【作者】 张瑾；

【导师】 王国栋； 惠竹梅；

【作者基本信息】 西北农林科技大学 ， 生物物理学， 2009， 硕士

【摘要】 葡萄汁中含有碳源、氮源,及其他利于酵母菌生存的微量化学物质。氮素营养是酵母菌进行正常酒精发酵所需的大量重要营养元素。葡萄中基本氮素化合物包括:游离α-氨基酸,铵态氮和少量小分子多肽。脯氨酸是亚氨基酸,不能在无氧条件下被酵母利用。因此,除脯氨酸外,游离α-氨基酸、铵态氮和小分子的多肽被称为酵母菌可同化氮(Yeast Assimilable Nitrogen, YAN)。葡萄汁中,酵母菌对含氮化合物可能的代谢途径有3条:在生物合成中被直接利用;被转变成相应的化合物后,再用于生物合成;通过转氨基作用(Transamination)将含氮化合物降解释放出游离态的铵离子或结合氮。在3条途径中,含氮化合物的骨架将作为代谢废物。葡萄醪中的氮素营养可以增加酵母菌的生物量,提高糖的利用效率,进而提高酵母菌的酒精发酵速率。酵母菌进行酒精发酵时,需要一定浓度水平的氮源,特别是酵母菌可同化氮(Yeast Assimilable Nitrogen,YAN)。酯类、高级醇和挥发性脂肪酸是发酵香气的主要贡献者。这些挥发性成分主要是酵母菌的糖和氨基酸代谢产生的。而且,氨基酸是挥发性成分的前体物质。酵母菌完成酒精发酵过程需要一定量的氮素营养,如果酵母菌可同化氮含量不足,在酒精发酵过程中会造成发酵停滞,以及H2S的产生。酒精发酵开始前或中期,向发酵体系中添加无机氮(铵盐类,如:磷酸氢二铵)、有机氮(氨基酸)可避免酒精发酵停滞带来的诸多问题。国外一些研究者指出添加氨基酸和铵态氮会对发酵速率产生影响,同时会影响醇类、酯类物质的生成。然而国内针对添加氨基酸和铵态氮对酒精发酵的挥发性成分的研究尚无报道。基于以上事实,本研究综合添加可同化氮浓度范围和种类(氨基酸和铵态氮),对酵母菌酒精发酵速率和发酵结束后挥发性成分进行测定分析,为酿造过程中氨基酸和铵态氮的添加提供依据。本研究通过对不同浓度可同化氮的模拟葡萄汁进行酒精发酵实验,分别对发酵过程中酵母菌的菌群数量,还原糖的消耗进行监测,以及对酒精发酵结束后挥发性成分进行测定。结果发现:随着可同化氮浓度升高,酵母菌群体数量增多,还原糖消耗增大,酒精发酵速率增大;进行二次添加的发酵汁,酒精发酵速率明显高于同一水平处理组;二次添加处理的高级醇含量明显高于一次添加处理组;随着可同化氮浓度的升高,乙酸、琥珀酸、高级醇、乙酸乙酯、辛酸乙酯、辛酸和十二酸的含量也升高;2-辛酮含量随可同化氮浓度的升高而降低。酵母菌可同化氮是酵母菌进行正常酒精发酵必需的大量营养元素,可同化氮浓度低于150mgN/L时,添加有机氮(氨基酸)和无机氮(铵态氮)均能促进酵母菌完成酒精发酵;可同化氮浓度升高,生成较多的有机酸和酯类。由此说明,酒精发酵前,通过对葡萄汁中可同化氮进行控制和适当的添加,可改善和促进酒精发酵结束后葡萄酒挥发性成分的生成,这对葡萄酒酿造和管理的意义重大。另外,酵母菌菌群数量关于可同化氮浓度的回归关系达到显著水平(p=0.021<0.05);酵母菌菌群数量关于时间的回归关系达到显著水平(p=0.025<0.05);糖的浓度关于可同化氮浓度的回归关系达到显著水平(p=0.041<0.05);糖的浓度关于时间的回归关系达到极显著水平(p=0.0001<0.01)。更多还原

【Abstract】 Grape juice contains carbon, nitrogen, and other micro-chemical substances which are conducive to the survival of the yeast. Nitrogen nutrition, a large number of important nutrients, is normal for wine yeasts during alcoholic fermentation. Freeα-amino acids, ammonium nitrogen and a small amount of low molecular peptides are basic nitrogen compounds in grape juice. Proline is imino acid that is not be used in anaerobic condition by wine yeasts. Therefore, in addition to proline, the freeα-amino acids, ammonium nitrogen and small molecules peptides are yeast assimilable nitrogen (Yeast Assimilable Nitrogen, YAN). There are three metabolic pathways by wine yeasts using nitrogen compounds: (1) It is in the direct use during biosynthesis; (2) It is transformed into the corresponding compounds, and then used for biosynthesis. (3) The nitrogen compounds release free ammonium ion or combined nitrogen through degradation. In these three ways, the framework of compounds which contains nitrogen will serve as the metabolic waste. The nitrogen nutrition in must can increase the wine yeast biomass, and it also improve the efficiency of the usage of sugar, thereby enhancing the alcohol fermentation rate. During alcohol fermentation, there should be a certation level of nitrogen for wine yeasts, particularly, yeast assimilable nitrogen(Yeast Assimilable nitrogen, YAN). Esters, alcohols and volatile fatty acids are major contributors to fermentation aroma. These volatile components are mainly produced through sugar and amino acid metabolism by wine yeasts. Moreover, the amino acid is the precursor substances of volatile components. Unless full of yeast assimilable nitrogen in must, the stuck happens during alcoholic fermentation which will cause the form of H2S gas. Before the start of alcoholic fermentation, or the mid-fermentation, addtion inorganic nitrogen (ammonium, such as: diammonium hydrogen phosphate) and organic nitrogen (amino acids) to system can avoid the stagnation problems caused by low yeast assimilable nitrogen. Some foreign researchers pointed out that amino acids and ammonium nitrogen will have an impact on fermentation rate, the production of alcohols and esters. However, the effect of addition amino acids and ammonium on volatile components in alcoholic fermentation has not been reported yet. According to these facts, this study analysed the impact of addintion yeast assimilable nitrogen (amino acids and ammonium) on the rate of alcoholic fermentation, and the volatile components content after fermentation, which will provide the basis for the enology industry. This research is on the alcoholic fermentation of chemically defined must. The determinations were carried out on the number of wine yeasts, the comsumption of sugar and the volatile compounds. It demonstrated that during fermentation the yeast number was increasing as the assimilable nitrogen content was increased and was added. And the fermentation rate of additional treatments were higher than others which the additional treatments were not carried out. After fermentation ended, the higher alcohol in additional treatments were higher than unprocessed group. The content of acetic acid, succinic acid, higher alcohol, octanoic acid ethyl ester, octanoic acid and dodecanoic acid increased with the increasing of assimilable nitrogen level, while 2-octanone production decreased. The yeast assimilable nitrogen was one of the necessary nutrients for yeasts in alcoholic fermentation. When the content of yeast assimilable nitrogen was lower than 150mgN/L, addition amino acids and ammonium promoted the yeast to finish the alcoholic fermentation. The productions of organic acid and esters were higher as the levels of yeast assimilable nitrogen increased. In addition, the regression relationships between the yeast number and assimilable nitrogen concentration is at a significant level (p=0.021<0.05). The regression relationship between the number of yeast and time reaches to the sigenificant level (p=0.025<0.05). The regression relationship between sugar concentration and assimilable nigtrogen concentration is also at a significant level (p=0.041<0.05). The regression relationship between sugar concentration and time is at a very significant level (p=0.0001<0.01).更多还原