Fermentation Preparation Of Nisin By Lactic Acid Bacteria 6032 In Media With Enzymatic Hydrolysis Of Potato Juice

Nisin is a kind of antibacterial peptides,which is produced by Lactococcus lactis or Streptococcus lactis.It can inhibit growth and reproduction of most gram-positive bacteria,especially for the inhibitory effect of staphylococcus aureus,hemolytic streptococcus.It was safe,non-toxic,effective antibacterial actions to the human body.In recent years,it is expected to be as a substitute for antibiotics in the treatment of diseases in the pharmaceutical industry.Potato starch is abundant biomass resources,a variety of products can be obtained by potato starch biological refining.Based on the potato as raw material,Nis in was synthesized by Lactic Acid Bacteria 6032.This work not only extended the potato industry chain,but also increased the kinds of biological refining products.The main research contents and results of this paper were as follows:(1)The process of potato processing browning inhibitors was studied,the best inhibitor collaboration was determined.The scope of different browning inhibitors were determined through the single factor experiment.On the basis of the experiments result,various affecting factors were optimized by orthogonal experiment.Browning inhibitors composition were as follows: 0.40% of citric acid,0.15% of L-cysteine,D-0.30% ascorbic acid and 0.80% sodium chloride.Under these conditions,browning degree was 0.221.(2)The various factors hydrolysis of potato juice which were studied b y single factor experiment.Effect of these factors were optimized by orthogonal experiments.This experimental result showed that ?-amylase was 0.90%,p H6.00,temperature 60.00?,time 2.00 h.Under the optimum conditions,the enzymolysis of potato starch DE value was 14.661.At the same time,the concentration of CaCl2 had influence on stability of ?-amylase which was studied,when the concentration of CaCl2 was 0.15%,the ?-amylase have better stability compared to other levels.(3)Original potato starch,hydrolyzed potato starch by ?-amylase and hydrolyzed residue partic les of potato starch by glucoamylase which were analyzed by X-ray diffraction patterns and SEM diagram.And the experimental result shows that particle had been destroyed in the process of digestion of starch crystal shape.It was not only the zymohydrolys is on the surface of the starch granules,but also internal to form hole,which showed starch degradation had been to form small molecules at the same time.(4)Potatoes can be used as a raw material for production of Nis in fermentation.The experiment results showed that the titer unit of Nis in was higher by using enzymatic hydrolysis of potato juice as fermentation medium than enzymatic hydrolys is of potato starch.So enzymatic hydrolys is potato juice was be used to production of Nisin as fermentation medium.The appropriate nutrients were added into hydrolyzed potato juice,the concentration of various nutrients were determined by single factor experiment.On the bas is of the experiment,according to the Box-Behnken group and design princ iple,the various nutrients were optimized by using response surface analys is.The results showed that the optima nutrients composition were as follows : ammonium sulfate 10.40 g/L,dipotassium hydrogen phosphate 5.00 g/L,corn steep liquor 10.70%.Under these conditions,the titer unit of Nisin was 4421.00 IU/m L,OD600 was 0.6007.Initial p H of fermentation had influenced for the titer unit of Nsin,the result showed that the titer unit of Nis in raised to 4635.00 IU/m L on the time interval for 6.00 h by regulating p H of fermenting broth to initial p H.(5)Potato starch can be applied to production of Nisin by simultaneous saccharification and fermentation(SSF).The various factors to influence fermentation was determined through experiment.On base of this,the fermentation conditions were optimized by Box-Behnken group and design princ iple of Design-Expert 8.0.The experiment results showed that the fermentation conditions were as follows,medium initial p H4.70,fermentation temperature 32.00?,inoculation amount 8.20%,the titer unit of Nisin was 4093.00 IU/mL,OD600 was 0.5699,in which the experimental values of the titer of Nisin were closed to predicted value.These results showded that the optimal conditions were feas ible and reasonable.(6)Molecular biology identification of 16 S r DNA was employed to fermentation strains.Phylogenetic treesic tree was constructed based on 16 S rDNA gene sequences,The results showed that the fermentation strains was located in the lineage of Enterococcus and had 99% sequence similarity to the type strain of faecium.