Study On Enzymatic Preparation Of Nanocellulose And Optimization Of Enzymatic Sysem

With the rapid development of economy and depletion of non-renewable resource,the problems of resources and environment have become increasingly prominent.The search for green and renewable resources is even more urgent.As one of the most abundant renewable resources on the planet,cellulose has received increasing attention.The research and application of cellulosic materials become more in-depth and extensive.Nanocellulose,because of its strong physical and chemical properties,was used for producing various nanomaterials as additives to improve the properties of composite materials.At present,the common methods for preparing nanocellulose includes physical and chemical methods,but their disadvantages such as high energy consumption,high requirements for equipments,difficult in treatment of strong acidic waste water,limited industrial application of the two methods.Enzymatic preparation of nanocellulose has been considered as one of the most potential methods because of their some advantages such as mild preparation conditions and no pollution.However,up to now,the efficiency of enzymatic preparation process is poor because of lack of effective enzymatic system for preparing nanocellulose.Now,enzymatic treatment is often used as a pretreatment method of raw materials for decreasing consumption of energy and chemicals in some studies.In this thesis,the feasibility of enzymatic preparation of nanocellulose was studied by using different cellulase and pine chemical pulp as a substrate,and the enzymatic system used for preparing nanocellulose was preliminarily optimized by adding different components of cellulase and hemicellulase.The results were as follows:During the process of enzymatic preparation of nanocellulose,the ultrasonictreatment step was necessary for effective separating cellulosic nano-particles and increasing efficiency of enzymatic process.The ultrasonic treatment after enzymatictreatment was better than that before enzymatic hydrolysis for preparation ofnanocellulose.On another hand,a suitable strength of ultrasonic treatment was alsonecessary and the particles are more likely to aggregation if the ultrasonic time too long.Pine chemical pulp was hydrolyzed by nine cellulases for preparing nanocellulose and their effect on enzymatic preparation were evaluated respectively.It was found that there were different effect on enzymatic preparation for the enzyme with different enzyme components,in which,commercial enzyme Fiber care D,the cellulase from Penicillium oxalicum I1-13 and the cellulase from Trichoderma reesei SCB18 were better than other enzyme for enzymatic preparation of cellulosic nano-particles.Overall,based on the enzymatic process efficiency,particle sizes,etc.the cellulase from Trichoderma reesei SCB18 was more suitable for being used to prepare nanocellulose when pine chemical pulp as substrate.The enzyme system of cellulase for preparing nanocellulose was optimized by adding extra different enzyme components to the crude cellulase solution from Trichoderma reesei SCB18.Firstly,four enzyme components including ?-glucosidase(BG),endo-glucanase(EG),cellobiohydrolase ?(CBH ?)respective from Penicillium oxalicum and Trichoderma reesei were added respectively,and found that the effect of addition of BG and EG were better than that of addition of CBH ?.The addition of BG significantly reduced the particle size and increased the yield of nanoparticles,which maybe because the increase in enzyme activity of BG decreased the inhibition of hydrolysis products for enzymatic hydrolysis,thus therefore promoted the enzymatic degradation of cellulose.The effect of the addition of EG on the particle size reduction was not obvious,though the yield of nanoparticles increased,should due to the rapid hydrolysis of cellulose in amorphous region under a higher EG dosage.Four hemicellulase components,xylanase(Xyn),?-xylosidase(Xyl),acetylxylan esterase(Axe),mannanase(Man),were also respectively added to the crude cellulase from Trichoderma reesei SCB18 for evaluating their effect on enzymatic preparation of nanocellulose in the study.It was found that the four hemicellulase components had little effect on particle size.The addition of Xyn,Xyl and Axe increased the yields of nano-particles,in which,the Xyn was the most effective.It should because the degradation of more hemicelluloses promoted the degradation of cellulose on a certain extent.Compared to other three hemicellulase components,the addition of mannanase(Man)resulted in a higher glucose consistency in hydrolysis liquor after enzymatic hydrolysis,should due to the high content of glucan mannan in pine chemical pulp.The effect of addition of two kinds of accessory proteins,lytic pohysaccharide monooxygenases and swollenin,were also compared.It was found that addition of the two proteins obviously reduced particle size,but didn't significantly affect the yield of nanocellulose.