Thesis (MScFoodSc)--Stellenbosch University, 2004. / ENGLISH ABSTRACT: The regular consumption of probiotics is becoming a recognized trend in the food industry due to several reported health benefits. A probiotic is defined as a live microbial feed supplement that beneficially affects the host animal by improving its intestinal microbial balance. A wide variety of probiotic food products are available on the South African market and comprise an assortment of fermented milks, as well as lyophilized preparations in tablet or capsule form. Strains of Lactobacillus acidophilus and Bifidobacterium species are mostly used as probiotic microbes in the industry due to their health enhancing effect. The survival of sensitive probiotic microbial species in food matrices are influenced by various factors such as oxygen concentration, pH levels and manufacturing and storage conditions. These should be considered and monitored as the South African food and health regulations stipulate that probiotic microbes should be present at a concentration of 10⁶ cfu.ml ̄ ¹' in order to exert a beneficial effect. Some health benefits are also correlated to specific microbial species and strains and these factors have resulted in the need for the rapid and accurate identification of probiotic microbes present in food products. The probiotic microbes present in probiotic yoghurts and supplements have in the past been identified using traditional methods such as growth on selective media, morphological, physiological and biochemical characteristics. However, even some of the most sophisticated cultural-dependant techniques are not always sufficient for the identification and classification of especially Bifidobacterium, as well as closely related Lactobacillus species. Molecular techniques are more often employed for the rapid and accurate detection, identification and characterization of microbial species present in food products. The aim of this study was to detect and identify the probiotic species present in various commercial South African yoghurts and lyophilized preparations using peR-based DGGE analysis. A 200 bp fragment of the V2-V3 region of the 16S rRNA gene was amplified and the peR fragments were resolved by DGGE. The unique fingerprints obtained for each product were compared to two reference markers A and B in order to identify the bands present. The results obtained were verified by species-specific peR, as well as sequence analyses of bands that could not be identified when compared to the reference markers. Only 54.5% of the South African probiotic yoghurts that were tested did contain all the microbial species as were mentioned on the labels of these products, compared to merely one third (33.3%) of the lyophilized probiotic food supplements. Some Bifidobacterium species were incorrectly identified according to some product labels, while other products contained various microbes that were not mentioned on the label. Sequence analysis confirmed the presence of a potential pathogenic Streptococcus species in one of the yoghurt products and in some instances the probiotic species claimed on the labels were non-scientific and misleading. The data obtained in this study showed that the various South African probiotic products tested were of poor quality and did not conform to the South African regulations. peR-based DGGE analysis proofed to be a valuable approach for the rapid and accurate detection and identification of the microbial species present in South African probiotic products. This could help with future implementation of quality control procedures in order to ensure a reliable and safe probiotic product to the consumer. / AFRIKAANSE OPSOMMING: Die gereelde inname van probiotiese produkte is besig om In erkende tendens in die voedselindustrie te word, as gevolg van verskeie gesondheidsvoordele wat daaraan gekoppel word. In Probiotika word gedefinieer as In voedingsaanvulling wat uit lewendige mikrobes bestaan en wat In voordelige effek op mens of dier het deur In optimale mikrobiese balans in die ingewande te handhaaf. In Wye verskeidenheid probiotiese voedselprodukte is tans beskikbaar op die Suid- Afrikaanse mark. Hierdie bestaan hoofsaaklik uit verskeie gefermenteerde melkprodukte asook 'n reeks tablette en kapsules wat probiotiese mikrobes in gevriesdroogde vorm bevat. Lactobacillus acidophilus tipes en Bifidobacterium spesies word die algemeenste in die voedselindustrie gebruik aangesien hierdie spesifieke mikrobes bekend is om goeie gesondheid te bevorder. Die oorlewing van sensitiewe probiotiese mikrobiese spesies in voedsel matrikse word beïnvloed deur faktore soos suurstof konsentrasie, pH-vlakke en vervaardigings- en opbergings kondisies. Hierdie faktore moet in aanmerking geneem word en verkieslik gemonitor word aangesien die Suid-Afrikaanse voedsel en gesondheids regulasies stipuleer dat probiotiese mikrobes teen In konsentrasie van 10⁶ kolonie vormende eenhede per ml teenwoordig moet wees om In voordelige effek te toon. Sommige gesondheidsvoordele word direk gekoppel aan spesifieke mikrobiese spesies en spesie-tipes. Hierdie faktore het gelei tot In groot aanvraag na vinnige en akkurate metodes vir die identifikasie van probioties mikrobes in voedselprodukte. Die probiotiese mikrobes teenwoordig in probiotiese joghurts en ook die gevriesdroogde vorms in tablette en kapsules, was al geïdentifiseer deur gebruik te maak van tradisionele metodes soos groei op selektiewe media, morfologiese, fisiologiese en biochemiese eienskappe. Selfs van die mees gesofistikeerde kultuur-afhanklike tegnieke is egter nie altyd voldoende vir die identifikasie en klassifikasie van veral Bifidobacterium en na-verwante Lactobacillus spesies nie. Molekulêre metodes word dikwels aangewend vir die vinnige en akkurate deteksie, identifikasie en karakterisering van mikrobes teenwoordig in voedselprodukte. Die doel van hierdie studie was om die probiotiese mikrobes teenwoordig in verskeie Suid-Afrikaanse joghurts en gevriesdroogde aanvullings, te identifiseer deur gebruik te maak van polimerase kettingreaksie (PKR)-gebaseerde denaturerende gradiënt jelelektroforese (DGGE) analise. 'n PKR fragment van 200 bp van die V2-V3 gedeelte van die 16S ribosomale RNS (rRNS) geen is geamplifiseer, en die PKR fragmente is geskei met behulp van DGGE. Die unieke vingerafdrukke wat verkry is vir elke produk is teen twee verwysings merkers A en B vegelyk om die bande teenwoordig in die profiele te identifiseer. Die resultate is bevestig deur spesies-spesifieke PKR en ook deur die ketting volgordes van die DNS fragmente te bepaal wat nie geïdentifiseer kon word deur vergelyking met die verwysings merkers nie. Slegs 54.5% van die Suid-Afrikaanse probiotiese joghurts wat getoets is het al die mikrobiese spesies bevat soos aangedui was op die etikette van hierdie produkte, teenoor slegs 'n derde (33.3%) van die gevriesdroogde voedingsaanvullings. Sekere Bifidobacterium spesies is verkeerd geïdentifiseer op sommige van die produk etikette, terwyl ander produkte verskeie mikrobes bevat het wat nie op die etiket aangedui was nie. 'n Potensiële patogeniese Streptococcus spesie is in een van die joghurt produkte gevind soos bevestig deur DNS kettingvolgorde bepalings. In sommige gevalle was die probiotiese spesienaam wat aangedui is op die etiket onwetenskaplik en misleidend. Die resultate wat uit hierdie studie verkry is dui aan dat die Suid-Afrikaanse probiotiese produkte wat getoets is van 'n swak gehalte is en nie aan die Suid- Afrikaanse regulasies voldoen nie. Daar is getoon dat PKR-gebaseerde DGGE analise 'n waardevolle tegniek kan wees vir die akkurate deteksie en identifisering van die mikrobiese spesies teenwoordig in probiotiese produkte. Dit kan help met die toekomstige implementering van kwaliteitskontrolerings prosedures om 'n mikrobiologiese betroubare en veilige produk aan die verbruiker te verseker.

Lors du développement et de l’introduction d’un nouvel aliment, la compréhension des croyances, attitudes et préférences sensorielles des consommateurs est très importante. Cette thèse présente une série d'études cross-culturelles, France vs. Vietnam, sur la représentation des consommateurs vis-à-vis de produits à base soja et sur la perception et la préférence des yaourts à base de soja, un produit dérivé du soja récemment développé pour le marché français et n'existant pas sur le marché vietnamien. Malgré une image favorable du soja sur le plan nutritionnel et sur la santé, les participants français ont rapporté ne pas consommer ce type de produit, la barrière à sa consommation étant son goût de soja. Ce résultat a été confirmé par un test de consommateurs au cours duquel les participants français ont donné une très faible évaluation hédonique pour les yaourts de soja. Les consommateurs vietnamiens, consommateurs traditionnels du soja, n'ont également pas apprécié ces produits. Une série de yaourts mixtes dans lesquels le lait de vache a été partiellement remplacé par celui de soja a été fabriquée. Les tests consommateurs ont montré que pour les deux pays, le yaourt fabriqué avec une proportion de lait de soja inférieure à 50% a reçu une note moyenne supérieure à 5 sur une échelle de 1 à 9. Le rôle de la familiarité sur les préférences alimentaires ainsi que l'impact de l'exposition culturelle sur les changements des croyances et attitudes à l'âge adulte ont été confirmés. La présente étude contribue à la caractérisation des deux cultures sur leur acceptabilité d'un nouvel aliment / Understanding consumers’ beliefs, attitudes and sensory preference is the key to a successful development and introduction of a new food. This thesis carried out a series of cross-cultural studies, France vs. Vietnam, on consumers’ representation of soy foods and on sensory perception and preference of soy yogurts, a soy derived product recently developed for French market which does not exist on the Vietnamese market. Despite a favourable image of soy in terms of nutrition and health benefices, French participants reported that the barrier to its consumption was the taste of soy. This finding was confirmed by a consumer test in which French participants gave very low hedonic evaluations to soy yogurts. Interestingly, Vietnamese consumers, soy traditional consumers, did not appreciate these products as well. A series of mix yogurts in which cow milk was partially replaced by soy milk was formulated. Consumer tests showed that for both countries, a mix yogurt with proportion of soy milk lower than 50% received an average hedonic score higher than 5 on a scale going from 1 to 9. The role of the familiarity on food preference and the impact of cultural exposure on changing in beliefs and attitudes’ change at adult age were confirmed. The present study contributes to the characterisation of the two cultures on their acceptability of a new food

Este trabalho de pesquisa descreve três estudos de utilização de métodos quimiométricos para a classificação e caracterização de óleos comestíveis vegetais e seus parâmetros de qualidade através das técnicas de espectrometria de absorção molecular no infravermelho médio com transformada de Fourier e de espectrometria no infravermelho próximo, e o monitoramento da qualidade e estabilidade oxidativa do iogurte usando espectrometria de fluorescência molecular. O primeiro e segundo estudos visam à classificação e caracterização de parâmetros de qualidade de óleos comestíveis vegetais utilizando espectrometria no infravermelho médio com transformada de Fourier (FT-MIR) e no infravermelho próximo (NIR). O algoritmo de Kennard-Stone foi usado para a seleção do conjunto de validação após análise de componentes principais (PCA). A discriminação entre os óleos de canola, girassol, milho e soja foi investigada usando SVM-DA, SIMCA e PLS-DA. A predição dos parâmetros de qualidade, índice de refração e densidade relativa dos óleos, foi investigada usando os métodos de calibração multivariada dos mínimos quadrados parciais (PLS), iPLS e SVM para os dados de FT-MIR e NIR. Vários tipos de pré-processamentos, primeira derivada, correção do sinal multiplicativo (MSC), dados centrados na média, correção do sinal ortogonal (OSC) e variação normal padrão (SNV) foram utilizados, usando a raiz quadrada do erro médio quadrático de validação cruzada (RMSECV) e de predição (RMSEP) como parâmetros de avaliação. A metodologia desenvolvida para determinação de índice de refração e densidade relativa e classificação dos óleos vegetais é rápida e direta. O terceiro estudo visa à avaliação da estabilidade oxidativa e qualidade do iogurte armazenado a 4C submetido à luz direta e mantido no escuro, usando a análise dos fatores paralelos (PARAFAC) na luminescência exibida por três fluoróforos presentes no iogurte, onde pelo menos um deles está fortemente relacionado com as condições de armazenamento. O sinal fluorescente foi identificado pelo espectro de emissão e excitação das substâncias fluorescentes puras, que foram sugeridas serem vitamina A, triptofano e riboflavina. Modelos de regressão baseados nos escores do PARAFAC para a riboflavina foram desenvolvidos usando os escores obtidos no primeiro dia como variável dependente e os escores obtidos durante o armazenamento como variável independente. Foi visível o decaimento da curva analítica com o decurso do tempo da experimentação. Portanto, o teor de riboflavina pode ser considerado um bom indicador para a estabilidade do iogurte. Assim, é possível concluir que a espectroscopia de fluorescência combinada com métodos quimiométricos é um método rápido para monitorar a estabilidade oxidativa e a qualidade do iogurte / This research work describes three studies of chemometric methods employed for the classification and characterization of edible oils and its quality parameters through Fourier Transform mid infrared spectroscopy and near infrared spectroscopy, and for the monitoring the oxidative stability and quality of yogurt using fluorescence spectroscopy . The first and second studies aimed the classification and characterization of edible oil and its quality parameters using Fourier Transform mid infrared spectroscopy (FT-MIR) and near infrared spectroscopy (NIR) measurements, respectively. Kennard-Stone algorithm was used for selecting the training set, after a principal component analysis (PCA) was applied. The discrimination of canola oils from sunflower, corn and soybean was investigated using SVM-DA, SIMCA and PLS-DA. The quality parameters refraction index and relative density of edible oil was investigated using partial least squares (PLS), iPLS, LS- SVM multivariate calibration of FT-MIR and NIR data were evaluated. Several preprocessing alternatives (first derivative, multiplicative scatter correction, mean centering, orthogonal signal correction and standard normal variate) were investigated by using the root mean square error of validation cross validation (RMSECV) and prediction (RMSEP), as control parameters. In fact, the methodology developed is proposed for direct relative density and refraction index in edible oils and their classification, requiring a few minutes per sample without any previous treatment. The third study aimed to evaluate the oxidative stability and quality of yogurt stored at 40C with light or dark using the combined parallel factor (PARAFAC) analysis and fluorescence spectroscopy. PARAFAC analysis of the fluorescence landscapes exhibited three fluorophores present in the yogurt, where, at least one of them was strongly related to the storage conditions. The fluorescence signal was resolved into excitation and emission profiles of the pure fluorescent compounds, which are suggested to be vitamin A, tryptophan and riboflavin. Regression model based on PARAFAC scores for riboflavin were built using the scores obtained in the first day as dependent variable and the scores obtained during the storage as independent variable. It was clear demonstrated that the slope of the analytical curve has become smaller throughout the experiment. Therefore, riboflavin level could be considered a good indicator for the yogurt stability. Thus, it is concluded that fluorescence spectroscopy in combination with chemometrics has a potential as a fast method for monitoring the oxidative stability and quality of yogurt

Este trabalho de pesquisa descreve três estudos de utilização de métodos quimiométricos para a classificação e caracterização de óleos comestíveis vegetais e seus parâmetros de qualidade através das técnicas de espectrometria de absorção molecular no infravermelho médio com transformada de Fourier e de espectrometria no infravermelho próximo, e o monitoramento da qualidade e estabilidade oxidativa do iogurte usando espectrometria de fluorescência molecular. O primeiro e segundo estudos visam à classificação e caracterização de parâmetros de qualidade de óleos comestíveis vegetais utilizando espectrometria no infravermelho médio com transformada de Fourier (FT-MIR) e no infravermelho próximo (NIR). O algoritmo de Kennard-Stone foi usado para a seleção do conjunto de validação após análise de componentes principais (PCA). A discriminação entre os óleos de canola, girassol, milho e soja foi investigada usando SVM-DA, SIMCA e PLS-DA. A predição dos parâmetros de qualidade, índice de refração e densidade relativa dos óleos, foi investigada usando os métodos de calibração multivariada dos mínimos quadrados parciais (PLS), iPLS e SVM para os dados de FT-MIR e NIR. Vários tipos de pré-processamentos, primeira derivada, correção do sinal multiplicativo (MSC), dados centrados na média, correção do sinal ortogonal (OSC) e variação normal padrão (SNV) foram utilizados, usando a raiz quadrada do erro médio quadrático de validação cruzada (RMSECV) e de predição (RMSEP) como parâmetros de avaliação. A metodologia desenvolvida para determinação de índice de refração e densidade relativa e classificação dos óleos vegetais é rápida e direta. O terceiro estudo visa à avaliação da estabilidade oxidativa e qualidade do iogurte armazenado a 4C submetido à luz direta e mantido no escuro, usando a análise dos fatores paralelos (PARAFAC) na luminescência exibida por três fluoróforos presentes no iogurte, onde pelo menos um deles está fortemente relacionado com as condições de armazenamento. O sinal fluorescente foi identificado pelo espectro de emissão e excitação das substâncias fluorescentes puras, que foram sugeridas serem vitamina A, triptofano e riboflavina. Modelos de regressão baseados nos escores do PARAFAC para a riboflavina foram desenvolvidos usando os escores obtidos no primeiro dia como variável dependente e os escores obtidos durante o armazenamento como variável independente. Foi visível o decaimento da curva analítica com o decurso do tempo da experimentação. Portanto, o teor de riboflavina pode ser considerado um bom indicador para a estabilidade do iogurte. Assim, é possível concluir que a espectroscopia de fluorescência combinada com métodos quimiométricos é um método rápido para monitorar a estabilidade oxidativa e a qualidade do iogurte / This research work describes three studies of chemometric methods employed for the classification and characterization of edible oils and its quality parameters through Fourier Transform mid infrared spectroscopy and near infrared spectroscopy, and for the monitoring the oxidative stability and quality of yogurt using fluorescence spectroscopy . The first and second studies aimed the classification and characterization of edible oil and its quality parameters using Fourier Transform mid infrared spectroscopy (FT-MIR) and near infrared spectroscopy (NIR) measurements, respectively. Kennard-Stone algorithm was used for selecting the training set, after a principal component analysis (PCA) was applied. The discrimination of canola oils from sunflower, corn and soybean was investigated using SVM-DA, SIMCA and PLS-DA. The quality parameters refraction index and relative density of edible oil was investigated using partial least squares (PLS), iPLS, LS- SVM multivariate calibration of FT-MIR and NIR data were evaluated. Several preprocessing alternatives (first derivative, multiplicative scatter correction, mean centering, orthogonal signal correction and standard normal variate) were investigated by using the root mean square error of validation cross validation (RMSECV) and prediction (RMSEP), as control parameters. In fact, the methodology developed is proposed for direct relative density and refraction index in edible oils and their classification, requiring a few minutes per sample without any previous treatment. The third study aimed to evaluate the oxidative stability and quality of yogurt stored at 40C with light or dark using the combined parallel factor (PARAFAC) analysis and fluorescence spectroscopy. PARAFAC analysis of the fluorescence landscapes exhibited three fluorophores present in the yogurt, where, at least one of them was strongly related to the storage conditions. The fluorescence signal was resolved into excitation and emission profiles of the pure fluorescent compounds, which are suggested to be vitamin A, tryptophan and riboflavin. Regression model based on PARAFAC scores for riboflavin were built using the scores obtained in the first day as dependent variable and the scores obtained during the storage as independent variable. It was clear demonstrated that the slope of the analytical curve has become smaller throughout the experiment. Therefore, riboflavin level could be considered a good indicator for the yogurt stability. Thus, it is concluded that fluorescence spectroscopy in combination with chemometrics has a potential as a fast method for monitoring the oxidative stability and quality of yogurt