【作者】 王玉霞；

【导师】 刘树文；

【作者基本信息】 西北农林科技大学 ， 葡萄与葡萄酒学， 2008， 硕士

【摘要】 在影响葡萄酒质量与品质的诸多因素中,生态条件与葡萄品种、工艺条件、酿造设备和发酵微生物这四个方面起着决定性的作用。在其他三个因素相对稳定的情况下,葡萄酒微生物的酿造学特性则是葡萄酒质量和感观风格的决定性因素。酵母菌是葡萄酒酿造中重要的微生物,其多样性和分类鉴定是选育优良酿酒酵母菌株的关键和基础。从葡萄品种原产地的环境中筛选出可以充分展现该品种个性、土壤特点、地域特性的酵母,并酿出风格独特、具有品种典型性的葡萄酒。目前国际市场上出现了适用于不同产地、不同葡萄酒类型和葡萄品种的“特种”或“专用”酵母,而且已经开始逐渐取代“广适性”酵母的应用。由此可见优良的酵母菌种,对葡萄酒的质量和品质起着至关重要的作用。河北沙城地区是我国主要的葡萄酒产区之一,历史悠久,酒厂众多。选取在该产区代表性的中国长城葡萄酒有限公司及中法庄园进行菌种采集工作。从酒厂环境、葡萄园土壤、葡萄浆果、不同浓度SO2和酒精处理以及未处理的自然发酵醪液中分离酵母菌株,研究SO2和酒精处理醪液中酵母菌群的变化。利用WL培养基对菌株进行初步分类,选取有代表性酵母菌的纯培养,通过5.8S-ITS区的PCR/RFLP分析对所筛选菌种进行鉴定。鉴定属于酿酒酵母的菌株,则用来进行优良起泡葡萄酒酵母菌株各项指标的测试,筛选适合该产区特定生态地理条件及工业条件、具有鲜明风格特征的优良起泡葡萄酒酵母菌株。通过一系列试验得到以下结果:菌种的分离和鉴定试验共得到葡萄酒相关酵母562株,所有菌株在WL营养培养基上培养5d后形成的菌落颜色和形态共分为15大类。根据每一类别中所包含的菌株数量,适当选取一定数量的菌株进行5.8S-ITS区RFLP分子鉴定。在所有采集的菌株中,酿酒酵母(Saccharomyces cerevisiae)占的比例最大,共分离得到291株,占总数的51.7%,绝大部分来自自然发酵;其次为柠檬形克勒克酵母(Kloeckera apiculata),共有44株,占总数的7.8%;最少的是酒香酵母(Brettanomyces bruxellensis),共有6株,占总数的1.1%。SO2和酒精处理对自然发酵中酵母菌群的影响在添加SO2和酒精处理的自然发酵的试验中,首先启动发酵的是一些对SO2不太敏感,和对酒精耐受性差的酵母菌,随后是较敏感的种类如毕赤酵母和隐球酵母等。发酵初始阶段酵母的群体消长情况既受SO2的影响,又受发酵产物酒精浓度的影响。在发酵中期至后期,绝大部分酵母为酿酒酵母,但也能分离到少量的耐SO2和酒精的拜耳接合酵母和贝酵母。优良起泡葡萄酒酵母菌筛选通过分子鉴定,以291株酿酒酵母(Saccharomyces. cerevisiae)为试材,进行起泡葡萄酒酵母筛选的各项测验试验。在产气能力初筛实验中筛选出了62株泡沫、细腻度、持久性都相对较好的菌株。通过自溶及絮凝性测试、产气复筛、耐酒精能力测试、SO2耐性试验、耐低温能力和耐压力测试后,最终筛选得出Z27、Z25、Z28为优良菌株。起泡葡萄酒品评与分析通过二次发酵起泡酒及基酒样品香气成分的气相色谱-质谱分析,共检测到了48种不同成分的香气物质。在基酒中共鉴定出了45种香气物质,对照、Z25及Z28菌株酒样中各有46种香气物质,Z27菌株酒样中有47种香气物质。在所有香气成分中,醇、酸、酯类化合物种类最多,占总类别的82.8%。且醇类物质在酒样的香气物质中占据主导地位,其次是酯类和酸类化合物。所有5个酒样都含有的香气物质中,经过二次发酵后,绝大多数香气化合物的相对含量都较基酒中的有所增加,而不同菌株之间的增幅有所差异。更多还原

【Abstract】 The quality and character of wine are mainly determined by four factors: ecological condition and grape varieties, technology, brewing equipment and fermentation microbiology. The zymurgy’s character of microbiology is the dicisive factor under the hypothesis that the others are relative steady. Wine yeasts are important microorganisms in winemaking. Their diversities and identification are the main bases to screen excellent stains. Strains isolated from wine producing regions are usually more adapted to their own climatic conditions, grapes and also partially responsible for wine quality and style. At present, more and more special or exclusive yeasts has been used in the market, which just were applicable to special wines and particular regions and replaced regularly eurytropy yeasts. We can not deny that wine yeasts are very important for wine quality and style.Shacheng is one of famous and long history wine producing region in China. Many wineries located in here. Yeast strains were isolated and screened in Great Wall Wine Co., LTD and Sino-France chateau. Yeast strains were isolated from winery environment, earth of vineyard, grapes, and SO2 and alcohol treatments and untreatment spontaneousness fermentation. The influence of SO2 and alcohol treatments on yeast floras was also researched. Strains were preliminarily classified on WL medium. Then typical and pure cultured yeasts were chose and identificated by PCR/RFLP of 5.8S-ITS method. All the Strains belonging to Saccharomyces cerevisiae were used to screen excellent sparkling wine yeasts.Isolation and IdentificationThe 562 yeasts isolated in our experiment were belonging to 15 classified types after 5 days on WL medium according to the colony colour and morphology. Then some strains were chose in accordance to the strain numbers of every types and identified by PCR/RFLP analysis of the amplification fragments of 5.8S-ITS.The proportion of strains identified as S. cerevisiae was 51.7%, 291strains in all yeast strains, mostly came from spontaneous fermentation. There were 44 strains belonging to Kloeckera apiculata, 7.8% proportion. The least was Brettanomyces bruxellensis, 6 strains, just 1.1%.The Influence on Yeast Floras from Spontaneous Fermentation under Different SO2 and Alcohol Treatment MethodsIn the spontaneous fermentation experimentation under SO2 and alcohol treatments, the yeast floras firstly starting fermentation were insensitive to SO2 and lower tolerance to alcohol, subsequently, such as Pichia and Cryptococcus, were sensitive to SO2. The rise and decline of yeast floras during prime fermentation phase was influenced by not only SO2 but also alcohol concentration. From middle to final stage, S. cerevisiae were almost majority, but still some strains which tolerance to SO2 and alcohol were higher were isolated also, for instance Saccharomyces bayanus and Zygosaccharomyces bailii.Screening of Excellent Sparkling Wine Yeast StrainsThere were 291 strains identified as S. cerevisiae, which been used to screening experiment through various tests. After prescreening on producing gas capability, 62 wine yeast strains has been selected which bubble was fine and permanence. Z27、Z25、Z28 were the final excellent three yeast trains after autolysis, flocculability, producing gas capability and tolerance of alcohol, SO2 and low temperature fermentation capability tests.Tasting and Analysis of Sparkling WineTotal 48 kinds aroma compounds were detected by GC/MS analysis from sparkling wines and base wine, of which 45 kinds in base, 46 kinds in sparkling wines fermented by strains Z25, Z28 and CK. The content of alcohol, acid and ester total up to 82.8% in all aroma composition, alcohol was the leading composition, next were acid and ester. The content of aroma compounds in all five wine samples increased compare with that of existing in base wine after second fermentation, but the quantity depends on strains.更多还原