| The cost of cellulase in the fuel ethanol production accounts for a high proportion. The production of cellulase from effective pretreatment of lignocellulosic biomass offers a lower cost than lactose, microcrystalline cellulose, etc. And optimization of culture technology can effectively reduce the production cost of cellulase. In this thesis, we used the rice straw as raw materials, varied the saturated steam pretreatment parameters such as NaOH concentration, steam pressure, incubation time and obtained the pretreatment lignocellulosic variants with different contents of cellulose, hemicellulose & lignin and macro & micro-structures.These variants were used as the starting carbon source & inducer precursors of Trichoderma reesei RutC-30 to syudy the cellulase & hemicellulase productivities with recalcitrancy of lignocellulosics due to micro-structure shielding & contents of cellulose, hemicellulose & lignin that might eventually change the inducers compositions & contents for cellulase & hemicellulase induction. The main research results are as follows:The morphology of rice straw has been changed dramatically by saturated steam pretreatment. Much slimmer and dark straw was formed. Scanning Electronic Microscope(SEM) data showed that rough & messy surface appeared offer the saturated steam pretreatment in contrast with the raw straw that has neatly & smoothly surface. With severity of the pretreatment increased, the lignocellulosic structure became even more irregularly. For few of them, the structure was almost near to collapse.Compositions analysis showed that hemicellulose in straw reached 4.01%, dropped 85.81%, the remove rate reached 89.49% and the weight loss rate remained only 24.44% during the pretreatment of two stadges 0.5MPa 10 min, then 2.0MPa 10 min. Under the pretreated conditions(5-100g/L NaOH, 0.5MPa, 10min), the weight loss rate, lignin & hemicellulose remove rate increase and the range of variation are(12.67%-51.33%),(18.03%-85.31%) &(31.16%-74.91%) respectively; the lignin and hemicellulose content decrease(24.25%-7.32%) &(28.26%-14.74%) sharply. Under the pretreated condition(100g/L NaOH, 0.5MPa, 10min), the weight loss rate, lignin & hemicellulose remove rate reached up to 51.33%, 85.31% & 74.91 respectively. Under the pretreated conditions(25g/L NaOH, 0.5-2.0MPa, 5-30min) the cellulose content kept the similar level(44.16%-46.19%) regardless of the severity of pretreatment, the weight loss rate was about 25% and remove rates of lignin and hemicellulose were over 50%. The weight loss of cellulose remained stable(~2%) throughout the entire pretreated process and cellulose was almost preserved.Cellulase is produced as the secondary metabotile at the growth phase of fungus and lignocellulosic substrate provide the the starting carbon source & inducer precursors. However, as we anticipated that non-pretreated rice straw barely induced any cellulase & hemicellulase presumably because the insusceptibility of the rice straw to the Trichoderma originated from the shielding structure of cellulose render the tough start-up of the fungus growth. Much more different pretreated rice straw with different severity of pretreatment were used as the sole carbon source for enzyme production. It appeared that those rice straw pretreated of two stadges(0.5MPa 10min+2.0MPa 10min) as shown from macro-and micro-structure & residual hemicell--ulose content has the highest cellulase activities. FPA, CMCA and xylanase activity reached 0.72 IU/mL, 11.59 IU/mL and 25.01 IU/mL, respectively. In this case, fungus growth havily relied on the level of de-construction of lignocellulose structure. Therefore, we designed the additional experiments. we used the rice straw easy start-up of the fungus growth as the carbon source and added rice strae with different composition and structure at 3th, 4th, 5th & 6th day in fed-batch, respectively and we found that cellulase production improved most at 5th day in fed-batch. Presumably, many enzymes has been formed at 5th day, the shielding structure has been weakened and composition and content of inducer played a crucial role in the final enzyme activity. Furthermore, cellulase production improved significantly added 3g/L pretreated straw(0.5MPa 10min+2.0MPa 10min) at 5th day in fed-batch. FPA and CMCA reached 1.01 IU/mL and 19.06 IU/mL, respectively; Addition 3g/L hemicellulose(78.23%) improved xylanase activity reaching 43.67IU/mL.10L-fermenter was used for cellulase production with Trichoderma reesei C-30 using straw(0.5MPa 10min+2.0MPa 10 min pretreated) as the starting carbon. The optimal fermentation condition was the same as shake-flask experiment. The cellulase production in fermenter was higher than that in shake-flask. The FPA and CMCA reached 1.31 IU/mL and 20.22 IU/mL by adding 5g/L pretreated straw(0.5MPa 10min+2.0MPa 10min) in fermenter. Addition 3g/L hemicellulose(78.23%) improved xylanase activity reaching46.65IU/mL. |
PDF Full Text Request