Fermentation Optimization,Purification And Enzymatic Characteristics Of Fucoidanase From Marine Penicillium Expansum

Pectinase refered to the kind of enzyme that could decomposite pectin,including PG,PE,PL,etc.Pectinase was widely used in juice clarification,wood preservation,hemp degumming,cotton fabric scouring,natural drug active ingredient extraction,feed processing,tea fermentation,environmental protection and other fields.In this paper,we studied the Marine Penicillium expansum from the laboratory preservation,and its producing pectinase was studied.Solid fermentation conditions of pectinase from marine Penicillium expansum were optimized by single factor and orthogonal experiments to improve the production of pectinase.The optimal condition of PG is composed of bran 7 g,jelly powders 0.42 g,ammonium chloride 0.28 g and artificial seawater 8.4 m L,and was cultured at 28 ? for 72 h with 3 mL?107/mL?inoculation in each 250 mL triangle flask.The pectinase activity could reach 6643.49 U/g,which was 4.36 times more than that at the initial conditions with 1239.80 U/g.The optimal condition of PE was composed of bran 7 g,starch and jelly powders 0.14 g,ammonium chloride 0.28 g and artificial seawater 10.5 mL,and was cultured at 28 ? for 72 h with 3mL?107/mL?inoculation in each 250 mL triangle flask.The PE activity could reach178.25 U/g,which was 4.44 times more than that at the initial conditions with 40.12U/g.The optimal condition of PL was composed of bran 7 g,jelly powders 0.42 g,urea 0.28 g and artificial seawater 10.5 m L,and was cultured at 32 ? for 72 h with3 mL?107/mL?inoculation in each 250 mL triangle flask.The PE activity could reach5.12 U/g,which was 5.95 times more than that at the initial conditions with 0.86 U/g.And enzyme process of PG was studied by using the optimal fermentation conditions on the yield of PG enzyme process was studied,the results showed that PG was synchronous synthetic enzyme.According to the obtained optimal solid-state fermentation conditions of PG to cultivation of Marine extension mold,extract enzyme with buffer,then centrifugat at10000 r/min,separate and purificate PG by ultrafiltration,DEAE Sephadex A-50 ion exchange chromatography,Sephadex G-100 gel filtration chromatography,final purification ratio was 24.13,the recovery rate was 36.32%,the SDS-PAGEelectrophoresis was shown as a single strip,and the molecular of PG was about 63.96 kDa.Results of PG enzymology properties revealed that the optimum reaction temperature was 50?,and the optimum pH was 5.4.Pectinase was stable in the range of pH4.6-6.2.Activation on pectinase activity was that Mg²⁺>Ca²⁺>Na⁺>Sr²⁺> Co²⁺>K⁺,Mg²⁺,Ca²⁺ had obvious activation on pectinase activity,and Cu²⁺ had obvious inhibition.Mn²⁺ almost did no effect on the activity of PG.Vmax and Km for jelly powder were 393.56 ?g/?min?m L?and 3.34 mg/mL respectively.