Despite the fact that the yeast Saccharomyces cerevisiae has frequently been the focus of genomic and proteomic research over the past 50 years and is the single organism to have been completely sequenced, there is still no method available for accurately determining the physiological state of a brewer’s yeast during the various steps of beer production. The vitality of yeast is currently evaluated using common analysis methods, such as yeast proliferation rate, fermentation rate, pH decline, etc., none of which provides insight into the operations inside the cell. Testing yeast performance through fermentation in the laboratory is possible, however, the results are first available three days later at the earliest, at which time the yeast is already in use. Therefore, subsequent corrections are impossible, even in the case of weakly fermenting yeast. Further disadvantages of the current analysis methods include their high cost and the need for trained staff to operate the required equipment and carry out any additional work. With the help of uncomplicated methods for measuring enzyme kinetics, we attempted to gain insight into the inner workings of the cell. The primary focus of this analysis was the enzyme activity of maltase (MAL), pyruvate dehydrogenase (PDH), pyruvate decarboxylase (PDC) and alcohol dehydrogenase (ADH), all of which have a key function in yeast metabolism. Due to the fact that past studies of these enzymes were conducted with laboratory strains, our first task was to gather data on yeast enzyme activity during all stages of beer production. For this purpose, a number of laboratory-scale fermentations were performed at the institute and the enzyme activity was measured and further supplemented with analysis results from a brewery. With the help of a formula for calculating fermentation performance, it was possible to compare the various fermentations conducted as part of this research. The activity of maltase is significant for the utilization of maltose, the primary sugar present in wort. Maltase was evaluated according to the following criteria: the level of initial activity, the effects of enzyme repression, analysis of the enzyme maxima as well as relationship between maltase activity and fermentation performance. The initial maltase activity can be used to predict the nature of the subsequent fermentation. Higher levels generally lead to a more rapid fermentation during which an increase in fermentation performance is observed, given that other factors, such as the composition of the wort or the absence of aeration, do not strongly impact the growth of the yeast. During propagation, higher activity results in an increase in cell concentration. If the maltase activity is too high, the yeast exhibit a tendency to start fermenting at the beginning of yeast multiplication. Through continuous yeast multiplication, we were able to show that the existence of lower ADH-activity, regardless of the level of the maltase activity, lead to poor fermentation performance. The performance of pitching yeast containing abundant enzymes is also subject to the technological conditions present during fermentation, which can also have an impact on enzyme activity. Therefore, maltase can be evaluated by measuring the following: the duration of the repression phase, the decrease in activity during the repression phase, the time and level of the first maximum after repression, as well as the overall level of enzyme activity. More rapid fermentations showed a distinctly shorter repression phase. During propagation, the duration of the lag-phase was dependant on the duration of repression and the reduction in activity. Better compensation for this reduction in enzyme activity and correspondingly a stronger induction of maltase after repression was observed when the initial enzyme activity was high. Furthermore, an early first maximum after repression and the level of this maximum are important factors in determining the rate of fermentation and the degree of yeast proliferation, respectively. During fermentation in cylindroconical tanks, it was observed that maltase activity exceeding the initial activity was indicative of an improvement in the physiological state of the yeast. In contrast, maltase activity during propagation always remained below the initial activity. This confirms the fact that under aerobic conditions, maltose is utilized to a lesser degree for energy yield. Basic correlations were evident between the level of enzyme activity and the estimated fermentation performance during propagation and fermentation. In order to find out whether pyruvate, the end-product of glycolysis, is used as effectively under these circumstances as it is during fermentation and propagation, the enzymes of pyruvate metabolism (PDH, PDC and ADH) were examined. Originally, it was assumed that PDH activity is highest under aerobic conditions. However, this is not the case, because the low pyruvate concentrations enable the PDH, in spite of its low km-value, to also catabolise the pyruvate in conjunction with PDC. This, together with the high PDC activity, has a positive effect on yeast cell count. In contrast, no increase in PDH activity was detected during propagation. In this manner, PDH can be used as an indicator for the existence of anaerobic conditions. Also, through examination of ADH activity, it is possible to determine, whether the acetaldehyde formed by PDC has been sent through the PDH bypass to acetyl-CoA or whether it has served as a hydrogen acceptor in the last step of alcoholic fermentation. The presence of high ADH activity is indicative of an overloaded PDH bypass, which results in an increase in alcoholic fermentation and suppressed yeast multiplication. This was evident through an increase in fermentation performance. Conversely, when low ADH activity was coupled with high PDC activity, a stronger formation of acetyl-CoA for lipid synthesis took place, as was the case during propagation. During yeast propagation a contraction of the lag phase was observed with yeast which exhibited high PDH, PDC and ADH activity. This may have enabled the yeast to adapt more readily to the Crabtree environment. These results show that measuring the enzyme activity of key yeast enzymes provides important clues about intracellular processes. Further research could be conducted on the enzyme activity involved in the PDH bypass (acetaldehyde dehydrogenase; acetyl-CoA synthetase), to clarify the interactions of the processes responsible for pyruvate metabolism during fermentation and propagation. Furthermore, information from the fields of metabolomics, proteomics, transcriptomics and genomics could contribute to a better understanding of cell metabolism, all of which combine to form “metabolic engineering”. Were the relationships among these various fields of study well-known, the physiological state of yeast could be effectively evaluated on the basis of data from one area alone. This would significantly alter the current methods of yeast management practiced within the brewing industry and would bring with it both technological and economic advantages.
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