Optimization Of Gnicotinic Acid Fermentation Conditions And Strain Identification

Niacin, also known as nicotinic acid, vitamin B₃, pyridin-3-carboxylic acid, is essential for peopleto maintain the physiological functions. At present, niacin is mainly used for feed additives, pharmaceutical,food industry, chemical industry.The main methods used in niacin synthesis include chemical synthesis and biological synthesis.Chemical synthesis method is the principal means on the market currently, but it’s high cost of production,strict to the equipment, and its process of purified Recycling is complex, moreover it will produce seriouspollution to the environment. As biosynthesis can overcome the unfavorable factors which made by thechemical synthesis, it has become more promising. Biosynthesis of niacin use nitrilase which yielded byorganism as catalyst, catalyticing the substrate nicotinic acid3-cyanopyridine to niacin. In this study, weresearch the biosynthesis of niacin which subject of production nitrilase strains produce niacin do theresearch, as follows:1. Screening for a strain which yield nitrilase. Soil or water samples with many nitrile compoundswere coated on the medium which made3-cyanopyridine as the sole nitrogen source.3days later,72strains which can grow on the filter medium were obtained. The concentration of the production of thesestrains was determined by micro-Berthelot method （phenol-hypochlorite method）, and strain T-1withhighest activity was rescreenied finally.2. Identification of the strains. T-1was analysised by colony morphology, microscopicobservation, physiological and biochemical reaction. Though comparing T-1with standard strain with《Bergey bacterial identification manual》, T-1was identificated as Klebsiella pneumoniae subsp finally.3. Optimization of fermentation conditions on the T-1. First. the compositions and conditions ofthe fermentation medium were optimized by single factor test, determining the optimum value of variousfactors: glucose1.2%, yeast extract0.6%, K₂HPO₄0.05%, KH₂PO₄0.05%, MgSO₄·7H₂O0.02%,caprolactam0.2%, nicotinonitrile0.2%, optimum fermentation temperature28℃, pH7.0, shaker speed200rpm. Furthermore, response surface regression analysis and orthogonal experiment were used, and theresult as follow: glucose1.0%, yeast extract0.77%, K₂HPO₄0.05%, KH₂PO₄0.05%, MgSO₄·7H₂O 0.02%,0.2%caprolactam,0.11%nicotinonitrile, optimum fermentation temperature28℃, pH7.0, shakerspeed200rpm. And the acticity of nitrilase was20.82U/mL after optimized, increasing about113.5%.