Doctoral Degrees (Microbial, Biochemical and Food Biotechnology)
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Browsing Doctoral Degrees (Microbial, Biochemical and Food Biotechnology) by Advisor "De Wit, M."
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Item Open Access Fractionation and characerisation of a commercial yeast extract to facilitate acceleration of yogurt fermentation(University of the Free State, 2013-11) Smith, Esti-Andrine; Myburgh, J.; Osthoff, G.; De Wit, M.English: In order to study the decrease of yogurt fermentation time, the effects of a wide range of supplements on yogurt fermentation time were evaluated. YE was identified as the only supplement which showed potential. Unfortunately it resulted in a product with an unacceptable flavour. It was therefore important to identify and isolate the specific component responsible for the decrease in yogurt fermentation time. YE was fractionated with size exclusion chromatography and it was subsequently determined that the accelerating fraction had a low molecular weight. It was important to establish whether the accelerating component was of mineral, vitamin or amino acid origin. Three cocktails containing the most common minerals, vitamins and amino acids were prepared and their respective effects on yogurt fermentation were determined. Results indicated that when compared to the respective controls, no decrease in fermentation time was observed with the mineral and vitamin cocktails. A decrease in fermentation time was observed with the amino acid cocktail, indicating that the accelerating component present in YE was of amino acid origin. It was however not clear whether it was a single amino acid or a peptide. The accelerating fraction was further analysed by SDS-PAGE and due to no visible bands in the respective region (<1kDa), it could not be analysed with mass spectrometry. The fraction obtained directly after size exclusion chromatography was however analysed by using mass spectrometry in order to determine the total amino acid content of the accelerating fraction after which it was evident that the fraction containing the accelerant contained an abundance of peptides. The individual effects of the 17 identified amino acids were determined in respective yogurt fermentations. Results indicated that no single amino acid was responsible for the decrease in yogurt fermentation time. Although a combination of the 17 amino acids in one fermentation run resulted in a decrease in fermentation time in comparison to unsupplemented yogurt, the decrease was not as considerable as that of the accelerating fraction obtained after size exclusion chromatography. Due to the fact that it was not practical to evaluate the effect of all possible combinations of the 17 amino acids, the combinations that were evaluated were based on literature reports on stimulation of Streptococcus thermophilus growth. Focus was placed on the growth of Streptococcus thermophilus due to this organism being the growth limiting organism between the two starter organisms used for yogurt fermentation. None of the evaluated amino acid combinations decreased yogurt fermentation time, and it was therefore concluded that the accelerating component was a peptide and not a free amino acid. In order to establish the mechanism of acceleration of the isolated YE fraction, it was important to determine whether the addition of YE to milk increased the rate of starter bacteria growth or whether it completed the growth requirements of starter bacteria. The latter could either result in earlier initiation of lactic acid production or in increased levels of lactic acid production. Results indicated that YE had no effect on the total growth rate of the starter bacteria. However, when examining the effect on the respective bacteria individually, it became clear that YE increased Streptococcus thermophilus cell numbers in comparison to the unsupplemented control. Using resazurin, it seems YE increased the metabolic rate of the starter bacteria. Although supplementation of yogurt with YE did not influence combined starter bacteria growth, it influenced lactic acid production. The addition of YE to yogurt resulted in increased lactic acid levels as well as an increase in lactic acid production rate. Lactic acid was also initiated earlier in the fermentation process, resulting in higher lactic acid levels in comparison to unsupplemented yogurt. It was therefore evident from this study that YE did not provide nutrients that are not already present in milk, but rather provided nutrients in a readily available form at the beginning of the yogurt fermentation process resulting in the reduction of the lag phase of lactic acid production.