Abstract:

In the rice paddy field ecosystem, the increase of hydrogen’s concentration can not only promote the metabolic activity of soil microorganisms, but also promote the growth of rice and reduce the accumulation of heavy metals in rice. So, it’s of great significance to study the hydrogen metabolism of hydrogen-producing bacteria in the paddy soil environment. As iron oxide is widely distributed in rice paddy fields, ferrihydrite has been shown to promote microbial hydrogen production. However, the buffering effect of high concentration of ferrihydrite on pH in existing studies has restricted the in-depth analysis of its mechanism. Therefore, this study focused on low concentration (0-50 mg?L^?1 concentration range) ferrihydrite. First, we studied the effect of ferrihydrite at different concentrations on the hydrogen production of Clostridium, the enzymatic activity and biomass of hydrogen-producing bacteria. In addition, we monitored the changes of main metabolites, glucose content and Fe (II) content in the fermentation broth. The results showed that in the low amount range, ferrihydrite had no obvious pH buffering effect on the dark fermentation system, but could effectively promote the production of hydrogen. The optimum addition amount of ferrihydrite was 15 mg?L^?1. Under this condition, hydrogen production increased by 85.34% compared with the blank control group. The utilization rate of glucose increased from 77.66% of the blank control group to 91.40%. The maximum biomass and hydrogen-producing enzyme activity of Clostridium were 1.82 times and 1.80 times that of the blank control group, respectively. The production of acetic acid and butyric acid of Clostridium increased, while the production of lactic acid decreased. Compared with the blank group, the production of acetic acid and butyric acid increased by 71.42% and 28.98%, respectively, and the production of lactic acid decreased by 12.55%. The Fe (II) content in the culture system increased with the reaction time, with the highest concentration of 6.36 mg?L^?1, but the total electron consumption for iron reduction only accounted for 0.12% of the electron consumption for hydrogen production. In conclusion, low concentration ferrihydrite can still significantly promote the hydrogen production of Clostridium without obvious pH buffering effect. The potential mechanisms of low concentration ferrihydrite to promote hydrogen-producing of Clostridium are as follows: (1) to promote the growth of Clostridium and the utilization of glucose; (2) to increase the enzymatic activity of hydrogenase; and (3) to regulate its metabolism in favor of hydrogen-producing. The results can provide a theoretical basis for in-depth understanding of the effect of ferrihydrite on hydrogen-producing microorganisms in the natural environment.

Key words: rice fields, ferrihydrite, Clostridium, hydrogen production, iron reduction, pH

摘要：

在水稻田生态系统中，氢气浓度的增加不仅可以提升土壤微生物的代谢活性，还可以促进水稻生长，降低水稻对重金属的积累，因而研究水稻田中产氢菌的产氢代谢具有重要的意义。水铁矿作为水稻田中广泛分布的铁氧化物，已有研究表明它可以促进微生物产氢，但现有研究中高浓度水铁矿对pH的缓冲效果制约了对其机制的深入剖析。因此，该研究以低量（0—50 mg?L^?1浓度范围内）水铁矿为研究重点，以稻田来源产氢梭菌为研究对象，通过研究不同浓度的水铁矿对梭菌的产氢量、产氢酶酶活性和生物量的影响，结合发酵液中的主要代谢产物、葡萄糖含量和Fe(II) 含量变化，对水铁矿的作用机制进行深入探究。研究表明，在低量范围内，水铁矿对梭菌暗发酵体系无明显的pH缓冲效果，但能有效促进氢气的产生。水铁矿最佳投加量为15 mg?L^?1，此条件下总产氢量比空白对照组高85.34%；葡萄糖利用率从空白对照组的77.66%提升至91.40%；梭菌最大生物量是空白对照组的1.82倍；产氢酶酶活性是空白对照组的1.80倍；梭菌产乙酸和产丁酸量增加，产乳酸量降低，其中乙酸和丁酸产量分别较空白组提高71.42%、28.98%，乳酸产量降低12.55%；培养体系Fe(II) 含量随反应时间不断增加，最高浓度为6.36 mg?L^?1，但铁还原电子消耗总量仅占产氢电子消耗的0.12%。综上，低量水铁矿在无明显pH缓冲效果的情况下仍能够显著促进梭菌产氢，其促进梭菌产氢的潜在机制有：（1）促进梭菌的生长及葡萄糖的利用；（2）提高产氢酶酶活性；（3）调节梭菌代谢向有利于产氢的方向转变。研究结果可为深入了解水铁矿在自然环境中对产氢微生物的影响提供理论基础。

关键词: 水稻土, 水铁矿, 梭菌, 产氢, 铁还原, pH