Study On Hot Compressed Water Pretreatment, Enzymolysis And Succinic Acid Fermentation Of Corn Stover

In China, annual corn stover output is about 0.17 billion tons. Large amouts of corn stover was incinerated, which caused searious environmental pollution. Corn stover contains large amounts of cellulose and semicellulose, which can be transformed to biochemicals. The key step was the conversion of lignocellulose to fermentable sugars. However, the structure of lignocellulose, which was composed of lignin, cellulose and hemicellulose, was very tight. Pretreatment could destruct lignin and hemicelluloses, thereby increasing the contact area of the enzyme molecule. There are many problems existed currently in the whole process. The pretreatment effect was low. Fermentation inhibitors produced is in high concentration. The cost of cellulase is high. The yield of succinic acid is low. In the meantime, waste water contains sulfate from pretreament and succinic acid fermentation needed to be treated. This dissertation aimed to optimize the pretreatment process, improve the rate of enzymatic hydrolysis, increase succinic acid yield and treat waste water by microbial desulfurization.First, hot compressed water pretreatment (HCW) of corn stover was established. SEM photos indicated that the HCW pretreatment could effectively destroy the structure of lignocellulose. More than 90% of hemicellulose was removed with HCW pretreatment. Ammonium sulfate added in the process of HCW pretreatment could thoroughly destroy inner structure of lignin in corn stover. Therefore, cellulose crystalline region was greatly exposed, which benefited to the adhesion of cellulase. The optimum reaction temperature and solid-liquid ratio was 185℃ and 10%, respectively. In the process of HCW pretreatment, only trace of fermentation inhibitors were found. The enzymolysis rate reached 85% in fed batch and the glucose concentration reached 40 g/LSecondly, anaerobic fermentation process with A. succinogenes BE-1 was optimized. Feeding strategy had been optimized. The highest conversion rate of succinic acid had been observed when the initial glucose concentration was 40～50 g/L and using 10 g/L yeast extract and 6 g/L CSL compound (C/N=30) as nitrogen source. Low concentration (0.05～0.1 mol/L) of Na+ had a certain role in promoting on the growth and acid production by A succinogenes BE-1, but high concentration (0.2-0.3 mol/L) of Na+ had an obvious inhibitory effect. In the anaerobic fermentation, MgCO3 was used as pH-buffer, which could effectively alleviate the inhibitory effect of high concentrations sodium ion, and thereby improved the succinic acid synthesis. Succinic acid titer in the fed-batch fermentation, which was 60 g/L 48h, increased by 100% compared to the batch fermentation, which was 30 g/L after 48h.Thirdly, a sulfate reducing strain, Citrobacter sp. HCSR (CGMCC NO.4660), was isolated. It was a facultative anaerobe.16S rDNA gene sequence tree showed that it belonged to the genus Citrobacter. The optimum growth temperature and pH were 30℃ and 6.4-6.8, respectively. The optimum initial pH was 6.4 for desulfurization reactions. The maximum OD600 under aerobic conditions was 3.5 times higher than under anaerobic conditions. But the desulfurization ability of strain HCSR was much lower than anaerobic condition. Using the dual-phase method, the total sulfate removal reached 21 mmol/L after 7 days, which was 79% higer than at anaerobic conditions.Finally, Fe3O4 magnetic nanoparticles were synthesized with coprecipitation method. Then, SiO2 was coated on the MNPs surface layer with the sol-gel method. The dispersion degree of the particles was greatly improved. Sulfate removal activity of HCSR coated with MNPs was studied. The maximum amount of desulfurization reached 35 mmol/L with immobilized cells, which was 75% hihger than free cells. The effects of SiO2/Fe3O4 ratio on the desulfurization efficiency of the immobilized cells were studied. The cells immobilized with MNPs coated with 300% SiO2 showed the highest desulfurization activity. The sulfate removal rate reached 92% by 7 days. Four batches of desulfurization with immobilized cells were studied and the sulfate removal rates were 94%,82%,57% and 24%, respectively. The overall reaction time lasted 24 days and the total desulfurization concentration of SO42- reached 65.5 mmol/L, which was 450% higher than free cells.