Evaluating the influence of winemaking practices on biogenic amine production by wine ...

172 Pages·2008·1.76 MB·English

by  S Baard

Evaluating the influence of
winemaking practices on biogenic
amine production by wine
microorganisms

by

Anita Yolandi Smit

Thesis presented in partial fulfilment of the requirements for the degree of
Master of AgriSciences at Stellenbosch University.

December 2007

Supervisor:
Dr. M. du Toit

Co-supervisor:
Dr. W.J. du Toit
DECLARATION

I, the undersigned, hereby declare that the work contained in this thesis is my own
original work and that I have not previously in its entirety or in part submitted it at any
university for a degree.

____________________ ______________
Anita Yolandi Smit Date

Copyright ©2007 Stellenbosch University
All rights reserved
SUMMARY
Biogenic amines are nitrogenous compounds of low molecular weight found in most
fermented foods, including wine. These biologically produced amines are essential at
low concentrations for normal metabolic and physiological functions in animals, plants
and micro-organisms. However, biogenic amines can have adverse effects at high
concentrations and pose a health risk for sensitive individuals. Symptoms include
nausea, hot flushes, headaches, red rashes, respiratory distress and fluctuations in
blood pressure. A number of countries have implemented upper limits for histamine in
food and wine. This development has already started to threaten commercial export
transactions and may become more serious in the near future, especially in the
competitive wine industry of today. The most important biogenic amines in wine include
histamine, tyramine, putrescine, cadaverine and phenylethylamine which are produced
from the amino acids histidine, tyrosine, ornithine, lysine and phenylalanine
respectively.
Biogenic amines are mainly produced in wine by microbial decarboxylation of the
corresponding precursor amino acid. It may be produced by yeast during alcoholic
fermentation, by lactic acid bacteria during malolactic fermentation, or potentially by
spoilage microbes such as acetic acid bacteria and Brettanomyces. However, lactic acid
bacteria are widely accepted as the main causative agents.
Inoculation with commercial malolactic fermentation starter cultures that do not
possess the relevant decarboxylase genes may inhibit the growth and activity of
decarboxylase positive indigenous bacteria and as such control the production of
biogenic amines in wine. In this study it was shown that co-inoculation of malolactic
starter cultures together with alcoholic fermentation could reduce the incidence of
biogenic amines in wine compared to conventional inoculation protocols; presumably
because undesirable activities were restrained at an earlier stage during co-inoculation.
It was also indicated in this work that in some cases the effect of co-inoculation on
biogenic amine reduction may only be visible after a period of ageing. The frequency of
biogenic amine occurrence in wines aged for a short period was generally higher in the
presence of fermentation lees than in its absence.
This work also included a preliminary investigation into the contribution of
commercial wine yeast starter cultures to biogenic amine production. Diamines and
polyamines (putrescine, spermidine and cadaverine) were produced to variable extents
by all yeasts with very little or no production of physiologically important biogenic
amines (histamine, tyramine and phenylethylamine).
Another objective of this study was to evaluate the influence of common
winemaking practices on biogenic amine production under winemaking conditions. We
have shown that biogenic amine production by lactic acid bacteria could be influenced,
amongst others, by the presence of precursor amino acids in the grape must or wine,
the time of contact between juice or wine and grape skins, the time of contact between
wine and yeast lees, the presence of microbial nutrients, wine pH, sulphite and ethanol
levels, the phenolic composition of the wine and the number of decarboxylase positive
lactic acid bacteria present in the wine.
Lately, the wine industry is under increasing pressure to increase measures to
ensure food safety and security and to eliminate any compound, present even in trace
amounts that could reduce the wholesomeness of the wine. The need arises for a rapid
and inexpensive method for quality control. In this study we investigated the potential to
use Fourier transform infrared spectroscopy to rapidly screen for the presence of
elevated levels of biogenic amines. This presents a novel method for the detection and
quantification of total biogenic amines in wines.
OPSOMMING
Biogeenamiene is stikstofbevattende verbindings met ‘n lae molekulêre gewig wat in
meeste gefermenteerde voedselsoorte, insluitend wyn, voorkom. Hierdie biologies
aktiewe amiene is noodsaaklik teen lae konsentrasies vir nomale metabolisme en
fisiologiese funksies in diere, plante en mikroörganismes.
Biogeen amiene kan egter nadelige gevolge hê wanneer dit ingeneem word teen
hoë konsentrasies en kan ‘n ernstige gesondheidsrisiko vir sensitiewe individue inhou.
Algemene simptome van biogeenamiene sluit naarheid, warm gloede, hoofpyn, ‘n rooi
uitslag, respiratoriese afwykings en fluktuasies in bloeddruk in. ‘n Aantal lande het reeds
maksimum limiete vir die konsentrasie van histamien teenwoordig in voedsel en wyn
geïmplimenteer. Hierdie ontwikkeling het alreeds begin om kommersiële
uitvoertransaksies te bedreig en mag in erns toeneem in die nabye toekoms, veral
weens die kompeterende aard van die hedendaagse wynindustrie. Die mees belangrike
biogeenamiene in wyn sluit in histamien, tiramien, putresien, kadawerien and
fenieletielamien, wat onderskeidelik geproduseer word vanaf die aminosuur voorlopers
histidien, tirosien, ornitien, lisien en fenielalanien.
In wyn word biogeenamiene hoofsaaklik geproduseer deur die mikrobiese
dekarboksilasie van die ooreenstemmende aminosuurvoorloper. Biogeenamiene kan
deur gis geproduseer word tydens alkoholiese fermentasie, deur melksuurbakterieë
gedurende appelmelksuurgisting of potensieël deur bederforganismes soos
asynsuurbakterieë en Brettanomyces. Melksuurbakterieë word egter wyd aanvaar as
die hoofbron van biogeen amien produksie.
Inokulasie van wyn vir appelmelksuurgisting met kommersiële aanvangskulture wat
nie die relevante dekarboksilase gene besit nie kan die groei en aktiwiteit van
dekarboksilase positiewe inheemse melksuurbakterieë inhibeer en sodoende die
probleem van biogeenamiene in wyn beheer. In hierdie werk het ons gewys dat die ko-
inokulasie van appelmelksuurgisting met alkoholiese fermentasie die voorkoms van
biogeen amiene kan verlaag in vergelyking met konvensionele inokulasie-metodes;
waarskynlik omdat ongewenste aktiwiteit reeds op ‘n vroeë stadium uitgeskakel word
met ko-inokulasie. Dit is ook in hierdie werk gewys dat die effek van ko-inokulasie op
biogeen amien vermindering soms eers na ‘n verouderingsperiode waarneembaar is.
Die voorkoms van biogeen amiene in wyne wat vir ‘n kort tydperk veroudering
ondergaan het was oor die algemeen hoër in die teenwoordigheid van gismoer as in die
afwesigheid daarvan.
‘n Voorlopige ondersoek in die bydrae wat kommersiële wyngisaanvangskulture tot
biogeen amien produksie kan lewer word ook in die werk vervat. Diamiene en
poliamiene (putresien, spermidien en kadawerien) is in wissellende vlakke deur alle
giste geproduseer, met min tot geen produksie van fisiologies belangrike
biogeenamiene (histamien, tiramien en fenieletielamien).
Nog ‘n oogmerk van hierdie studie was om die invloed van algemeen toegepaste
wynmaakpraktyke op biogeen amien produksie onder wynmaakkondisies te evalueer.
Ons het gewys dat biogeen amien produksie deur melksuurbakterieë onder meer
beïnvloed kan word deur die teenwoordigheid van voorloper aminosure in die
druiwemos of wyn, die kontaktyd tussen sap of wyn en die druiwedoppe, die kontaktyd
tussen die wyn en die gismoer, die teenwoordigheid van mikrobiese voedingstowwe,
wyn pH, sulfiet- en etanolvlakke, die fenoliese samestelling van die wyn en die aantal
dekarboksilase positiewe melksuurbakterieë teenwoordig in die wyn.
Die wynindustrie is tans onder toenemende druk om maatstawwe in plek te stel om
voedselveiligheid en -sekuriteit te verseker, en om die teenwoordigheid van enige
komponent wat die heilsaamheid van wyn kan verminder te beperk, selfs in
spoorhoeveelhede. Die behoefte ontstaan dus vir ‘n vinnige en goedkoop metode om
kwaliteitskontrole toe te pas. In hierdie studie het ons ondersoek ingestel om Fourier-
transformasie-infrarooi-spektroskopie te gebruik as vinnige siftingsmetode om hoë
vlakke van biogeen amiene in wyn aan te dui. Hiermee rapporteer ons dan ook ‘n nuwe
metode om totale biogeen amiene in wyn op te spoor en te kwantifiseer.
This thesis is dedicated to my family
Hierdie tesis is opgedra aan my familie
BIOGRAPHICAL SKETCH
Anita Smit was born on 25 March 1983 and lived for most of her childhood in George.
She matriculated at Oakhill School, Knysna in 2001. Anita obtained her BScAgric
degree (Oenology Specialised) at the Stellenbosch University in 2005. In 2006, she
enrolled at the same university for a Masters degree in Oenology.
ACKNOWLEDGEMENTS

I wish to express my sincere gratitude and appreciation to the following persons and
institutions:

DR. M DU TOIT, (Institute for Wine Biotechnology, Department of Viticulture and
Oenology, Stellenbosch University) who acted as supervisor and provided endless
patience, encouragement, support and motivation, for the opportunity to be one of her
students, and for the critical evaluation of this manuscript;

DR. W.J. DU TOIT, (Department of Viticulture and Oenology, Stellenbosch University)
for his interest in my work, for very valuable insights into the winemaking process and
the chemistry of wine, and for the critical evaluation of this manuscript;

FELLOW COLLEAGUES, (Institute for Wine Biotechnology, Department of Viticulture
and Oenology, Stellenbosch University) for providing valuable training, advice and
assistance in the laboratory;

THE NATIONAL RESEARCH FOUNDATION, WINETECH, THRIP, POSTGRADUATE
MERIT BURSARY AND HARRY CROSSLEY FOUNDATION, for financial support;

ACHOR YEAST, LALLEMAND AND CHR. HANSEN, for providing commercial
products;

DR. M.A. STANDER, (Central analytical facility, Stellenbosch University) for her
enthusiasm to develop new analytical methods and for sample analyses;

CHEMOMETRICS GROUP, (Institute for Wine Biotechnology, Department of Viticulture
and Oenology, Stellenbosch University) for assistance with multivariate data analysis;

CELLAR STAFF, (Experimental cellar, Department of Viticulture and Oenology,
Stellenbosch University) for assistance and support in the cellar;

MY PARENTS AND SISTER, Johann, Marietjie and Elzaan for their continuous
encouragement, motivation and understanding;

MY CLOSE FRIENDS, for keeping my life in balance; and

GOD ALMIGHTY, for blessing me with a passion for this work.
PREFACE

This thesis is presented as a compilation of seven chapters. Each chapter is introduced
separately and is written according to the style of the South African Journal of Enology
and Viticulture.

Chapter 1 GENERAL INTRODUCTION AND PROJECT AIMS

Chapter 2 LITERATURE REVIEW
Biogenic amines in wine: Understanding the headache

Chapter 3 RESEARCH RESULTS
Evaluating the influence of malolactic fermentation inoculation times and
ageing on lees on the production of biogenic amines by wine lactic acid
bacteria and preliminary quantification of biogenic amines by Fourier
transform infrared spectroscopy

Chapter 4 RESEARCH RESULTS
Evaluating the influence of maceration practices on biogenic amine
production by wine lactic acid bacteria

Chapter 5 RESEARCH RESULTS
Evaluating the influence of complex commercial yeast and bacterial
nutrients on biogenic amine production by wine microorganisms

Chapter 6 RESEARCH RESULTS
Preliminary screening of commercial yeast starter cultures for biogenic
amine production

Chapter 7 GENERAL DISCUSSION AND CONCLUSIONS