Study On Screening And Enological Characteristics Of Malolactic Bacteria With High Malolactic Enzyme Activity

Malolactic fermentation (MLF) is a bacterial fermentation which occurs in wine and results in the conversion of malic acid to lactic acid and dioxide.This research was based on the malolactic bacterium isolation system established by past studies. In this paper, in order to obtained instinguished strain, isolation and screening of malolactic bacteria were carried out in main wine-making area in Ningxia Province. By testing enological characteristics, adaptivity and malolactic enzyme (MLE) activity of isolates, a strain with high MLE activity were obtained. The results showed that:In vinage of 2002 and 2003, 12 isolates of malolactic bacteria from main wine-producing regions of Ningxia Province were isolated and screened. After conformed the isolates on malolactic fermentation (MLF) ability, 12 isolates were tested for their capacity to resist to the individual stress condition of low pH, high ethanol and SO2 content. After testing the selected strain,s growth under the combination of external stress physical conditions of ethanol, pH and SO2, the results showed that finally selected strain of No. N7-03 had good adaptation to adverse conditions. The isolate had much high adaptability to stress environments than commercial strain O. oeni 31DH with high malolactic enzyme (MLE) activity of 117.05mg malate/log(cfu/ml). The characteristics of cell and colony morphology of No.N7-03 were observed. The phenetic characteristics including Gram stain, catalase test, the adaptation to different stress conditions and producing acid from carbohydrates were tested. The strain was identified as Oenococcus oeni (formly Leuconostoc oenos), the malolactic enzyme gene(mle) sequence of the isolate was also confirmed the determinative result.A DNA fragment from No.N7-03 genome about 530bps was amplified from geneme DNA of No.N7-03 using PCR technique. The fragment was then cloned into pCU19 vector and subsequently transformed into E. coli JM109. The sequencing result demonstrated that the cloned DNA fragment is malolactic enzyme gene (mle). It is an ORF of 527bp including signal peptide enconding region, and it has the same sequence as described in literature.The volatile components of Cabernet Sauvignon dry red wines were analyzed by gas chomatography-mass spectrometry after carried out MLF. The results showed that the main aroma components before and after MLF were volatile acids, ketones, alcohols, esters, phenols,et al..In these components, 2-methyl-1-propanol, ethyl acetate, 3-methyl-1- propanol, 2,3-butanediol, phenylethyl alcohol, butanedioic acid diethyl ester, ethyl hydrogen succinate, 2-hydroxy-ethyl propanoic acid and 4-hydroxy-benzeneethanol were main aroma components, these volatile componds provided the typical sensory style of wine. The MLF by commercial starer cultures and No.N7-03 only had flaver modification effect.By comparison flaver modification effect after MLF by No.N7-03 and commercial starer cultures, the results showed that there were slightly differences in aroma components and contents after MLF. The wine sensory style after MLF were also similar. These results also indicated that No.N7-03 had commercially potentiality in wine-making.