【作者】 谢天卉；

【导师】 任南琪；

【作者基本信息】 哈尔滨工业大学 ， 环境科学与工程， 2006， 硕士

【摘要】 提高产氢效能是发酵法生物制氢技术工业化面临的关键问题之一。利用自凝集产氢细菌可以避免菌种流失,保证生物量,有利于提高系统产氢效能。因此研究自凝集产氢细菌的凝集机理和连续流扩大培养,作为发酵法生物制氢技术的辅助工艺,将具有重要的工程意义。本文利用离子交换树脂提取了哈尔滨产乙醇杆菌（Ethanoligenens harbinense）的自凝集和非凝集细菌的胞外聚合物,分析了自凝集细菌YUAN-3的凝集机理;进行了菌株YUAN-3分批和补料培养;在非灭菌条件进行了该菌株的连续流扩大培养,并通过DGGE对微生物群落进行了分析。胞外聚合物的最佳提取条件为树脂用量30g/g-CDW,搅拌机转速为400rpm,提取时间12h。胞外聚合物分析表明,与其他同属的非凝集产氢细菌相比,菌株YUAN-3的胞外聚合物在其自凝集过程中发挥了重要作用。在胞外聚合物中,蛋白质、碳水化合物、DNA的含量分别为20.1±0.2mg/g-CDW、67.5±0.8mg/g-CDW和27.4±0.5mg/g-CDW。在各种规模分批培养中,YUAN-3表现出相似的生长规律,即使扩大生产,仍然能维持较高的生长活性和产氢活性,生物量达0.33g/L,平均产氢速率最高达22.34mmol/（g-CDW?h）,比产氢率最高达1.4molH2/mol-葡萄糖。生物制氢反应器投菌的最佳时间是细菌培养30h左右。补料培养的最佳补料体积为发酵液体积的60%左右,最佳的补料时间为产气的高峰期,在此条件下投加细菌可以保证细菌具有较高的生长和产氢活性,并可以缩短延滞期。连续培养能够不断将发酵产物排出,引进新鲜培养基,避免底物的抑制。在YUAN-3扩大连续培养后,平均产氢速率为0.75mmolH₂/（g-CDW·h）⁵.60mmolH₂/（g-CDW·h）变化,比产氢率为0.35molH₂/mol-葡萄糖¹.84molH₂/mol-葡萄糖。DGGE分析表明,在运行初期就出现了很多非目的细菌,YUAN-3逐渐成为反应器的优势种群,在非灭菌和一定的生态条件下,该菌具有较强的生长竞争能力,可以成功的实现连续流产氢和培养,适合投加于大规模生物制氢反应器中。更多还原

【Abstract】 One of the key problems of industrial application of is how to increase the hydrogen producing efficiency. Addition of auto-aggregation hydrogen-producing will maintain the biomass and increase the hydrogen producing efficiency. Thus, research on aggregation mechanism of auto-aggregation hydrogen-producing and its continuous extended culture as assistant process of fermentive bio-hydrogen production technology will bring important practical value.Extracellular polymeric substances （EPS） of auto-aggregation and non- aggregation bacterium of Ethanoligenens harbinense were extracted by cation exchange resin, and aggregation mechanism was analysed. Bio-hydrogen progen production by pure culture and its extend culture of the strain of YUAN-3 were carried out by batch culture, fed-batch and continuous culture, and DGGE was used in monitoring the communities in continuous culture. The optimal conditions for extraction of EPS was resin addition of 30g/g-CDW, 400rpm, the extraction time was 12h. Compared with non-aggregation bacterium, EPS played important role in aggregation of YUAN-3, and in EPS, the content of protein, carbohydrates and DNA were 20.1±0.2mg/g-CDW, 67.5±0.8mg/g-CDW and 27.4±0.5mg/g-CDW, respectively.In different scale batch culture, YUAN-3 showed similar growth trends, though extended culture, YUAN-3 still kept higher growth and hydrogen-producing activity, the biomass was 0.33g/L, maximum specific hydrogen rate was 22.34mmol/（g-CDW·h）, maximum hydrogen yield was 1.4molH₂/mol-glucose. The best time of adding bacterium to the bio-hydrogen production reactors was around 30 hours after cultured. In fed-batch culture, the best fed volum was 60% of the working volum, the best time was peak time of hydrogen production, in above conditions, YUAN-3 kept higher growth and hydrogen-producing activity.Contiuous culture would discharge fermention products, indraught the fresh medium, avoid restraint of fermention products. When YUAN-3 were continuous extended culture, the specific hydrogen rates were between 0.75mmolH₂/（g-CDW·h）⁵.60mmolH₂/（g-CDW·h）, the hydrogen yield were between更多还原