Breeding Of Producing L-lactic Acid Bacteria And Research Of Fermentation Technology

L-lactic acid has anti-fatigue,repair,antibacterial,antioxidant effect and so on.Theproduct extends to a plurality of fields,such as medicine,food,chemical,environmentalprotection products and so on.In this paper, Lactobacillus rhamnosus of keeping in mylaboratory was mutated to obtain high production L-lactic acid strain. The fermentationconditions and medium were optimized, the aim was to improve the capacity of thisstrain to produce L-lactic acid. The specific research contents were as follows:1.Optimized the culture medium and cultivating condition of original strain. Seedliquid culture time using the logarithmic growth phase: seed age of Lactobacillusrhamnosus strain was10h. By one-factor-at-one-time experiment, results showed thatthe optimum fermentation conditions of initial pH, temperature,fermentation period,inoculation and liquid volume to produce L-lactic acid were6.6,39℃,82h,10%and50mL respectively.Based on the optimization culture conditions, medium compositionwas selected by single factor experiment, steepest ascent test methods and responsesurface optimization.The optimized medium components were: glucose132.27g/L,diammonium hydrogen citrate2.955g/L, sodium acetate7.43g/L, peptone0.625g/L,yeast extract FM8188.75g/L. Verification tests on the optimized formulation, L-lacticacid production reached56.7g/L, consistent with the model prediction.2.The original strain Lactobacillus rhamnosus D-10was mutated by diethylsulphate (DES). The best mutated time was35minutes and mutation strain D1wasobtained. After that,the strain DES was mutated by ultraviolet radiation (UV). The bestmutated time was40s and mutation strain UV7was obtained.Then the strain UV7wasmutated by compound mutagenesis (LiCl) and mutation strain L1was obtained. TheL-lactic acid production of mutation strain L1was69.26g/L, strain D-10was53.96g/L,which was enchaced28.35%compared with the strain D-10. By genetic stability tested,strian L1showed a good stability.While the Lactobacillus rhamnosus strain wasmutated,screening experiment of mutate strains resistent to glucose and lactic acid weredoing at the same time. The result showed that the mutants resistant to glucoseconcentration was from the original250g/L to300g/L, high calcium lactateconcentration was from the original120g/L to150g/L.3.Optimized the culture medium and cultivating condition of mutant strain L1.Seed liquid culture time using the logarithmic growth phase: seed age of Lactobacillusrhamnosus strain L1was9h. By one-factor-at-one-time experiment, results showed thatthe optimum fermentation conditions of initial pH, temperature,fermentation period,inoculation and liquid volume to produce L-lactic acid were6.3,39℃,78h,8%and50mL respectively.Based on the optimization culture conditions, medium compositionwas selected by single factor experiment, steepest ascent test methods and responsesurface optimization.The optimized medium components were:glucose90g/L,diammonium hydrogen citrate4.775g/L, sodium acetate3.08g/L, peptone0.827g/L,yeast extract FM8189g/L. Under the identified optimal conditions, the maximalL-lactic acid production of L1was76.56g/L, which was35.02%higher than that ofstrain D-10(56.7g/L).