Study On Culture Conditions Of Nitrile Hydratase And Acrylic Amide Crystal Method Of Preparation

Acrylamide is a very important chemical product in fine chemical field. It can be used to compose polyacrylamide. With the increasing demand of polyacrylamide and its monomer acrylamide at home and abroad, the study on the producing methods of acrylamide is becoming popular. There are many advantages in the application of immobilization technology for continuous production of acrylamide by microbial methods, such as simplifying process, longer use of immobilized cells, low cost, easy separation and recovery of catalyzer, etc.In my study, a strain with high nitrile hydratase activity was isolated and screened, which produced acrylamide from activated sludge of wastewater containing nitrile. Then the strain was acclimated and the affecting factors of nitrile hydratase activity were investigated.In the study of immobilization, nitrile hydratase producing strain strain was immobilized and continuous production of was conducted in self-made enzyme column reactor, aqueous solution of acrylamide produced from continuous flow reactor was used as material, the production and refine methods of acrylamide crystal were studies, and acrylamide crystal were analyzed by infrared spectra and Nuclear magnetic resonance. The mainly results were as followed:The nitrile hydratase producing strain U2is isolated and screened from activated sludge of wastewater with10-5dilution factor and nitrile concentration of2g/L on medium. Nitrile hydratase activity of strainU2is486.4U and It is renamed as BJHM-1. Acrylonitrile was used for the gradiently acclimation of strain. When the acrylonitrile mass concentration of culture medium reaches to6g/L, the nitrile hydratase activity is the highest, which improved to758.6U. In the study of affecting factors of nitrile hydratase activity, PH value and temperature on strains BJHM-1were studied, results showed that strain BJHM-1had the optimal enzymatic activity at culture temperature of30℃and PH7.5, whose enzyme activity was530.3U.1.0g/L nitrile hydratase producing strain was added to chloride agent for immobilization. The concentration of acrylamide solution get from immobilized continuous catalysis reactor was7.5xlO-5g/L. The concentration of the water solution ratio is closer to100:1, the temperature was control at65℃, the crystallization temperature was0℃-5℃, join the seed to induced crystallization happen. Nuclear magnetic resonance (NMR) C spectrum demonstrated that,-CH2in128nm mark peak,-CH sign appeared in131nm peak,-CO in170nm mark peak and no other carbon impurity appear. According to the infrared spectra, in3370,3185cm-1of the double peak for N-H characteristic peak, In2818cm-1place for the peak of a double C-H telescopic vibration characteristic peak1676cm-1place of strong peak for C=O asymmetric telescopic vibrating peaks.1448cm-1place for the peak of a double C-H bending characteristic peak, which can determine that the substance we obtained from microbial immobilization reactor is acrylamide.