Yeast viability during fermentation and sur lie ageing of a defined medium and subsequent growth of Oenococcus oeni

Interactions between the yeast strain used for primary oenological fermentation and the bacterium used to conduct subsequent malolactic fermentation were studied under model winemaking conditions. A commercial Saccharomyces cerevisiae wine yeast (strains, EC 1118, AWRI 835 and CY-3079) was grown in a defined medium whose composition approximated grape juice. Fermentations by all strains reached dryness, and retained a cell viability of greater than 90% upon completion of fermentation. Highest total viable cell number and percentage of viable cells were recorded for EC 1118. A sur lie ageing of the fermented medium over a 12 week period revealed a bi-phasic decay of culture viability for all strains. Thus 99% of cells had died within 2 weeks post-fermentation. Viabilities were then stable for the subsequent 4–6 week period before a second decline phase ensued and ended in either a minimal ( ca 100 CFU/mL, EC 1118) or no viable cells being detected at 12 weeks of ageing. The growth response of an Oenococcus oeni inoculum to yeast culture supernatants, previously aged for up to 12 weeks in the presence or absence of yeast lees, was evaluated in a bio-assay. In this way, yeast strains could be designated as being either inhibitory, neutral or stimulatory to the growth of O. oeni (strain Lc5p). Inhibition by supernatants of strain EC 1118 was evident, but found to be reduced by ageing the supernatant (with or without lees). Conversely, longer ageing on yeast lees increased the magnitude of the stimulatory response in O. oeni (strain Lc5p) to the supernatant from the wine yeast (strain CY-3079).     

树脂原位吸附提高酒明串珠菌不对称合成(R)-2-辛醇的催化效率

通过运用原位吸附技术构建水/树脂反应系统,利用树脂的原位吸附来提高酒明串珠菌Oenococcus oeni CECT 4730不对称合成(R)-2-辛醇的催化效率。通过比较6种不同型号的吸附树脂对反应的影响,选出了最适的树脂AB-8。通过考察树脂的用量及水/树脂相中底物浓度对反应的影响,发现反应体系中加入一定量合适的吸附树脂,能够显著减小底物和产物的抑制,大幅度的提高初始底物浓度和反应产率。AB-8树脂加量为15g/L的水/树脂系统中,底物浓度为30g/L时,反应产率达到58%,较之水相系统提高了1.7倍。     

酒酒球菌高密度培养条件的优化

酒酒球菌（Oenococcus oeni）主导的苹果酸-乳酸（MLF）发酵是优质葡萄酒生产的重要工艺环节,制备高活菌数的酒酒球菌发酵剂是保障该环节顺利进行的重要前提之一。研究发现,一定量的L-苹果酸可以有效促进酒酒球菌的生长,并通过高密度培养条件优化提高酒酒球菌菌体密度。结果表明,通过正交试验得出的最佳高密度培养条件为初始pH值5.1、接种量3%、L-苹果酸添加量1 g/L。在此优化条件下,酒酒球菌ES-1的菌体密度较高,为（7.33±0.40）×109 CFU/mL;进一步结合化学中和法和半连续培养法,可使最终菌体密度达（1.67±0.11）×1010 CFU/mL,是对照组的11倍。该研究获得的高密度培养优化方案可为制备优质本土酒酒球菌发酵剂奠定基础。     

Protease activity of Oenococcus oeni viable cells on red wine nitrogenous macromolecular fraction in presence of SO2 and ethanol

The protease activity of Oenococcus oeni X₂L viable cells on red wine nitrogenous macromolecular fraction (NMF) as sole nitrogen source was determined. The enzyme releases growth factors for Oenococcus oeni. In presence of SO₂ and ethanol the rate of amino acids liberation was twofold higher. A peptide peak analyzed by HPLC with a retention time of 47 min diminishes markedly during the first 2 h incubation. O. oeni X₂L living cells are able to produce the exoprotease and to act on the red wine NMF in the presence of SO₂ and ethanol releasing essential amino acids for its survival.     

Hydrolysis of glycosidically bound flavour compounds from oak wood by Oenococcus oeni

Malolactic fermentation (MLF), which is conducted by lactic acid bacteria (LAB), has a significant influence on the stability and organoleptic quality of wine. Recent studies have shown that when MLF is carried out in oak wood barrels, LAB were also able to interact with wood and increase volatile compound contents such as vanillin during MLF. The release of these compounds indicates that LAB may convert vanillin precursors present in oak wood. In this work, the effect of commercial glycosidases on the released vanillin was firstly studied. This aldehyde is present in wood extracts in monoglycosidic forms where the major glycones are arabinose and xylose. Other aglycons released during MLF in barrels, syringaldehyde and whisky-lactones, can be considered as other sources of aroma. Secondly, strains selected with high activities toward glycoside substrates could hydrolyse vanillin glycoside precursors from oak wood with the same efficiency as commercial enzymes.     

β-Glucoside metabolism in Oenococcus oeni: Cloning and characterisation of the phospho-β-glucosidase bglD

The structural gene for a phospho-β-glucosidase from the oenologically important lactic acid bacterium (LAB) Oenococcus oeni has been cloned and its protein product characterised. This gene is found in a putative β-glucosidase operon of 2178 base pairs encoding 4 genes designated bglA to bglD. The bglA, B and C genes were not cloned and characterised, however, are thought to be phosphoenolpyruvate dependent phospho transferase system (PEP-PTS) components IIC, IIA and IIB which regulate the uptake, phosphorylation and translocation of β-glucosides across the cytoplasmic membrane. The cloned bglD was sequenced and expressed in Escherichia coli followed by purification. The purified bglD protein has 480 residues, a molecular mass of 55.5 kDa and shows high homology to known phospho-β-glucosidases. bglD exhibited high activity towards the phosphorylated β-glucoside para-nitrophenol-β-d-glucopyranoside-6-phosphate with a pH optimum of 5.5 and maintained similar levels of activity between temperatures of 4 °C and 40 °C. The enzyme was not active against non-phosphorylated β-glucosides.     

酒酒球菌抗酒精基因RAPD标记的研究

以抗酒精酒酒球菌和酒精敏性酒酒球菌为材料,提取基因组DNA构建抗性与敏性基因池,对RAPD-PCR反应体系进行优化,利用RAPD分子标记技术对45条随机引物进行筛选。结果引物S90在750bp附近扩增出了差异条带,且重复性好,这表明S90 750是与酒酒球菌抗酒精基因相连锁的RAPD标记。