Isolation And Multi-enzyme Ptoduction Of Non-Saccharomyces Yeasts

Non-Saccharomyces is limited to the first 2 or 3 days of fermentation after which they die as a result of ethanol toxicity. As these yeasts disappear, highly fermentative strains of the species Saccharomyces cerevisiae begin to multiply until they become solely responsible for alcoholic fermentation. Despite growing only during the first few days of fermentation, non-Saccharomyces yeasts produce a large number of compounds that can have a significant influence on the quality of the wine. The different between non-Saccharomyces yeasts and Saccharomyces yeasts is the production of enzymes. By interacting with substrates in the medium, these enzymes can increase yield and color extraction and enhance the characteristics of the wine. The aim of this work was to determine the ability of non-Saccharomyces yeasts to produce extracellular enzymes, and design of mixed starter cultures, for replacing a guided fermentation using pure Saccharomyces cerevisiae.The conclusions are as follows:(1)228 yeast strains were isolated from apples, grapes and dairy products, 47 yeast strains were Zygoascue.sp, 67 yeast strains were Hanseniaspora.sp, 29 yeast strains were Candida.sp, 76 yeast strains were Pichia.sp, only 9 yeast strains Yarrowia.sp.(2)Non-Saccharomyces yeasts has unique hydrolytic activity, selected 11 non-Saccharomyces cerevisiae strain, the activities of pectolytic enzymes, proteolytic enzymes, β-glucanases, xylanase andβ-glucosidase were determined:(1)Selected 3 non-Saccharomyces yeast strain have the ability to produce pectolytic enzymes, XYN A4 、 XYN N8 and XYN Y5, the enzyme activity was 41.94±3.12U/mL, 24.97±0.03U/mL and16.54±0.21U/mL respectively. The optimum pH were: 4.0, 5.0, 4.0, and optimum temperature were: 25℃,30℃, 25℃ respectively.(2)Selected 3 non-Saccharomyces yeast strain have the ability to produce proteolytic enzymes, XYN N8, XYN Y9 and XYN A4, the enzyme activity was 53.16±1.20U/mL, 31.04±2.01U/mL and70.27±1.50U/mL respectively. The optimum pH were: 5.0, 8.0, 5.0, and optimum temperature were: 30℃,35℃, 30℃ respectively.(3)Selected only 1 non-Saccharomyces yeast strain have the ability to produce β-glucanases, XYN A4 and the enzyme activity was 157.36±1.21U/mL. The optimum pH was 4.0 and optimum temperature was40℃.(4)Selected only 1 non-Saccharomyces yeast strain have the ability to produce xylanase, XYN A4 and the enzyme activity was 377.58±0.91U/mL. The optimum pH was 4.0 and optimum temperature was 40℃.(5)Selected 4 non-Saccharomyces yeast strain have the ability to produce β-glucosidase, XYN A8,XYN Y17, XYN N8 and XYN A4, the enzyme activity was 162.17±1.20U/mL, 130.92±2.01U/mL,273.84±3.50U/mL and 291.04±4.52U/mL respectively. The optimum pH were: 7.0, 5.0, 5.0, 7.0, andoptimum temperature were: 50℃, 60℃, 60℃, 40℃ respectively.(3)The present work was to detemine the ability of the yeasts to produce enzymes in pure and mixed cultures. Throughout the alcoholic fermentation, non-Saccharomyces yeast XYN A4(Candida) and industrial Saccharomyces cerevisiae ALG804 at an inoculation ratio 1:100(MSC1) mixed fermentation,whose has the higher enzymatic activity and presented throughout the alcoholic fermentation. MSC1 complete alcoholic fermentation with Saccharomyces cerevisiae pure fermentationsame same time was 7days. Thus, in beginning wine fermentation, add Saccharomyces cerevisiae ALG804 with cell concentration 105cells/mL and non-Saccharomyces yeast XYN A4 Candida.sp with cell concentration107cells/mL mixed fermentation could be an interesting alternative to a guided fermentation using a sterter culture of pure Saccharomyces cerevisiae ALG804.