Fermentation Of D-1,2,4-butanetriol By Recombinant Escherichia Coli And Its Extraction

D-1,2,4-butanetriol(BT)is an important precursor for the synthesis of energetic materials butanetriol trinitrate and several pharmaceutical ingredients.However,as a non-natural biological product,the synthesis processes of butanetriol mainly include chemical and biological methods.Although the former has a relatively large market share,chemical method has various drawbacks such as harsh reaction conditions,heavy environmental pollution,more by-products,and difficulties in subsequent separation and purification processes.Compared with biological methods,its technical prospects are obviously lagging.On the other hand,the current high-density fermentation of BT with microbial methods still has various shortcomings such as immature technology,substandard output,and high economic cost.Therefore,exploring the high-density fermentation and separation and purification technology of BT raw materials has important theoretical guiding significance and industrial application value to meet the needs of industrial production.This subject introduces the recombinant E.coli strain MJ138k-1A,realizes the microbial fermentation synthesis of BT products based on shake flask experiment and high-density fermentation process,explores the optimal process method for high BT production,and then uses thin film evaporation and molecular distillation separation technology.Explored a new method of direct concentration and purification of BT in fermentation broth.The research conclusions are as follows:1?The strain MJ138k-1A with the highest ability to synthesize BT was used as the fermentation strain,and the fermentation conditions were optimized in the shake flask experiment.Finally,it was determined that the fermentation temperature was33 ?,and 10 g/L CaCO3 was added to maintain the p H of the fermentation broth,and the fluid volume was 60 m L/250 m L,the medium was 1.5×LB,and the inoculation volume was 10% of the fluid volume.Under these conditions,the BT yield of recombinant strain MJ138k-1A after 72 hours of fermentation can reach 10.4 g/L,which is 25.3% higher than before optimization.2?To further increase the production of BT,the scale-up culture of E.coli was carried out in a 7 L fermentor based on the shake flask experiment to achieve highdensity fermentation of E.coli to produce BT.The effects of feeding strategy,dissolved oxygen concentration(DO)and inducer concentration(IPTG)on the yield of BT at the end of the fermentation were investigated;the results showed that when using dissolved oxygen feedback feeding,maintaining the DO at 15% and the inducer concentration at0.8 m M,the yield of BT can reach 20.5 g/L.3 ? There are few studies on the separation and purification of BT from fermentation broth.The paper tries to study the separation of BT by using a combination of carbon fiber column adsorption,thin film evaporation and molecular distillation;the optimal separation conditions are obtained by optimizing the adsorption rate,evaporation temperature and other parameters.The adsorption rate is 0.6 L/min,the film evaporation temperature is 95 ?,the scraper speed is 200 r/min,the molecular distillation temperature is 120 ?,and the scraper speed is 223 r/min.Finally,the combined detection method of high-performance liquid chromatography and nuclear magnetic resonance was used to determine the purity of BT which was greater than or equal to 98%.Through the above research,a complete technical route for high-density synthesis of BT using biological methods and subsequent separation and purification of target products has been initially explored,laying a theoretical foundation for the industrial production of bio-based BT.