Study On Producing L-Ornithine By Arginase In Microorganism

L-ornithine, which exists in many kinds of tissue and cell, plays important roles in organism metabolism. It is intermediate product of urea producing and precursors of many amino acids such as arginine and citrulline. L-ornithine takes part in the urea cycle and detoxifies the high concentration of ammonia in the body. It is mainly used in preparing the compound amino acid infusion which shows effects on liver protecting and anti-fatigue. At present, there are four methods to prepare L-ornithine: extraction, chemical, fermentation and enzymatic method. Enzymatic method is preparing ornithine by arginase which exists in animals, plants and microorganisms widely. This thesis researched the entire process for ornithine by enzymatic conversion. The process includes strains screening, fermentation, conversion and separation. We try to find a effective method for industrial production.By preliminary screening, mutagenizing and secondary screening, a strain with high activity of arginase named XT-025 was selected. The strain was identified as Bacillus thuringiensis according to its morphology, physiological characteristics and 16S ribosomal RNA gene sequence analysis.The fermentation medium included 2% glucose, 1.2% fish peptone, 0.1% inorganic salt. XT-025 was cultured at 35℃, with 180 r/min rotation speed, for 24 h. Then the cells were used to convert arginine into ornithine in 0.3 mol/l carbonate buffer (V(Na₂CO₃):V(NaHCO₃)=8:2) with 5% arginine and 0.1 mg/ml manganese ion at 30℃for 24 h. The final ornithine yield was 43.57 g/l and the molar conversion rate of arginine reached 90.13%.732 strongly acidic cation exchange resin was used to separate ornithine and arginine and pH value of sample solution was 7. The ornithine and arginine were eluted by different concentration of ammonia. The elution rate of ornithine was 98.71%. Finally, the purity of ornithine was 96.17% and the yield of ornithine in whole separation process reached 81.49%.