The Interaction Between Non Saccharomyces And Saccharomyces Cerevisiae And Its Effect On The Quality Of Haihong Wine

In order to enrich the aroma substances in Haihong fruit wine and enhance its quality,non-saccharomyces,Klockera apiculata（K.apiculata）was used to ferment Haihong fruit wine together with Saccharomyces cerevisiae（S.cerevisiae）in this study.On the basis of preliminary understanding the physiological and biochemical characteristics of Klockera apiculata（K.apiculata）and its fermentation characteristics,the interaction between Saccharomyces cerevisiae（S.cerevisiae）and K.apiculata in the mixed fermentation of Haihong wine was investigated furtherly and the effects of K.apiculata on the flavor components of Haihong wine were explored.The main research content,obtained results and conclusions are as follows:1)Study on the physiological characteristics of two kinds yeast The growth of two kinds of yeast in culture medium with different concentrations of SO₂,ethanol and tannin were studied.The results showed that K.apiculata was sensitive to SO₂ and ethanol in culture solution,and the maximum tolerable contents of SO₂ and ethanol were 100 mg/L and 3.6%vol respectively.When the content of SO₂ and ethanol in the culture medium are over these values,the growth of K.apiculata is threatened even death.When tannin content is less than 6g/L,the growth of the two yeasts is basically stable,whereas,when the content of tannin is higher than 6g/L,the growth of both yeasts is affected to a great extent.The determined content of tannin in Haihong fruit juice is 3.5 g/L,therefore,it does not affect the growth and reproduction of yeast.The enzyme-producing characteristics of two kinds of yeast used were explored using a select medium and results showed that K.apiculata can secrete extracellular pectinase.And S.cerevisiae can not secrete pectinase,but can secrete protease.In addition,K.apiculata has stroger ability to secreteβ-glucosidase more than S.cerevisiae.2)Study on the interaction between K.apiculata and S.cerevisiae in fermentation The mix inoculum with different ratio of K.apiculata and S.cerevisiae were used to ferment Haihong juice,using a pure yeast fermentation of K.apiculata or S.cerevisiae as a reference.During fermentation the changes of alcohol content,release of CO₂ and the bacteria number of two yeasts were measured to interpret the interaction between S.cerevisiae and K.apiculata in the fermentation.The results showed that the growth of K.apiculata was inhibited obviously after 48 hours fermentation with three kinds of mixture inoculation,and K.apiculata did not detected when fermented over 108 hours.In order to investigate whether there are other substances in the fermentation inhibiting the growth of K.apiculata besides ethanol,the effects of the fermented supernatant and the yeast cell crushing slurry,on the growth of K.apiculata were investigated,when the fermentations with the single S.cerevisiae and the mixed fermentation of two yeast were carried out to 36 hours.Then,the sensitivity of the fermented supernatant to protease and heat were explored.The results showed that when the fermentation of S.cerevisiae to 36hours,neither the supernatant nor the cytoplasm did not affect the growth of K.apiculata.However,the two yeasts were mixed and fermented to 36 hours,the supernatant obtained by centrifugation severely affected the growth of K.apiculata.Furtherly,the supernatant was treated with protease or heat,and then added to the culture medium and it was found that the inhibitory effect of K.apiculata basically disappeared.Therefore,it is speculated that the inhibitor in the supernatant may be a polypeptide.The above results prove that the mixed fermentation of S.cerevisiae and K.apiculata,S.cerevisiae can secrete both ethanol and extracellular polypeptides and they cause a serious threat to the growth of non-Saccharomyces cerevisiae such as K.apiculata.3)Study on the effect of K.apiculata on the aroma content of Haihong fruit wine The volatile components in the samples were extracted by headspace solid-phase microextraction（HS-SPME）,and the flavor components in wine samples were analyzed by GC-MS.The results showed that the aroma components of Haihong juice are dominated by aldehydes,while the aroma components of fermented Haihong wine are mainly alcohols and esters.The mixed fermentation of K.apiculata and S.cerevisiae could increase the types and contents of esters in the Haihong wine,while the content of higher alcohols decreased.With the extension of the interval between sequential inoculations,the content of esters in the final fermentation liquor and the proportion in the total aroma components are on the rise.The best wine sample is the fermented for 5 days by yeast of K.apiculata and then sequentially inoculated S.cerevisiae to continue fermentation until the end.Relative to the fermentation of single strains of Saccharomyces cerevisiae,the content of esters increased by72.3%,and the content of higher alcohols decreased by 62.8%.In the wine sample,typical aromas of citrus fruits such as orange and lemon were obviously given to fruit wine.In addition,the aromas of honey and bitter almond were also found in the wine samples.It can be concluded that:（1）Non-Saccharomyces cerevisiae can stimulate the self defense mechanism of Saccharomyces cerevisiae during mixed yeast fermentation,as a result,Saccharomyces cerevisiae secretes ethanol and other substances to inhibit the growth of non Saccharomyces cerevisiae and to ensure itself advantages growth;（2）The inhibit substances for non Saccharomyces cerevisiae are of polypeptides with thermal instability;（3）K.apiculata can enrich aroma substances and enhance the quality of Haihong fruit wine in mixed yeast fermentation of Haihong fruit wine;（4）The ideal way of mixed fermentation is a sequential fermentation in the order of K.apiculata to S.cerevisiae.