The selection and characterisation of lactic acid bacteria to be used as a mixed starter culture for malolactic fermentation

[摘要] ENGLISH ABSTRACT: The quality of wine is influenced and determined by various factors, one of which includes theprocess of malolactic fermentation (MLF). MLF plays an integral role in the flavour and sensoryprofile of most red wines as well as some white wines like Chardonnay. This process is conductedby lactic acid bacteria (LAB), specifically of the genera Oenococcus, Lactobacillus, Pediococcusand Leuconostoc. Of these, Oenococcus oeni is best adapted to survive in the harsh wineenvironment.MLF is defined as the conversion of L-malic acid to L-lactic acid and carbon dioxide. Theconversion of the dicarboxylic malic acid to the monocarboxylic lactic acid results in a decrease inacidity and an increase in pH, to give a softer mouthfeel and more favourable flavour profile. Afurther reason for conducting MLF in wine includes the improvement of microbial stability due tothe removal of malic acid as a possible substrate for microorganisms. Recently, research focus hasshifted to the ability of MLF and LAB to alter the aroma profile of wine via the production and/ormodification of certain aroma compounds.In order for wine LAB to conduct MLF, they need to be able to survive the harsh andchallenging wine environment. Conditions in South African wines are particularly challenging dueto the long, hot ripening seasons resulting in high sugar concentrations which give high ethanolconcentrations. Some LAB also struggle to adapt to an environment with high pH and low malicacid concentrations. These factors, combined with the use of sulphur dioxide, cause LAB tostruggle in conducting and completing successful MLF. Many of the commercial starter culturesthat are currently available contain LAB that have not been isolated from South African wine andare therefore not optimal for use under these challenging wine conditions. Oenococcus oeni is alsothe single LAB culture present in all commercially available starter cultures.The overriding goal of this study was to create a MLF starter culture containing a mixture ofLAB cultures, namely O. oeni and Lactobacillus plantarum, which can successfully convert malicacid to lactic acid, ensure microbial stability, but also make a positive contribution to the winearoma profile. Lactobacillus plantarum has previously been considered for possible use in acommercial starter culture. The LAB isolates used in this study were selected from the Institute forWine Biotechnology culture collection as well as isolated from spontaneous MLF.The first objective was to characterise these LAB strains for important traits and for possibleuse as a MLF starter culture. A total of 23 strains were identified as O. oeni and 19 strains asLb. plantarum. The identified strains were screened in a synthetic wine medium for their ability toconvert malic acid to lactic acid. Based on the LAB strain performance in the synthetic winemedium, seven strains of both O. oeni and Lb. plantarum were selected. These 14 strains werescreened for the presence of genes encoding for enzymes responsible for biogenic amineproduction and were found to contain none of the genes associated with the formation ofhistamine, tyramine or putrescine. The LAB strains were genetically screened for enzymes associated with aroma modification by LAB during MLF. The enzymes of interest that werescreened for included β-glucosidase, esterase, protease and phenolic acid decarboxylase (PAD).The Lb. plantarum strains were found to possess more diverse enzymatic profiles related to aromathan O. oeni. The biggest differences were observed for the presence of β-glucosidase and PAD.The second objective was to perform small-scale fermentations with the individual LABisolates. The individual isolates were evaluated in Pinotage and based on these results; threestrains of each O. oeni and Lb. plantarum were selected for evaluation in mixed culturefermentations. The mixed cultures were evaluated in Pinotage, Shiraz and Cabernet Sauvignon inthe 2008 vintage. As a third objective, the wines were also analytically and sensorially evaluated toinvestigate the changes in the aroma profile that could be attributed to the presence of the mixedLAB isolates. Based on the fermentation data as well as data pertaining to the aroma modification,three mixed cultures were selected for evaluation in the 2009 vintage in Pinotage, CabernetSauvignon and Chardonnay. The mixed cultures were able to successfully complete MLF infermentation periods comparable to that of a commercial culture used as control. The different LABcultures had distinct and diverse effects on the wine aroma profile. The O. oeni strain played alarger role in the ester concentration present after MLF, while the Lb. plantarum strain had a largereffect on the higher alcohol and volatile fatty acid concentration upon completion of MLF.The results generated by this novel study clearly indicate the potential of a mixed LAB starterculture for conducting MLF. The mixed cultures successfully completed MLF and made a positivecontribution to the wine aroma profile.

[发布机构] Stellenbosch University