The Research On Laccase Of Myrothecium Melanosporum

In this dissertation, systematic researchs were focused on the laccase by Myrothecium melanosporum.Two different termination methods of Myrothecium melanosporum laccase determination were compared in this paper. The results revealed that it was feasible to terminate the enzyme reaction by heat. Compared heating with freezing, the former was more reliable, repeatable, simple and convenient.The optimum culture medium and conditions in shaking flasks for laccase production by Myrothecium melanosporum were studied. To measure Myrothecium melanosporum laccase cativity in different culture medium and conditions through orthogonal experiment, the optimum conditions were:initial pH of medium is 5.5, 100mL medium in 250 ml-flask, inoculation 7.5%,mycelium age 9×24hr, nature light, 28℃. Myrothecium melanosporum laccase activity of submerged fermentation liguid improved remarkably in the optimum culture and conditions, with 2.49 times as non-optimized culture conditions. The optimum culture components were:Glucose 20g/L, cotton 0.1%, ammonium tartrate0.2%, peptone0.5%. After culture medium optimization, Myrothecium melanosporum laccase activity of submerged fermentation liquid is 3.58 times as basic liquid medium,1.07 times as optimum conditions.The biosynthesis of Myrothecium melanosporum laccase was affected in various extents by four metallic ions of Cu2+, Zn2+, Mn2+, Fe2+. Four metallic ions reacted on low concentration. The effect of Cu2+ was the most in four, then Mn2+. The effect of Zn2+ and Fe2+ were less. Above some concentration, the effects of four ions were changed into inhibitory action.Effect of four different metallic ions and inducers was detected in vegetation process of Myrothecium melanosporum.The induction of four inducers:vanillin, guaiacol, ferulic acid and O-tolidine were studied. The results indicated that, four inducer have some effect, but not obvious.From the PAGE zymogram, it was inferred that Myrothecium melanosporum laccase is a single enzyme and there was no other isoenzyme expressed in the process of liguid fermentation.