【作者】 李丹；

【导师】 秦智；

【作者基本信息】 哈尔滨理工大学 ， 应用化学， 2013， 硕士

【摘要】 生物制氢法弥补了传统制氢法的诸多缺点而得到重视,厌氧发酵制氢是生物制氢的一种方法,具有产氢效率高、产氢效果稳定且价格低廉等优点。从本实验室CSTR中分离到一株产氢菌,命名为Pectinatus sp.L6。本实验考察了在间歇产氢装置中培养温度对产氢菌L6发酵产氢的影响,并研究了在五种不同培养温度下pH值、干重、液相末端产物的变化规律,同时研究了产氢菌L6脱氢酶活性即DHA的测定条件以及培养温度对DHA的影响。产氢菌L6产氢能力的研究结果表明:产氢菌L6在五个不同培养温度下生长及产氢能力最好的是35℃,其它培养温度下产氢菌L6生长及产氢能力由大到小的顺序依次为30℃、40℃、25℃、45℃。在35℃的培养温度下,产氢菌L6的产气速率在培养33h时有最大值84.7mL/h,产氢菌L6的产氢速率在培养7h时迅速增加并在培养17h时基本稳定,产氢速率在培养34h时有最大值72.1mL/(L·h),在培养64h时产氢菌L6几乎不产氢;在培养40h时,产氢菌L6单位时间干重的增加量有最大值579mg/L;产氢菌L6的初始pH值为6.13,培养12h时pH值急剧下降至4.37,培养64h时基本稳定在4.45;产氢菌L6液相末端发酵产物的百分含量由大到小的顺序依次为丁酸、乙酸、丙酸、乙醇,培养28h时丁酸的百分含量达最大值86.1%,此时乙酸、丙酸和乙醇的百分含量依次为8.3%、3.9%和1.3%,产氢菌L6的发酵类型为丁酸型发酵。其它四个培养温度下各指标虽然没有培养35℃时的效果好,但是基本规律与培养35℃时保持一致,在培养温度分别为3℃、40℃、25℃和45℃时,产氢菌L6的最大产氢速率分别为59.0mL/(L·h)、51.0mL/(L·h)、31.8mL/(L·h)、23.9mL/(L·h)。通过对DHA的四种测定方法优缺点的对比,最终选择TTC-DHA法测定产氢菌L6的DHA。研究结果表明,TTC-DHA法中产氢菌L6的DHA最佳测定条件为:菌液的离心转速为6000rpm,离心时间为10min,最佳吸收波长为480nm,水浴温度35℃,显色反应时间4h,TF萃取时间为1Omin,萃取剂丙酮的最佳加入量为5mL,TTC-DHA法标准曲线的还原时间为10min,Tris-HC1(pH=8.4)、TTC 标液、Na2SO3(0.36%)溶液和 Na2S204 的最佳加入量依次为2mL、2mL、1mL和5mg。产氢菌L6在五个不同培养温度下DHA由大到小的顺序依次是:35°C、30°C、40°C、25°C、45°C,说明产氢菌L6的产氢能力及生长状况越好,其DHA也越高。培养35°C时产氢菌L6的DHA最高,DHA增加、稳定在最佳范围内及有所下降的三个时期分别为:Oh至24h,28h至48h,52h至64h。初始TF值为27mg/L,在培养32h时有最大值133mg/L,在培养64h时下降至94mg/L。更多还原

【Abstract】 Bio-hydrogen production was paid attention to its advantages,anaerobic fermentation hydrogen production has the advantages of high hydrogen production capacity,stable hydrogen production and et al.A hydrogen producing bacterium was isolated from the CSTR,and was termed as Pectinatus sp.L6.Batch culture for fermentation hydrogen production of hydrogen producing bacteria L6 were examined with cultured temperature,and the hydrogen producing bacteria L6 was researched in five cultured temperature,including pH value,dry weight,terminal aqueous productions.The determination condition and cultured temperature of DHA was researched simultaneously.The results show that the optimal culture temperature was 35℃,and the order of culture temperature of hydrogen production capacity was as follow:30℃ 40℃,25℃,45℃.Under the temperature of 35℃ condition,The maximum biogas production rate and hydrogen production rate of hydrogen producing bacteria L6 was 84.7mL/h and 72.1mL/(L-h)at 33h,respectively.The maximum increment of dry weight per unit time was 579mg/L at 40h.The pH value was declined to 4.37 at 12h from the initial 6.13,and was stabilized to 4.45 at 64h.The order of percentage of the terminal aqueous productions was as follow:butyric acid,acetic acid,propionic acid and ethanol.The maximum percentage of butyric acid was 86.1%at 28h,and at the same time the percentage of acetic acid,propionic acid,ethanol was 8.3%、3.9%,1.3%,respectively.The fermentation type was butyric acid type.The rules of biogas production rate,hydrogen production rate,pH value,dry weight and terminal aqueous productions at other temperatures was similar with 35℃.But the hydrogen producting was the best at 35℃.At 30℃,40℃,25℃ and 45℃ the maximum hydrogen production rate was 59.0mL/(L·h),51.0mL/(L·h),31.8mL/(L·h),23.9mL/(L-h),respectively.The TTC-DHA method was choosed by comparing with the four measured methods of DHA.The results show that the optimum conditions of TTC-DHA were as follow:the centrifugal revolving speed was 6000rpm,the centrifugal time was 10min,the maximum absorption peak was at the point of 480nm,the bath temperature was 35℃,the reaction time was 4h,the extraction time was 10min,the optimal addition of extractant was 5mL,the reducing time of the standard curve of TTC-DHA method was 10 minutes,and the best dosage of Tris-HCl(pH=8.4),TTC,Na2SO3(0.36%)and Na2S2O4 was 2mL,2mL,1mL and 5mg.The order of culture temperature of DHA was as follow:30℃,40 ℃,25 ℃,45 ℃.The DHA of hydrogen producing bacteria L6 increased with increasing hydrogen production capacity and the DHA was the highest at 35℃.The increasing period,the stable period and the decreasing period of DHA was 0-24h,28-48h,52-64h,respectively.The initial TF value was 27mg/L,and the maximum TF value was 133mg/L at 32h,and declined to 94mg/L at 64h.更多还原