Research On The Immobilization Of Xylanase By Coconut Fiber And Its Enzymatic Hydrolysis Kinetics

In this paper,coconut fiber was studied as raw material,after acid-base treatment and oxidation of sodium periodate,the covalent fiber-bound xylanase was prepared by covalent bonding method,and the xylan in coconut shell powder was directly hydrolyzed to study the effect of immobilized xylanase catalyzed hydrolyzed wood Glycan reaction mechanism and kinetic analysis.Using dilute hydrochloric acid hydrolysis method to remove the hemicellulose on the surface of the coconut fiber,the optimal combination parameters are:dilute hydrochloric acid concentration was 1.7mol/L,extraction time was 2h,extraction temperature was 104 ?.Under these conditions,The best removal rate was 18.30%,the standard recovery of xylose was 88.5%?97.375%,and the relative standard deviation of repeatability test was ± 1.09%.The method was simple,convenient,reproducible and quick,the relative error was small.Hemicellulose can be extracted from coconut fiber using dilute sodium hydroxide solution,and its structure is analyzed and characterized.The results showed that the optimal combination parameters were as follows:the concentration of sodium hydroxide was 5%,the extraction time was 4h,the extraction temperature was 65 ?,the yield of hemicellulose on the surface of coconut fiber was the highest(14.33%).The resulting sample was analyzed by infrared spectroscopy and confirmed as coconut hemicellulose.The number average molecular weight was 49049 and the weight average molecular weight was 368,354 by GPC gel permeation chromatography.The coconut hemicellulose hydrolyzed with dilute sulfuric acid and then analyzed by high performance liquid chromatography,the retention time consistent with the standard xylose retention time.After the acid hydrolysis of the washed coconut fiber,the aldehyde-based coconut fiber can be prepared by the oxidation of sodium periodate solution.The optimum combination parameters were as follows:the concentration of sodium periodate solution was 0.7mol/L,the reaction time was 3.5h,the reaction temperature was 45 ?,periodic acid solution pH was 2.3,under the conditions of oxidation,coconut fiber surface aldehyde concentration of 2.74mmol/g.The prepared aldehyde-based coconut fiber can be covalently bound to xylanase through Schiff reaction to prepare coconut fiber-immobilized xylanase.The results showed that the optimal combination parameters were as follows:the concentration of xylanase was 200U/mL,the immobilization time was 24h,the immobilization temperature was 39 ? and the pH of the buffer was 4.2.Under this process condition,The activity of the coconut fiber-immobilized xylanase was 18.6 U.Compared with the free enzyme,after immobilization,the optimum reaction temperature of the enzyme increased by 5 ? and the optimal pH was 0.6 units to the acidic direction,the thermal stability and acid and alkali resistance were significantly improved,and after repeated use for 7 times,it still has about 80%residual enzyme activity,with good reusability.Xylanase is catalyzed by immobilized xylanase using coconut fiber.The results showed that the optimal combination parameters were as follows:the hydrolysis time was 60min,the amount of imobilized enzyme was 0.1400g,the substrate concentration was 40mg/mL,the hydrolysis temperature was 55 ?,the pH value of the buffer was 4.5.Under this condition,Liquid xylan conversion rate(DE)reached 27.24%.The results of kinetic analysis showed that the concentration of xylan solution and the reaction speed of the immobilized enzyme catalyzed hydrolysis satisfy the "hyperbola"characteristic of the Mie equation,which is consistent with the first order reaction law.Using the formula deduction,the Michaelis constant Km is 14.06 mg/mL,Vm ?0.1465mg/(mL.min),find the Mie equation is V = 0.1465S/S+14.06.