Study On The Application Of Cellulase For Fiber Modification And Its Mechanism

The increasing widely application of environmental friendly processes in pulping and papermaking industry promots the study and development of biotechnology to be one of the most advancing frant research hotspots. The attractiveness of biotechnology lies in its potential of increasing the specificity of the enzymatic reaction, providing the environmental friendly processes which lead to the reduction of cost. Today, there are increasing researches involve in the application of cellulases in pulping and papermaking industry, and the modification of the cellulose fibers with cellulases can gain some favorable effects that other methods can hardly approach. The modification of cellulose fibers with cellulases is of highly application values, e.g. improving the drainage of the recycled fibers and speeding up the paper mechine; decreasing the energy requirement of the beating/refining process; modifying some mechanical properties of the papers; increasing the portion of filler in papers and saving the fiber materials; assisting the deinking and bleaching of the recycled fibers.The poplar alkaline peroxide mechanical pulp (APMP), softwood bleached sulfate Kraft pulp (NBKP) and mixed office waste (MOW) were modified with different kinds of enzymes so as to select the applicable enzymes for fibers modification and gain benefits according to the properties of the fibers and its end use. Optimization and statistics analysis of the process parameters with the response surface methodology (RSM) for investigating the effects of enzymatic modification of fibers on the saving of refining energy requirement, enhancing the drainage of the recycled fibers and improving of the paper physical properties were based on the single factor experiment of the major influence factors. The roles of cellulose binding domains (CBD) of the cellulases on the modification of the fibers were studied by digesting the molecular structure of the cellulase and separating the CBD from the cellulase molecular structure. The mechanisms of the fibers modification with cellulases were investigated with approaches of isothermal adsorption experiments, scanning electron microscope analysis, X-ray diffraction (XRD) analysis and thermogravimetric analyzer (TGA).The cellulase Nov476was proved to be the most effective enzyme for enzymatic refining of APMP, and its optimal processes conditions were:PFI refining time of90s, enzyme dosage of30u/g, temperature of55℃, pulp consistency of3%and enzymatic pretreatment time of1.5h. Enzymatic refining under these conditions could greatly improve the beatability of the pulp and save the refining energy. The beating degree of the Nov476pretreated pulp under the optimal conditions increased9unites after refining under a giver PFI revoluntions. And when the pretreated pulps were refined to the same beating degree with the control sample, approximate22%of the refining energy could be saved and without detectable influence on the mechanical properties of the handsheets. Observation of the fibers by optical microscope, SEM and analysis of fiber morphology indicated that after the enzymatic beating, the fibers surface showed apparent external fibrillation, fibers were widened due to the increased swollen, more fines and fiber fragments were detected.Cellulase Refinase M and NS51101were selected for the enzymatic refining of the NBKP, and the former contains CBD in it. Through the single factor experiments and RSM analysis, the optimal conditions of the enzymatic refining of the NBKP were reveled as: enzyme dosage of NS51101was0.03%and the contact time was97min; enzyme dosage of Refinase M was0.02%and the contact time was100min. The predicted beating degrees of the pulp from the RSM model were in aggrement with the actual values, indicating that the model can be used to predict the design space. Approximate18.5%and15.8%of the refining energy could be saved when pulps were pretreated with the cellulase Refinase M and NS51101and refined to the same or close beating degree. However, pretreatment with cellulase of Refinase M was proved to be more energy efficient when the pulps were refined under the same PFI revolutions. SEM and fiber morphology analysis suggested that after the enzymatic refining, the fibers lumen were collapsed and flattened, the fiber surface became rough and attached with microfiber, fibers was shortened and straightened (kink index decreased and shape factor increased). Analysis of the water resistency of the handsheets by dynamic contact angle showed that enzymatic refining increased the beating degree and consequently enhanced the water resistency, and the cellulase Refinase M pretreated sample exhibited the best water resistency.Cellulase Refinase S and NS51101were proved to be the most effective enzymes for inproving the pulp drainage and decreasing the fiber hornification degree. Through the single factor experiments and RSM analysis, the optimal conditions for the enzymatic modification of the pulp drainage were reveled as:enzyme dosage of NS51101was0.01%, pulp consistency was4.90%and contact time was34.6min; enzyme dosage of Refinase S was0.02%, pulp consistency was3.00%and contact time was40.0min. Both the two cellulase pretreatment decreasd the pulp beating degree approximate by10unites and decreased the fiber hornification. The average fiber length were increased and fines content decreased-after the cellulases pretreatments, and tear indexes of the handsheets decreased slightly but the tensile indexes remained unchanged even under lower beating degree. Dynamic drainage time of the pulps decreased to17.4s and15.6s from23.2s after the cellulase NS51101and Refinase S pretreatment, i.e. dynamic drainage property increased25%and32.8%respectively. During the single factor experiments, by comparision of the two cellulases for modification of the pulp we found that the cellulase NS51101was more effective in improving the drainage and Refinase S was more effective in decreasing the hornification of the fibers. SEM analysis of the fiber surface suggested that the cellulases pretreatment could greatly destroy the crust covered the fiber surface, restoring and activing the rough fiber surface.The cellulases rapidly adsorbed onto the fibers surface at the beginning of the adsorption (within10min) and reached an adsorption equilibrium at30min. Formula derivation and linear fitting between the cellulase adsorption amount and substrate consistency revelaed that the cellulase adsorption parameter of a were0.4279for Refinase M which contains CBD and0.2972for NS51101which lacks of CBD. CBD solution and cellulase lack of CBD were separated by limited digestion of Refinase M which contains CBD with papain, and the adsorption parameter (a) of intact cellulase, CBD and cellulase lack of CBD were0.3381,0.6569and0.2199respectively. Analysis of enzymatic properties indicated that the CBD showed almost no activity on the substrate, and the removing of CBD from the cellulase significantly decreased its activity on the insoluble substrate but had no influence on the soluble substrate. Cellulase lack of CBD required longer adsorption time to reach the adsorption equilibrium than the intact cellulase. Results of isothermal adsorption experiments of cellulase onto the fibers and analysis of the cellulose crystallinity by X-RD suggested that the roles of CBD in the modification of fibers with cellulase were:①to enhance the adsorption efficiency of cellulases onto the fibers, creating an stable surrounding for enzymatic catalytic hydrolysis.②to split the crystal cellulose, increasing the accessibility for hydrolyzing of cellulose. Analysis of the fiber hornification with thermal gravimetric analysis (TGA) showed that the hard-to-remove (HR) water content increased from1.42to2.41(g water/g o.v. fibers) after the cellulase pretreatment, confirming that the cellulases are capable of decreasing fiber hornification degree and increasing the fiber swollen ability.