Thesis (MSc)--University of Stellenbosch, 2002. / ENGLISH ABSTRACT: The production of wine is a complex process, which involves the conversion of sugar in
grape must to ethanol, carbon dioxide and other byproducts. The principal organism in
winemaking is yeast, of which Saccharomyces cerevisiae is the most important due to
its ability to survive winemaking conditions, its GRAS (Generally Regarded As Safe)
status and the favourable flavours it imparts during the winemaking process. However,
due to the demands of the consumer and the emergence of sophisticated wine markets,
a demand is developing for specialised yeast strains with enhanced and new
oenological properties. For these reasons, research into the contribution of wine yeast
to the aroma bouquet as well the influence of wine or brandy maturation in wood on the
aroma bouquet is important for consumer demands to be met.
The fruity aroma of wine is associated with esters, which are produced during the
alcoholic fermentation by yeast. Important acetate esters in wine and brandy are ethyl
acetate, which has a fruity, solvent-like aroma, and isoamyl acetate, which has a
banana-like aroma. These esters are produced through the action of acetyltransferases
(AATases), which catalyse the reaction between a higher alcohol and acyl Coenzyme A.
Esters are mainly a product of alcoholic fermentation. However, their concentration
changes during wood maturation and it has been found that the concentration of acetate
esters can increase during the maturation period.
In this study, the aim was to investigate the influence of AATase I and AATase II,
which are encoded by the ATF1 and ATF2 genes respectively, on the aroma bouquet of
wine and brandy. Therefore, the first objective of this study was to clone the ATF2 gene
from a commercial wine yeast strain and to overexpress this gene in a commercial wine
yeast strain and in a wine yeast strain that already has the A TF1 gene overexpressed.
Disruption cassettes were also designed in order to disrupt the ATF1 and ATF2 genes in
a commercial wine yeast strain. The resultant recombinant wine yeast strains were
used for the production of wine and brandy. GC analyses and tasting trials were
conducted to determine the effect of the overexpression or disruption of these genes on
the aroma bouquet of wine.
The results obtained indicated that there are differences in the aroma bouquet of
wine and brandy when changes are made in gene expression. The results indicated
that the A TF1 gene plays a large role in the production of ethyl and isoamyl acetate.
When this gene was overexpressed, the level of ethyl acetate was 5.6-fold more than
that of the control and the level of isoamyl acetate was 3.5-fold higher than that of the
control. However, no increase in ethyl acetate or isoamyl acetate was observed when
the A TF2 gene was overexpressed. An increase in 2-phenylethyl acetate and diethyl
succinate was observed in brandy, although there was a decrease in total ester
concentration. A decrease in acetic acid was also observed in the brandy produced,
which could be an indication of ester production. Similarly, no increase in ethyl acetate or isoamyl acetate was observed in the wine or brandy produced when both the ATF1
and ATF2 genes were overexpressed in a single yeast. Once again, a marked decrease
was observed in acetic acid concentration in both the wine and brandy.
In conclusion, it is clear that changes in gene expression can change the aroma
profile of wine or brandy. However, the role of the ATF2 gene still remains unclear and
further studies are needed to clarify its role in yeast. Future studies involving the effect
of wood maturation on ester concentration will also be of importance, so that the
winemaker or distiller can make a product that suits the ever-changing market. / AFRIKAANSE OPSOMMING: Die produksie van wyn is 'n komplekse proses wat die omskakeling van die suiker in
mos tot etanol, koolstofdioksied en ander byprodukte tot gevolg het. Die hooforganisme
betrokke in die wynmaakproses is gis, waarvan Saccharomyces cerevisiae as een van
die belangrikste geag word as gevolg van die vermoë daarvan om onder die
wynfermentasietoestande te kan oorleef, die "GRAS"-status (Generally Regarded As
Safe) daarvan en die invloed wat dit op die aroma van die uiteindelike produk het weens
die werking daarvan gedurende alkoholiese fermentasie. Die behoefte aan wyn met
nuwe, verbeterde eienskappe het die vraag na meer gespesialiseerde gisrasse deur
beide die verbruiker en nuwe wynmarkte gedurende die afgelope paar jaar drasties laat
toeneem. Dit is om dié redes dat navorsing oor die bydrae van wyngis en
houtveroudering tot die aroma van beide wyn en brandewyn so belangrik geag word.
Die vrugtige aroma van wyn word geassosieer met die esters wat gedurende die
alkoholiese fermentasie deur gis gevorm word. Die belangrikste asetaatesters in wyn en
brandewyn is etielasetaat, wat vir 'n oplosmiddelagtige, vrugtige aroma bekend is, en
isoamielasetaat, wat 'n piesangaroma veroorsaak. Die esters word geproduseer deur
die werking van asetieltransferases (AATases), wat as katalis in die reaksie tussen 'n
hoër alkohol en asetiel-Ko-ensiem A optree. Alhoewel esters hoofsaaklik 'n produk van
alkoholiese fermentasie is, wissel die konsentrasie daarvan gedurende houtveroudering.
Daar is gevind dat die konsentrasie van die asetaatesters gedurende die
verouderingsproses kan verhoog.
Die studie het ten doelom die invloed van AATase I en AATase II, wat
onderskeidelik deur die ATF1- en ATF2-gene geënkodeer word, op die aroma van wyn
en brandewyn te ondersoek. Die eerste doelwit van die studie was vervolgens om die
ATF2-geen vanaf 'n kommersiële wyngisras te kloneer en dit daarna te ooruitdruk in 'n
kommersiële wyngisras, asook die geen te ooruitdruk in 'n kommersiële wyngisras wat
reeds die ATF1-geen ooruitdruk. Disrupsiekassette is ook vir die disrupsie van die
ATF1- en ATF2-gene in 'n kommersiële wyngisras ontwerp. Die rekombinante
wyngisrasse wat gedurnde die studie gemaak is, is vir die produksie van wyn en
brandewyn gebruik. Gas chromatografise-ontledings en sensoriese evaluerings is ook
op die wyn en brandewyn uitgevoer.
Die resultate van die studie het bewys dat daar wel veranderings plaasvind
wanneer 'n verandering in geenuitdrukking gemaak is. Die resultate het weereens
bevestig dat die ATF1-geen 'n belangrike rol in die produksie van etiel- en isoamielasetaat
speel. Wanneer die ATF1-geen ooruitgedruk is, is die etielasetaatproduksie 5.6
keer meer en die isoamielasetaatproduksie 3.5 keer meer as in die kontrole. Die
ooruitdrukking van die ATF2-geen het geen verhoging in etielasetaat of isoamielasetaat
of in totale esters in die wyn getoon nie, alhoewel die ras 2.7 keer meer diëtielsuksinaat
geproduseer het. In die brandewyn wat geproduseer is met die gisras waarin ATF2 ooruitgedruk is, was daar wel 'n verlaging in die asynsuur, wat 'n aanduiding van
estervorming kan wees, alhoewel die totale esters wat geproduseer is minder was as in
die kontrole. 'n Verhoging in diëtielsuksinaat en 2-fenielasetaat is ook gevind. Daar is
geen verhoging in etiel- of isoamielasetaat getoon wanneer die ATF1- en ATF2-geen
saam ooruitgedruk is nie. Die ras het minder totale sure in wyn en brandewyn
geproduseer en ook geen verhoging in totale esters getoon nie.
Uit die resultate is dit duidelik dat veranderings in geenuitdrukking 'n verandering
in die aromaprofiel van wyn en brandewyn kan veroorsaak. Die rol van dié A TF2-geen
is nog steeds onduidelik en verdere studies sal moet plaasvind om die rol van die geen
te verduidelik. Studies wat konsentreer op die invloed van houtveroudering op
esterkonsentrasie is ook belangrik vir die toekoms, want dit sal die wyn- of
brandewynmaker meer beheer oor die uiteindelike produk gee en daardeur die wyn- of
brandewynmaker help om 'n produk te vervaardig wat sy mark bevredig.
| Identifer | oai:union.ndltd.org:netd.ac.za/oai:union.ndltd.org:sun/oai:scholar.sun.ac.za:10019.1/52926 |
| Date | 12 1900 |
| Creators | Bayly, Jennifer Carr,1977- |
| Contributors | Du Toit, M., Pretorius, I. S., Lambrechts, M. G., Stellenbosch University. Faculty of AgriSciences. Dept. of Viticulture and Oenology. Institute for Wine Biotechnology. |
| Publisher | Stellenbosch : Stellenbosch University |
| Source Sets | South African National ETD Portal |
| Language | en_ZA |
| Detected Language | English |
| Type | Thesis |
| Format | 79 p. : ill. |
| Rights | Stellenbosch University |
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