attracted to and adsorb onto the surface of the
bentonite sheets. This process upsets the charge balance of the wine
medium, as positively charged ions (i.e. the wine proteins) are being
removed from the wine. This charge imbalance is corrected by the
concerted migration of cations (either Na+
or
Ca2+,
or other trace cations) from the inter-laminar region of the
bentonite into the wine. The action of bentonite, therefore, is an
ion-exchange process. In effect, when discussing the protein removal
capacity of bentonite we are really discussing its ion-exchange
capacity.
In consideration of this, wines for
export to markets where maximum sodium levels apply (such as the EU)
might require protein stabilisation with calcium rather than sodium
bentonite.
Parameters to consider when
choosing bentonite
Knowing
something about the structures of proteins and bentonite allows us to
better predict our fining needs. Firstly, if protein removal is your
priority then sodium bentonite is more effective. If you are
concerned about introducing extra sodium into your wine, then you
should consider using a calcium bentonite at the expense of protein
removal. If processing time is at a minimum then calcium bentonite
will sediment more rapidly. Variations in varietal protein contents
and compositions will have an effect, as well as climate differences
between growing regions and seasonal variations from year to year.
Wine pH also plays a role (Figure
6),
since at lower pH wine proteins will carry greater positive charge
density, which will reduce the bentonite requirement to achieve
stability. In effect, the correct product you should use depends on
your priorities with respect to the wine. It is a balancing act that
all winemakers must perform.
Speciality bentonite – aroma
retention
Bentonite
is well known for causing the loss of aroma compounds at higher rates
of application (Figure
7)
(Ribéreau-Gayon et al, 2006a), which reinforces the absolute
requirement for bench trials to be conducted on every wine before any
additions are made. Since bentonite is a natural product, being a
mineral that is obtained from mining operations in various locations
throughout the world, there can be significant variation between
products. Some bentonites will display greater protein removal
capacity, whilst others will have better lees compaction. In order to
make the best products available to its customers, Laffort Oenologie,
through its SARCO research facility, recently invested heavily in an
exhaustive examination of over 50 natural bentonites (all Laffort
Oenologie products are certified GMO-free and we use natural products
wherever possible). The samples were investigated for all parameters
that impact on wine quality and protein efficiency, such as protein
removal capacity, lees compaction and wine varietal aroma retention.
Some examples of the results are given in Figure
8.
The
result of this investigation was the development of two new products
in the Laffort Oenologie range: Microcol Alpha™
and
Microcol CL™
(Microcol™
is
an existing product sold in the EU). Microcol CL™
and
Microcol Alpha™
display
the characteristics indicated in Table
1.
Figure
9
shows, for example, the excellent lees compaction of Microcol CL™
in
comparison with a sodium bentonite. Additional data are presented
below.
Aroma retention
Two
of the characteristic aroma molecules found in Sauvignon blanc wine
are 3-mercaptohexanol (3MH) and 4-mercapto-4-methylpentan-2-one
(4MMP), contributing passion
fruit/grapefruit and boxwood aromas respectively. Such compounds are
known to be removed by bentonite treatments. Microcol CL™
and
Microcol Alpha™
were
compared with other commercial bentonites (brands not disclosed) for
their respective abilities to permit the retention of these compounds
in wines containing this varietal.
In
Figure
10
the levels of 3MH remaining such a wine are illustrated, after
treatment with increasing levels of the designated bentonites. It can
be seen that Microcol Alpha and bentonite X perform very well in this
test, whilst Microcol CL removes slightly more 3MH. Bentonite Z,
performs relatively poorly, with a significant reduction in 3MH
levels in the wine at dosages above 50 g/hL.
Microcol
CL™
was
also compared with another bentonite (E) and Microcol™
for
the effect on 4MMP levels in a Sauvignon blanc wine (Figure
11).
Above dosages of 10 g/hL some reduction in 4MMP levels remaining in
the wine is noted for all samples, with bentonite E performing
relatively poorly. At a high dosage rate (100 g/hL) Microcol CL still
allows the retention of more than 80% of the original 4MMP
concentration in the wine, making it particularly effective for
retaining grapefruit/passion fruit characters in this varietal.
Dosage requirement and lees volume
In
addition to improvements in wine quality, winemakers are concerned
with increases in logistics and efficiency. For
these reasons Microcol CL™
and
Microcol Alpha™
were
compared with some other commercial bentonites regarding dosages
required to achieve protein stability and lees compaction. As can be
seen in Figure
12,
Microcol Alpha™, being a natural sodium bentonite, performs
exceptionally well at protein removal with a correspondingly higher
volume of lees. In comparison, Microcol CL™, being a natural
calcium bentonite, requires a higher dosage to effect protein
stability, yet displays excellent lees compaction.
Laffort
South Africa is represented by Grettchen Visagie and Morne Kemp
– they
can be contacted at grettchen.visagie@laffort.com and
morne.kemp@laffort.com respectively.
Summary
A
working understanding of protein and bentonite structures allows
winemakers to make informed choices on the products and methodologies
that are most likely to give the highest performance under a given
set of conditions. Laffort Oenologie
caters for variation in these parameters by providing natural
bentonite products that offer high performance in different areas,
such as the capacity for protein removal (i.e. ion exchange), lees
volume and, importantly for producing high quality wines, wine
varietal aroma retention. Winemakers, working in combination with
Laffort Oenologie, are therefore provided with understanding of the
technical issues, in addition to the products best suited to perform
the task required.
References
Bowyer,
P.K. (2001).
Bitartrate instability and the pH see-saw, The Australian Grapegrower
and Winemaker, September, 100 - 101.
Bowyer,
P.K. (2003). Molecular polarity – it’s behind more than
you think, The Australian Grapegrower and Winemaker, November, 89 -
91.
Colby,
C.B., Waters, E., Nordestgaard, S. & O’Neill, B.K. (2006).
Bentonite fining: can we improve performance and efficiency and
decrease value losses? The
Australian and New Zealand Grapegrower and Winemaker – Annual
Technical Issue 2006,
82 - 88.
Gougeon,
R.D., Soulard, M., Miehe-Brendle, J., Chezeau, J-M., Le Dred, R.,
Jeandet, P. & Marchal, R. (2003). Analysis of two bentonites of
enological interest before and after commercial activation by solid
Na2CO3.
Journal
of Agricultural and Food Chemistry,
51,
4096 - 4100.
Fukui,
M. & Yokoysuka, K. (2003). Content and origin of protein in white
and red wines: changes during fermentation and maturation. A.J.
Enol. Vitic.
54:
178 - 188.
Ribéreau-Gayon,
P., Glories, Y., Maujean, A. & Dubourdieu, D. (2006a). Handbook
of Enology Volume 2 – The Chemistry of Wine Stabilization and
Treatment. Wiley: Chichester, pp 132.
Ribéreau-Gayon,
P., Glories, Y., Maujean, A. & Dubourdieu, D. (2006a). Handbook
of Enology Volume 2 – The Chemistry of Wine Stabilization and
Treatment. Wiley: Chichester, pp 290.
Ruediger,
G.A., Pardon, K.H., Sas, A.N., Godden, P.W. & Pollnitz, A.P.
(2004). Removal of pesticides from red and white wine by the sue of
fining and filter agents. Australian
Journal of Grape and Wine Research,
10,
8 - 16.
Waters,
E.J., Alexander, G., Muhlack, R., Pocock, K.F., Colby, C., O’Neill,
B.K., Høj, P.B. & Jones, P. (2005). Preventing protein
haze in bottled wine. Australian
Journal of Grape and Wine Research,
11,
215 - 225.
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