Effect Of Low PH On Alcoholic Fermentation And Primary Study Of Acid Stress Response In Saccharomyces Cerevisiae

Constant intracellular pH is vital to maintain normal function of organism, for example enzyme reaction, protein folding and function are all dependent on intracellular pH. When the environmental pH is very low, intracellular pH of organisms will be decreased, which will seriously inhibit the growth and metabolism of organisms. The aim of this study is to investigate the influence of low pH value on growth and alcoholic fermentation of Saccharomyces cerevisiae, and further uncover the stress response of Saccharomyces cerevisiae on low pH stress.3S. cerevisiae strains Freddo, BH8, N°7303were selected from12yeast strains by the different tolerance to low pH value. And4pH value:4.50(pH value suitable for growth and alcoholic fermentation to yeast),3.00and2.75(pH value repressive to some extent for growth and fermentation to yeast),2.50(pH value repressive severely for growth and fermentation to yeast), different effect on yeast growth respectively, were chosen. Effect of these4pH values on growth and fermentation performance of these3S. cerevisiae strains was studied. Comparing with the optimal pH4.50, the production of ethanol and glycerol, the relative expression of ADH1, GPD1, ALD6(main gene encoding key enzyme in acetic acid formation), the response of HOG1-the encoding gene of the key kinase of HOG/MAPK pathway, its related regulatory gene FPS1,3genes HAA1, PMA1, PDR12, related to weak acid stress were observed under3low pH value3.00,2.75,2.50when they were inoculated into model synthetic medium of low sugar for1h and2h. Stationary phase cells of BH8in model synthetic medium of low sugar, logarithmic phase cells of BH8in model synthetic medium were also investigated. The main results are as follows:1. Among the12strains, strain Freddo was the most tolerant one to low pH stress, strain BH8was the middle one, and N°7303was the weakest one. pH4.50was the optimal pH value for growth and fermentation, while S. cerevisiae would encountered low pH stress under pH≤3.0. Among the4pH values, pH2.75and2.50had more vital effect on yeast growth and fermentation products in contrast with pH4.50and3.00. In general, low pH value showed the properties of prolonging yeast lag phase, decreasing accumulated mass loss, slowing down the consumption rate of total sugar, increasing final content of acetic acid and glycerol, and decreasing final content of ethanol and L-succinic acid, increasing catalase activity and the relative expression of CTT1in logarithmic phase cells of3strains.2. Both ethanol and glycerol production of the3strains decreased with the increase of low pH stress. Ethanol production of strain BH8in model synthetic medium was lower than of it that in model synthetic medium of low sugar, glycerol production of BH8in model synthetic medium exceeded than that of it in model synthetic medium of low sugar.3. In terms of the relative expression level of gene, the level of ADHl in3strains was all deduced with the increase of pH stress. While in logarithmic phase cells, the relative expression level of gene GPD1, HOG1, FPS1, HAA1, PMA1, PDR12was all related to pH tolerance of strain and kept a good consistency in the same strain. For Freddo, the level of these6genes almost was the highest in samples of pH3.00-2h or2.75-2h(except PMA1); For N°7303, the level was stable and low; For BH8, the level of samples in pH2.50incubated for2h was the highest, while the level of other samples was low. In stationary phase cells, the level of FPS1, HAA1, HOG1, PDR12at low pH was much higher deduced than the samples at pH4.50, while the level of GPD1, PMA1in all samples was exactly the same. As to model synthetic medium, there was no significant difference among the level of all genes except GPDl, which level induced by the pH stress.The response mechanism of S. cerevisiae to low pH stress in must was discussed according to the pH tolerance of these3strains. And the hypothesis was put forward that the response mechanism was both involved in general stress response pathway and special stress response pathway; it adhered to typical weak acid response, and other stress response meanwhile. Detail mechanism needs further researches.