The evaluation of lysozyme under winemaking conditions

Wassung, Riaan Frederick (2006-12)

Thesis (MScAgric)--Stellenbosch University, 2006.

Thesis

ENGLISH ABSTRACT: The use of lysozyme in oenology is a relative new concept. It forms with bacteriocins and glucose oxidase part of biopreservation of wine. Biopreservation refers to natural substances contributing to the stability of food and beverages. Lysozyme was approved in 2000 by the Office International de la Vigne et du Vin (OIV) for usage in wine. Lysozyme is commercially extracted from hen egg white and inhibits Gram-positive microorganisms such as lactic acid bacteria (LAB). Lysozyme's mode of action is through degradation of peptidoglycan and cleaves the p-(1-4)-glycosidic bond between N-acetylmuramic acid (MurNAc) and N-acetyl-D-glucosamine (GlcNAc) in the polysaccharide, which form peptidoglycan, the essential constituent of bacterial cell walls. The control of LAB during winemaking is especially important due to spoilage LAB that can produce biogenic amines, off-flavours, volatile acidity, ropiness, ethyl carbamate, bitterness, mannitol, geranium tone and they can also play a role in stuck alcoholic fermentation. One of the primary agents used for the control of microorganisms is sulphur dioxide (S02) due to its anti-oxidative and antimicrobial functions. However, the action of S02 is pH dependant and consumer demand has increased for lowered S02 levels in wine. This study therefore focused on the evaluation of lysozyme under South African winemaking conditions by investigating the influence of lysozyme on different strains of LAB and the effect on the alcoholic fermentation tempo (AFT) during small-scale fermentations. Secondly the effect of lysozyme on acetic acid bacteria (AAB) and LAB numbers during Pinotage, Cabernet Sauvignon and Shiraz red wine vinifications were evaluated. Other wine parameters monitored in the red wine vinifications included volatile components, biogenic amine levels, colour and total phenol content. It was shown that lysozyme was effective in lowering, or completely inhibiting, LAB growth with the exception of a few strains, thus indicating differences in sensitivity towards lysozyme between species and strains. Sensitivity to lysozyme treatments was seen for strains of L. nagelii, L. pentosus, L. vermiforme, L. paraplantaum, L. hilgradii, L. plantarum, L. paracasei and L. buchneri. Strains that did not show positive growth during the alcoholic fermentation (AF) included those of L. plantarum, L. fermentum, Pediococcus acidilactici, L. pentosus and Leuconostoc mesenteroides. This study showed that wine isolated Lactobacillus strains of L. brevis, L. buchneri and L. paracasei are more resistant to lysozyme concentrations during a controlled small-scale AF. No effect was observed for AFT. Furthermore, lysozyme treatment did not result in an increase in AAB growth during AF, however, in some cases resulted in lower AAB numbers for lysozyme treated wines. Differences in LAB and AAB numbers could be seen between the tested cultivars. No conclusion could be drawn from this study on the effect on the volatile compounds and biogenic amine and needs further investigation. Lysozyme treatment did not have any effect on colour or total phenol content of red wine.

AFRIKAANSE OPSOMMING: Die gebruik van lisosiem in wynkunde is 'n redelike nuwe konsep. Lisosiem vorm saam met bakteriosiene en glukose oksidase deel van biopreservering strategiee vir wyn. Biopreservering verwys na natuurlike middels wat bydra tot die stabiliteit van voedsel- en drankprodukte. Lisosiem is in 2000 goedgekeur deur die Office International de la Vigne et du Vin (OIV) vir die gebruik in wyn. Kommersieel word lisosiem uit eierwit geekstraheer en gebruik om Gram-positiewe organismis soos melksuurbakteriee (MSB) te inhibeer. Lisosiem se werking is deur die afbraak van peptidoglukaan en breek die 13-(1-4)glikosidiese verbinding tussen N-asetiel muramiensuur (MurNAc) en N-asetiel-Dglukosamien (GlcNAc) in die polisakkaried wat peptidoglukaan vorm, die noodsaaklike bousteen waaruit bakteriele selwande bestaan. Die beheer van MSB in wyn is veral belangrik vanwee die vermoe van bederf MSB om biogene amiene, afgeure, vlugtige suur, draadagtigheid, etielkarbamaat, bitterheid, mannitol en malva-agtigheid te vorm en MSB kan ook 'n rol speel by slepende/gestaakte alkoholiese 'gistings. Een van die belangrikste middels om mikroorganismes te beheer is swaweldioksied (S02) vanwee die antioksiderende en antimikrobiese werking van S02. Die werking van S02 is pH afhanklik en hedendaagse verbruikerstendense neig na verlaagde S02 vlakke in wyn. Hierdie studie het dus gefokus op die evaluasie van lisosiem onder Suid-Afrikaanse wynmaak toestande deur die invloed van lisosiem op verskillende rasse van MSB en gevolglik op die alkoholiese fermentasie tempo (AFT) te evalueer tydens kleinskaalse fermentasies. Tweedens is die effek van lisosiem op asynsuurbakteriee (ASB) en MSB getalle tydens Pinotage, Cabernet Sauvignon en Shiraz rooiwynmakery getoets. Ander parameters wat ook gemonitor is sluit vlugtige komponente, biogeniese amien vlakke, kleur en totale fenol vlakke in. Die studie het getoon dat lisosiem effektief was in die verlaging of algehele inhibisie van MSB groei met die uitsluiting van 'n paar rasse wat dus wys op sensitiwiteits verskille tussen rasse en spesies. Sensitiwiteit vir lisosiem behandeling is gesien vir rasse van L. nagelii, L. pentosus, L. vermiforme, L. paraplantarum, L. hilgardii, L. plantarum, L. paracasei en L. buchneri. Rasse wat nie positiewe groei getoon het tydens die alkoholiese fermentasie nie sluit die van L. plantarum, L. fermentum, Pediococcus acidilactici, L. pentosus en Leuconostoc mesenteroides. Hierdie studie het getoon dat meestal wynge·isoleerde rasse van Lactobacil/us in L. brevis, L. buchneri en L. paracasei meer weerstandbiedend is teen lisosiem toevoegings gedurende 'n beheerde kleinskaalse fermentasie. Geen effek op die AFT is waargeneem nie. Verder het lisosiem toediening nie gelei tot 'n verhoging in ASB getalle tydens alkoholiese fermentasie nie en wel in sekere gevalle laer ASB getalle vir lisosiem behandelde wyne getoon. Verskille in terme van MSB en ASB getalle kon gesien word tussen kultivars. Geen gevolgtrekking kon gemaak word in terme van die invloed van lisosiem op die vlugtige komponente en biogeniese amiene en benodig verdere navorsing. Lisosiem toediening het geen effek gehad op kleur en totale fenol inhoud van rooiwyn nie.

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