Copper Tolerance and Biosorption of <i>Saccharomyces cerevisiae</i> during Alcoholic Fermentation

<div><p>At high levels, copper in grape mash can inhibit yeast activity and cause stuck fermentations. Wine yeast has limited tolerance of copper and can reduce copper levels in wine during fermentation. This study aimed to understand copper tolerance of wine yeast and establish the mechanism by which yeast decreases copper in the must during fermentation. Three strains of <i>Saccharomyces cerevisiae</i> (lab selected strain BH8 and industrial strains AWRI R2 and Freddo) and a simple model fermentation system containing 0 to 1.50 mM Cu²⁺ were used. ICP-AES determined Cu ion concentration in the must decreasing differently by strains and initial copper levels during fermentation. Fermentation performance was heavily inhibited under copper stress, paralleled a decrease in viable cell numbers. Strain BH8 showed higher copper-tolerance than strain AWRI R2 and higher adsorption than Freddo. Yeast cell surface depression and intracellular structure deformation after copper treatment were observed by scanning electron microscopy and transmission electron microscopy; electronic differential system detected higher surface Cu and no intracellular Cu on 1.50 mM copper treated yeast cells. It is most probably that surface adsorption dominated the biosorption process of Cu²⁺ for strain BH8, with saturation being accomplished in 24 h. This study demonstrated that <i>Saccharomyces cerevisiae</i> strain BH8 has good tolerance and adsorption of Cu, and reduces Cu²⁺ concentrations during fermentation in simple model system mainly through surface adsorption. The results indicate that the strain selected from China’s stress-tolerant wine grape is copper tolerant and can reduce copper in must when fermenting in a copper rich simple model system, and provided information for studies on mechanisms of heavy metal stress.</p></div

Intracellular elemental composition of <i>S</i>. <i>cerevisiae</i> BH8 before (a) and after (b) cultivated in MSM with 1.50 mM Cu²⁺ for 48h.

<p>Intracellular elemental composition of <i>S</i>. <i>cerevisiae</i> BH8 before (a) and after (b) cultivated in MSM with 1.50 mM Cu²⁺ for 48h.</p

Intracellular images (×20000) of <i>S</i>. <i>cerevisiae</i> strain BH8 before (a) and after (b, c) culturing in MSM with 1.50 mM Cu²⁺ for 48 h; CW: cell wall; N: cell nuclear; PM: plasma membrane; V: vacuole.

<p>Intracellular images (×20000) of <i>S</i>. <i>cerevisiae</i> strain BH8 before (a) and after (b, c) culturing in MSM with 1.50 mM Cu²⁺ for 48 h; CW: cell wall; N: cell nuclear; PM: plasma membrane; V: vacuole.</p

Images (×10000) of yeast surface of <i>S. cerevisiae</i> strain BH8 cultivated in MSM with 0 (control), 0.50, 1.00 and 1.50 mM Cu²⁺ for 24 h and 48 h.

<p>Arrows indicate pits on individual cell surfaces.</p

Copper ion concentration (A, Control; B, strain A; C, strain B; D, strain F) of MSM during fermentation for <i>S</i>. <i>cerevisiae</i> strains in MSM with 0 (control) (A*, B*, F*), 0.50 (A*, B**, F***), 1.00 (A*, B**, F***) and 1.50 mM (A*, B**, F***) Cu²⁺.

<p>(*, **, and *** represent different statistical significance level, CI = 0.95, n = 3).</p

Fermentation must reducing sugar (A, strain A; B, strain B; C, strain F) and fermentation ethanol concentration (D, strain A; E, strain B; F, strain F) for <i>S. cerevisiae</i> strains in MSM with 0 (control) (A*, B*, F*), 0.50 (A*, B**, F***), 1.00 (A*, B**, F***) and 1.50 mM (A*, B**, F***) Cu²⁺.

<p>(*, **, and *** represent different statistical significance level, CI = 0.95, n = 3).</p

Elemental composition of yeast surface of <i>S. cerevisiae</i> strain BH8 cultivated in MSM with 0 (control), 0.50, 1.00 and 1.50 mM Cu²⁺ for 24 h and 48 h.

<p>Elemental composition of yeast surface of <i>S. cerevisiae</i> strain BH8 cultivated in MSM with 0 (control), 0.50, 1.00 and 1.50 mM Cu²⁺ for 24 h and 48 h.</p

Removal ratio η (A) and adsorption efficiency <i>A</i> (B) of Cu²⁺ on <i>S</i>. <i>cerevisiae</i> strains AWRI R2 (A), BH8 (B) and Freddo (F) at the end of alcoholic fermentation in MSM with 0.50, 1.00 and 1.50 mM Cu²⁺.

<p>Removal ratio η (A) and adsorption efficiency <i>A</i> (B) of Cu²⁺ on <i>S</i>. <i>cerevisiae</i> strains AWRI R2 (A), BH8 (B) and Freddo (F) at the end of alcoholic fermentation in MSM with 0.50, 1.00 and 1.50 mM Cu²⁺.</p