Test samples were left undisturbed for 15 min at room temperature. Thereafter, 100 μL aliquots were carefully withdrawn from just below the meniscus and added to 900 μL 100 mM EDTA, pH 7 contained in a cuvette. The absorbance of both control and test samples was determined at 600 nm. The average of five control

observations was obtained (denoted A). Each of the five individual test replicate observations (denoted B) was used to determine the replicate percentage sedimentation as follows: replicate % sedimentation=(A−B)/A. The percentage sedimentation of a sample was determined from an average of five replicate percentages as calculated above. The percentage sedimentation reported reflects the arithmetic mean of three independent determinations. Samples of fermented red wines containing lees were homogenized, diluted and filtered through 0.22-μm Durapore® membrane filters (Millipore Corporation, MA) and immediately frozen selleck by plunging into subcooled liquid nitrogen (‘slush’). Thereafter, filters containing wine lees samples were freeze dried, lightly sputter-coated with gold and viewed with an LEO 1450 scanning electron

microscope (Carl Zeiss, Jena, Germany) at 5 kV accelerating voltage. Turbidity of the wines after racking was evaluated using an LP2000 turbidity meter (Hannah Instruments, Bedfordshire, UK). The turbidity meter was calibrated before use as detailed in the instruction manual. Bottled

wines (five per replicate fermentation) were allowed to stand undisturbed for CH5424802 supplier 5 days after racking. Thereafter, a 10-mL aliquot was removed from below the meniscus from each bottle and dispensed down the inside of a clean cuvette to avoid the formation of air bubbles. All measurements were taken with samples equilibrated to room temperature. The turbidity of wines is presented as formazine turbidity units. Values reported in this Wilson disease protein study reflect the mean of experiments performed in triplicate (five measurements per replicate) and error bars represent SDs. In this study, paired t-tests or one-way anova were used to statistically compare data obtained for BM45 and VIN13 wild-type strains with that of transgenic yeast strains. Statistical tests were performed using graphpad instat version 3.05 32 bit for Windows 95/NT (GraphPad Software, San Diego, CA). Using a microsatellite PCR strain-typing method that targets δ sequences confirmed that alcoholic red wine fermentations were performed by the inoculated BM45 and VIN13 wild-type wine yeast strains. As reported previously (Govender et al., 2010), using a screening system that incorporated sensitivity to SM, flocculation ability (FLO5 transformants) and lack of invasiveness (HSP30p-FLO11 transformants) confirmed that alcoholic red wine fermentations were performed by the inoculated transgenic wine yeast strains.