Fed-batch Strategy Optimization And Metabolic Flux Analysis Of Thuringiensin

Thuringiensin is a kind of nucleotide derivative antibiotic and has a broad spectrum against agricultural pests, such as Coleptera, Diptera, Hymenoptera, Isoptera, Lepidoptera, Orthoptera, Hemiptera, aphid, nematode and mite.An improved pH-control fed-batch strategy for Bacillus thuringiensis subsp. darmstadiensis 032 producing thuringiensin was developed based on the analysis of the batch culture, constant rate fed-batch cultures and the original pH-control fed-batch. Having considered the pH variation and the glucose consumption status, the pH was adjusted from 6.5 to 7.0 by adding base in the late cultivation period of batch culture, and then the pH was kept at 7.0 by glucose feeding. The feeding was terminated when the pH couldn't be controlled by glucose feeding anymore. The proposed fed-batch strategy effectively avoided underfeeding or overfeeding, and it increased the thuringiensin yield and Y_P/X by 89.51% and 103.2% compared to that of the batch culture, respectively.The metabolic network of Bacillus thuringiensis subsp. darmstadiensis 032 producing thuringiensin was developed. The metabolic fluxes during the culture and the culture with tri-sodium citrate were analyzed by professional metabolic flux analysis software - FluxAnalyzer under Matlab environment. The result showed that the glucose-6-phosphate node and the acetate coenzyme A node were flexibility, and the a-ketoglutarate node was semi-flexibility. Based on the analysis and the former pH-control fed-batch strategy, 2 g/L tri-sodium citrate was added to the culture medium to adjust the metabolic fluxes distribution, and the final thuringiensin yield was improved by 154.2%.The thuringiensin degradation processes in aqueous under different conditions, where pH ranges from 5.0 to 9.0 and temperature ranges from 10℃ to 40℃, were systematically investigated. The half-life of thuringiensin was calculated by an exponential decay model, and the half-life was ranged from 2.72 to 16.19 d in the conditions we investigated. This indicated that thuringiensin was an easily degradable antibiotic. In this study, an artificial neural network (ANN) was employed to further describe the degradation processes. The radius basis neural network (RBF) model, which the accuracy was 0.1 and the SPREAD value was five, could well simulate and predict the degradation processes. The mean...