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fi rst introduced to wine in 1987 by Marshall and Williams [ 25 ] and
has also been widely used in food proteomics [ 26 ]. However, this
highly resolutive method has been poorly applied for wine protein
analysis to date [ 7 , 9 , 10 , 15 , 27 , 28 ].
Among these studies, different methods were employed to
extract soluble proteins from wine before 2D-E analysis. In 2006,
Okuda et al. [ 28 ] have undergone successive steps to extract solu-
ble proteins from a Chardonnay wine. Briefl y, the wine was con-
centrated by rotary evaporation at 40 °C, precipitated by
ammonium sulfate 80 %, ultracentrifugated, dialyzed, and fi nally
the lyophilized proteins were fractionated by Sephadex G-100
chromatography before 2D-E (16 cm, pH 3-6). The main disad-
vantage of this method might be the numerous steps that are time
consuming, though more than 300 spots were visualized by
Coomassie Brilliant Blue (CBB-R250) staining. More recently,
Batista et al. (2009) [ 7 ] purifi ed the proteins from an Arinto white
wine by FPLC cation exchange chromatography, subsequently
desalted and lyophilized before 2D-E (13 cm, pH 3-10). A few
number of spots was retrieved on 2D gels, likely due to the low
sensitivity of the staining method employed, i.e., CBB-R250 or an
insuffi cient amount of protein loaded (not specifi ed). More
recently, Sauvage et al. (2010) [ 15 ] employed 2D electrophoresis
(18 cm, pH 3-10) to monitor the impact of enological treatments
on the protein content of a Chardonnay wine. In this latter study,
a partial purifi cation was undergone through adsorption of poly-
phenols on Fractogel, removal of polysaccharides (>150 kDa) and
then, concentration of wine proteins (between 5 and 150 kDa) by
successive ultrafi ltrations. Only ten spots were visualized by col-
loidal CBB (G-250) staining, despite the high amount of protein
loaded on the IPG strips (i.e., 300
g).
Here, we present an alternative method to extract proteins
from a Champagne base wine with minor modifi cations as com-
pared to the protocol published in our previous studies [ 9 , 10 ].
Briefl y, a Champagne base wine (Chardonnay) was subjected to an
ultrafi ltration in order to concentrate ten times the wine protein
content. The retentate was precipitated in ethanol with 15 % (w/v)
of TCA (ET/TCA) and fi nally, the protein pellet was solubilized in
an appropriate rehydration buffer. The wine protein sample was
separated and analyzed by 2D-E, using two IPG strips (7 cm) with
different pH gradients, pH 3-10 and pH 3-6. Thus, we were able
to visualize around 30 and 50 spots on pH 3-10 and pH 3-6 gra-
dients, respectively, by using a sensitive colloidal CBB-G250 stain-
ing [ 29 ]. Our precipitation method is easy to conduct, well adapted
to wine and has been applied successfully to must and champagne
( data not shown ). Another advantage is the use of 7 cm IPG strips,
instead of 18 cm [ 9 , 10 ] that allows a lower protein load, without
disturbing the subsequent MS analysis. It is an important fact to
take into account, since Champagne wines generally contain a low
μ
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