Post-alcoholic fermentation micro-ecology of wines

: the role of phenolic composition and the effects on quality

Abstract

The research presented in this thesis explores the interactions between certain phenolic compounds that are naturally present in wines and the microorganisms that are generally present in this medium.The interactions studied include the effects on microbial growth and metabolism, the intra-specific diversity of Oenococcus oeni and the evolution of volatile and non-volatile compounds during malolactic fermentations (MLF) and subsequent storage. The ability of several strains of lactic acidbacteria (LAB) to release hydroxycinnamic acids (HCA) from their tartrate derivative forms((hydroxy)cinnamoyl-tartaric acids) was also studied. Metabarcoding analysis of 16 different wines analyzed at the post-malolactic stage was performed using sequence data from 16S (for bacteria) and ITS2 (for fungi) amplicons. Similar patterns were observed in all wines at this level of discrimination, with Saccharomyces cerevisiae yeasts andbacteria from the genera Acetobacter, Gluconobacter and Swaminathania being the most abundant taxa. Concerning the profiles of phenolics and volatiles, red and white wines were grouped separately, and French and Spanish wines tended to cluster together.The effects of kaempferol, trans-caffeic and trans-caftaric acids (added at 10 mg/L) on the microbialgrowth and metabolism were explored in non-inoculated and inoculated wine (O. oeni) and wine mixed with MRS (de Man, Rogosa & Sharpe). The strongest impact on microbial growth, malolacticand general metabolic activity in LAB, was noted for kaempferol, with trans-caftaric acid showing the weakest. The effects of each phenolic compound varied according to the medium used, the type of MLF (inoculated or not) and the O. oeni strain inoculated (OenosTM or CH35TM).Using concentrations normally encountered in wines as a reference, flavan-3-ols ((+)-catechin and(-)-epicatechin), flavonols (kaempferol and quercetin), HCA (trans-p-coumaric and trans-ferulicacids) and trans-resveratrol were studied in experiments with non-inoculated wines and wine sinoculated with OenosTM. Depending on its concentration, (+)-catechin positively impacted the yeast population, activated or delayed malic acid degradation or inhibited the growth of bacteria. All phenolics tested, at all concentrations tested, delayed citrate consumption in inoculated samples.The effect of flavonols and trans-resveratrol on the O. oeni diversity was dependent on the strainsunder study. An increase in trans-p-coumaric and trans-ferulic acid levels appeared to induce the release of trans-caffeic acid from one of its precursors, possibly via an increase in cinnamoyl esteraseactivity of some strains. Flavan-3-ols and flavonols inhibited the growth of these bacteria during storage. The phenolics tested affected specific enzymatic systems responsible for the production and degradation of important metabolites involved in the organoleptic quality of the wines. Cinnamoyl esterase (CE) activity of wine microbes can be relevant as it confers the capacity tomodulate the phenolic acid composition of a wine by liberating these from otherwise see minglybiologically unavailable, tartrate derivatives. Five commercial strains of O. oeni were studied in this respect, three exhibiting CE activity (OenosTM, CiNeTM and CH35TM) and two not (CH16TM andCH11TM). CE activity was detected in cell-free extracts of the three CE positive (CE+) strains and one of the CE negative (CE-) strains. From comparative genome analysis, no gene exclusive to the 3 CE+ was detected and the inference that membrane transport differences might be behind differences in CE activity was also not conclusively supported. This hypothesis needs to be further explored as does the possibility of the involvement of wine molecules and of more than one enzymein the CE activity.

Date of Award	25 Oct 2019
Original language	English
Awarding Institution	Universidade Católica Portuguesa
Supervisor	Timothy Hogg (Supervisor) & Francisco Manuel Morais Sarmento de Campos (Co-Supervisor)