Hydrogen Production from Unutilized Biomass by Anaerobic Bacteria

该研究是在爱知县名古屋市地区的大学和社会合作资助金（MEXT）的支持下完成的，用于通过使用几丁质分解厌氧菌Clostridium paraputrificum M-21将固体废物微生物转化为氢气。通过遗传操作和培养条件的优化，提高了该菌的发酵能力，从而提高了虾壳产气能力。具体如下：1. C.从三重大学校园土壤中分离到的类营养菌M-21能在虾壳中的甲壳素主要高分子聚合物上生长。在N-乙酰氨基葡萄糖上培养时，其倍增时间约为30 min。克隆了几丁质酶和氢化酶的编码基因，并测定了它们的DNA序列。对大肠杆菌转化子产生的这些酶进行了表征.产气荚膜梭菌的宿主-载体系统成功地应用于转化副腐败梭菌.通过电穿孔将几丁质酶和氢化酶的基因整合到生物体中来强化副营养梭菌的几丁质酶和氢化酶。各转化子显示出比野生型更高的几丁质降解和氢气产生活性。氢气产量增加至约3 mol H2/mol葡萄糖。当气体被充入燃料电池时，由螺旋桨和螺旋桨产生的沼气由氢气（65%）和二氧化碳（35%）组成。构建了由两个发酵罐组成的双容器发酵罐，一个用于氢气生产，一个用于甲烷生产，并用于分别在6和7下优化pH。甲烷发酵，然后使用双容器发酵罐氢气发酵提高气体产量和减少固体废物。

The research was fulfilled under the support of the Grant-in-Aid for University and Society Collaboration (MEXT) in Aichi Prefecture-Nagoya City Area for microbial conversion of solid waste to hydrogen gas by using a chitinolytic anaerobe Clostridium paraputrificum M-21. The productivity of the gas from shrimp shell was increased by improving fermentation ability of the organism by genetic manipulation and by optimization of its cultivation conditions. The details are as follows :1. C. paraputrificum M-21 isolated from Mie University Campus soil can grow on chitin main high polymer in shrimp shell. Its doubling time was about 30 min when cultivated on N-acetylglucosamin.2. The genes encoding chitinases and hydrogenase were cloned and their DNA sequences were determined. These enzymes produced by the Escherichia coli transformants were characterized.3. The host-vector system for Clostridium perfringens was successfully applied to transform C.paraputrificum.4. Chitinases and hydrogenase of C.paraputrificum were fortified by integrating their genes into the organism by electroporation. The respective transformants revealed higher activities of chitin degradation and hydrogen gas production than wild ones. Hydrogen production was increased up to about 3 mol H2/mol glucose.5. The biogas produced by the transformant consisted of hydrogen (65%) and carbon dioxide (35%) and screwed propeller when the gas was charged to a fuel cell.6. A two-vessel fermentor consisting of two fermentors, one for hydrogen and one for methane production was constructed and used to optimize pHs at 6 and 7, respectively.7. Methane fermentation followed by hydrogen fermentation using the two-vessel fermentor enhanced gas production and reduction of solid waste.