Fermentation Process Of Laeease Production From White Rot Fungi And Its Preliminary Appiication In Flour Products

The laccase (Laccasc, EC1.10.3.2.), which is one of multicopper polyphenol oxidase, was first discovered from the sap of the Japanese lacquer and then found that it is widely exist in plants, insects and higher fungi. Laccase can catalyze the oxidation of phenolic compounds, remove electrons or protons on hydroxyl radical formation, form free radicals, and make the degradation of phenols and lignin compounds as well as the molecular oxygen reduceing to water at the same time. It causes people growing interest and become a current research focus due to its special catalytic performance and broader substrates, appearing broad application in the field of biological, chemical and environmental.The most prominent feature of laccase are the degradation of the lignin in the pulp, bleaching wastewater and removal of toxic phenols in wastewater, and have a very large application prospects in the paper industry. At present, the domestic and foreign research on laccase has two directions, one is the optimization of technology and applications of laccase, the other is to find a suitable carrier for heterologous expression of laccase gene. But the high expression of white rot fungi in the active host was so difficult, because it is still in the stage of exploration, that on laccase focused screening strains, optimization of culture conditions, enzyme in bacteria immobilization and the pollutant treatment.The synthesis levels of fungi laccase is closely related to the ability of producing enzyme of the strains, and is affected by multiple factors of environmental conditions, such as the carbon source, nitrogen source and inducing agent, especially the inducing agent is added or not, adding amount and adding time, which all have a very significant effect on the activity of laccase.The present experiment studied the influence of various factors on the synthetic laccase ability in the liquid culture conditions, including inoculation volume, temperature, time, speed and pH value of medium, as well as the carbon source, a nitrogen source, ratio of carbon to nitrogen and inducing factors. Results show that, when the carbon source is glucose, yeast extract as nitrogen source, C/N ratio was15:1, the P. conchatus synthesis of laccase activity reached8795U·L-1; while adding1.5%wheat bran and1mmol/L Cu2+to induce laccase synthesis, in the initial, the laccase activity reached51004U·L-1; and while adding3mmol/L Cu2+after4days in the fermentation and other conditions remaining unchanged, enzyme activity reached101968U·L-1, which was nearly two times than the initial conditions of medium.To reference the process conditions of laccase production in shake flask and to enlarge the fermentor to5L, fermentation of white rot fungi attempts to expand production initially.From the flask to the fermentor, shear force and dissolved oxygen have undergone great changes, while the secretion activity of laccase was lower than shake flasks. Preliminary processing of fermentation products, the enzymatic properties of laccase showed that the optimum reaction temperature was45℃, at4℃preservation, presenting the best thermal stability, and relative activity reached96.4%after20days; the optimum reaction power of hydrogen adapted to pH7, at4℃, presenting the best stability of laccase when the pH value was8in surrounding, and the relative activity reached90.5%after20days; when the Mg2+, Mn2+, Zn2+and Cu2+at a concentration of1mmol/L, the white rot fungus P. conchatus presented a weak role in activation, however, the K+, Ca2+, Ba2+,Al3+, Fe3+, Mg2+and Fe2+exhibited obvious inhibition, especially in the concentration of5mmol/L, the inhibitory effect was more obvious.Common effect of both in the oxidation of laccase and laccase and ferulic acid in flour to form gel, laccase can be very good to improve the quality of flour products. In the production process of steamed bread,steamed bread of laccase can improve the good external and quality, make it more white, the structure is more uniform and fine, larger size, elastic is not easy to collapse; in the preservation process of steamed bread, laccase can reduce the volatilization rate and volatile content of moisture due to forming gels, and better maintain moisture in steamed bread; adding laccase in the production process of wet flour, can make bacteriostatic action better.