纳米铁—反硝化细菌复合体系修复地下水中NO_3~--N污染的研究
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摘要
大量氮肥的施用,生活污水和含氮工业废水的排放,使得饮用水源和地下水的硝酸盐氮污染日益严重,已成为全世界关注的重要的环境问题之一。地下水中的硝酸盐本身对人体并没有危害,但硝酸盐在人体内可经硝酸盐还原酶作用生成亚硝酸盐,从而对人体健康构成威胁。
有关地下水中硝酸盐污染物的去除技术国内外已有较多研究,主要可分为物理化学法、化学还原法和生物反硝化法。然而,物理化学方法只是简单地将硝酸盐进行浓缩或转移,并没有将其从地下水环境中去除；化学还原法对于产物中氨氮比例过高的问题始终找不到彻底解决的办法；生物反硝化法中异养微生物对于有机物的依赖限制了异养反硝化方法在地下水修复中的应用,而地下水中的自养微生物在反硝化过程中又缺乏足够的电子供体以还原硝酸盐。因此,单独使用任何一种方法都无法得到令人满意的处理效果。
针对以上问题,本文采用液相还原法制备出纳米铁颗粒,并将其与真养产碱杆菌(一种自养反硝化细菌)联合构建脱氮复合体系,以纳米铁在水中厌氧腐蚀过程中释放的氢气供给微生物反硝化,将硝酸盐氮转变为无害的氮气,可以同时解决脱氮产物中氨氮比例过高和生物反硝化缺少电子供体这两方面的缺陷。
本文通过一系列的实验来研究纳米铁-反硝化细菌复合体系的脱氮性能、影响因素、改进方法、动力学以及反应机理,并在此基础上尝试进行纳米铁和反硝化细菌的耦合共栖技术的开发,为实际地下水环境修复中的应用奠定基础。研究结果表明：
1.反硝化细菌的加入对于纳米铁去除硝酸盐的反应产生很明显的影响。单独使用纳米铁仅仅需要2天的时间即可将硝酸盐完全降解,但是其中大约95%的硝酸盐氮转变为氨氮；而加入反硝化细菌之后,虽然去除硝酸盐的反应时间从2天增加到8天,但是只有约为33%的硝酸盐氮转变为氨氮。在此过程中氨氮的含量并不是一直增加的,而是在前四天里,体系中的氨氮含量从0增加到35%,此后,氨氮含量一直稳定在33-36%之间。
2.反应体系的初始pH值对纳米铁-反硝化细菌复合体系还原硝酸盐的反应速率产生明显的影响,随初始pH值的升高反应速率呈现出减慢的趋势,但对脱氮产物中氨氮的生成量并无太大影响。此外,随复合体系温度的升高,硝酸盐降解速率加快,但脱氮产物中氨氮的产量也有所上升。而溶解氧的含量高低对复合体系中硝酸盐还原速率的影响很小,但溶解氧含量过高或过低均会增加脱氮产物中氨氮的比例,适宜的DO水平在0.4 mg·L-1左右。
3.SO42-和ClO4离子的存在都会对纳米铁-反硝化细菌复合体系的脱氮效果产生干扰作用。随着体系中存在的SO42-和ClO4离子浓度的增加,复合体系脱氮速率出现降低,而且脱氮产物中氨氮的比例也会随之降低。
4.Ni和Cu这两种催化剂的引入均可以提高纳米铁-反硝化细菌复合体系的脱氮反应速率,且催化效率相差不大。但是,对于由纳米Fe/Ni双金属所构建的复合体系,反应结束后氨氮比例高达69%。另一方面,纳米Fe/Cu双金属复合体系中仅有39%的硝酸盐氮转变为氨氮。
5.在纳米铁-反硝化细菌复合体系中,当反应进行到第四天时,体系中硝酸盐降解反应的准一级速率常数(kobs)从0.0969d-1突然增大到0.4213 d-1,复合体系的脱氮反应过程很难用一个准一级反应方程式来表示,整个硝酸盐降解过程应该分段使用两个不同的准一级反应式来表示,其中第一个准一级反应式主要描述反应初期纳米铁对硝酸盐的化学还原作用,第二个准一级反应式主要描述反应后期微生物的反硝化脱氮作用。
6.反硝化细菌的引入对于脱氮体系中纳米铁的形貌和成分均产生较大的影响。未加菌时,纳米铁在脱氮反应之后,其表面为一层致密的簇状氧化层所覆盖,此时纳米铁材料的主要成分为Fe、Fe2O3、Fe3O4的混合体。然而,加入细菌之后再进行脱氮反应时,纳米铁的表面变为一层云状的疏松结构,其主要成分为非晶态的FeOOH。
7.脱氮反应中,复合体系总RNA含量测定及平板菌落计数的结果均显示,在复合体系中的细菌数量出现了先降低后升高的趋势。这说明在反应初期纳米铁对反硝化细菌产生明显的毒害作用,导致细菌的大量死亡；而后,随着纳米铁的消耗和氧化,毒性降低,细菌得以逐渐适应新环境开始进行生长。TEM的结果显示,纳米铁可以破坏细菌的细胞膜从而进入到细胞内部,造成细菌死亡。
8.从纳米铁-反硝化细菌复合体系中分离出来的耦合体可以在5天的时间里将硝酸盐完全去除,同时,随着体系中硝酸盐的降低,反硝化细菌的数量逐渐增多。此外,脱氮反应结束之后,溶液中NO3--N、NO2--N、NH4+-N含量均低于检出限,而反应器顶部的气体中N20-N的含量达到21.60 mg·L-1,占到初始NO3-N含量的95.65%,这就是说,利用纳米铁-反硝化细菌耦合体去除硝酸盐时,脱氮产物几乎全部为气态氮,水体中不含NO2-、NH4+等有毒脱氮副产物。
9.油酸钠包覆型纳米铁-反硝化细菌耦合体可以在6天的时间里将约50 mg-N·L-1的硝酸盐完全去除,其中仅有7%左右转化为氨氮,剩余93%的氮素很可能通过生物反硝化作用以气态氮的形式从水相中分离出去。而且,当硝酸盐完全去除时,反应体系中的总RNA含量由初始的0ng·μL-1增加到551ng·μL-1这就是说随着脱氮反应的进行,反硝化细菌的数量在不断地增加,反硝化活性得以不断地增强。
With great use of nitrogen-enriched fertilizers and abundant discharge of domestic sewage and waste water, nitrate pollution of groundwater and surface water has become a serious environment problem in many parts of the world. At elevated concentrations, nitrate intake causes methemoglobinemia in infants, and may also react with secondary or tertiary amines to form carcinogenic nitrosamines.
Popular methods used for nitrate removal include physical chemical method, chemical reductive method and biological denitrifying method. However, physical chemical methods can only help to remove the nitrate from the groundwater with no help to reduce it. As to the chemical reductive methods, they may also lead to the generation of large amounts of ammonium, which may result in greater degree of toxicity than the nitrate itself. Biological denitrification, however, is also likely to lead to excessive biomass and soluble microbial products that make it necessary to do secondary treatment before using it, especially, when heterotrophic bacteria are involved. That is to say, no methods can be properly used for nitrate removal without other treatment.
Based on these problems, an integrated nitrate treatment using nanoscale zero-valent iron (NZVI) and Alcaligenes eutrophus, which is a kind of hydrogenotrophic denitrifying bacteria, was conducted to reduce nitrate and decrease ammonium generation. When NZVI particles are immersed in water, their oxidation is coupled with the reduction of water derived protons to form cathodic hydrogen, which can be utilized by denitrifying populations to remove nitrate from groundwater.
The objectives of this work were to study the denitrifying performance in NZVI-bacteria system, and also to evaluate the influence factors, improving methods, kinetics and mechanism of this integrated system. Then, development of NZVI-bacteria compound was also carried out. The conclusions from these experiments were as follows:
1. Within 8 d, nitrate was removed completely in the reactors containing NZVI particles and bacteria while the proportion of ammonium generated was only 33%. That is a lower reduction rate but a smaller proportion of ammonium relative to that in abiotic reactors. It was also found that ammonium generation experienced a two-step process, involving an increasing period and a stable period.
2. Increasing pH decreased the reduction rate slightly, but did not have a distinguishable effect on the generation of ammonium in the range of pH 7-10. In addition, with the temperature increasing, both the rate of the nitrate removal and the generation of ammonium increased. Moreover, the rate of the nitrate reduction had no change with increasing DO level in the solution. However, too high or too low DO level enhanced the generation of ammonium, so the proper DO may maintain around 0.4mg·L-1.
3. The presence of sulfate or perchlorate could both inhibit the nitrate reduction in the combined system. As the initial concentration of sulfate or perchlorate elevated, both rate of nitrate removal and ammonium proportion in final products declined.
4. Adding Ni or Cu into the NZVI-bacteria composed system could improve the denitrification efficiency, and the rates of nitrate removal in these two systems were very nearly the same. However, about 69% of nitrate was converted into ammonium in the Ni-catalyzed composed system, while there was only 39% of ammonium generated in the Cu-catalyzed composed system.
5. During the first 4 d, nitrate reduction can be in very good agreement with a pseudo first-order reaction (kobs=0.0969 d-1). Afterwards, kobs of the denitrification increased into 0.4213 d-1 abruptly. Based on the results obtained, two sequential pseudo first-order expressions should be used to describe the nitrate removal rates in this integrated system.
6. Unlike the Fe3O4 and Fe2O3 coating of NZVI particles generated in the abiotic reactors, an incompact wave structure was found on the surface of nanoparticles in the biological reactors, and the major component was testified to be non-crystal FeOOH, which was the corrosion product in the reaction between iron and water.
7. The results of total RNA content and enumeration by plate count method showed that the amount of the bacteria in the NZVI-bacteria system had experienced a first decreased and then increased process, which indicated NZVI might have toxicity on bacteria growth. Also, TEM image showed the plasmamembrane of bacteria could be destroyed by NZVI when the bacteria contacted with NZVI particles.
8. Complete removal of nitrate was observed within 5 d when the compound was utilized, which was separated from NZVI-bacteria composed system, and 95.65% of the nitrate was converted into N2O without ammonium and nitrite remained. That is to say, all the nitrate could be reduced via a biological process without chemical reduction when the compound was used.
9. Using the compound including sodium oleate coated NZVI particles and the autotrophic bacteria could remove all the nitrate within 6 d, while only 7% of nitrate was converted into ammonium. Also, the total RNA content increased from 0 ng·μL-1 to 551 ng·μL-1 when the nitrate was removed completely.
有关地下水中硝酸盐污染物的去除技术国内外已有较多研究,主要可分为物理化学法、化学还原法和生物反硝化法。然而,物理化学方法只是简单地将硝酸盐进行浓缩或转移,并没有将其从地下水环境中去除；化学还原法对于产物中氨氮比例过高的问题始终找不到彻底解决的办法；生物反硝化法中异养微生物对于有机物的依赖限制了异养反硝化方法在地下水修复中的应用,而地下水中的自养微生物在反硝化过程中又缺乏足够的电子供体以还原硝酸盐。因此,单独使用任何一种方法都无法得到令人满意的处理效果。
针对以上问题,本文采用液相还原法制备出纳米铁颗粒,并将其与真养产碱杆菌(一种自养反硝化细菌)联合构建脱氮复合体系,以纳米铁在水中厌氧腐蚀过程中释放的氢气供给微生物反硝化,将硝酸盐氮转变为无害的氮气,可以同时解决脱氮产物中氨氮比例过高和生物反硝化缺少电子供体这两方面的缺陷。
本文通过一系列的实验来研究纳米铁-反硝化细菌复合体系的脱氮性能、影响因素、改进方法、动力学以及反应机理,并在此基础上尝试进行纳米铁和反硝化细菌的耦合共栖技术的开发,为实际地下水环境修复中的应用奠定基础。研究结果表明：
1.反硝化细菌的加入对于纳米铁去除硝酸盐的反应产生很明显的影响。单独使用纳米铁仅仅需要2天的时间即可将硝酸盐完全降解,但是其中大约95%的硝酸盐氮转变为氨氮；而加入反硝化细菌之后,虽然去除硝酸盐的反应时间从2天增加到8天,但是只有约为33%的硝酸盐氮转变为氨氮。在此过程中氨氮的含量并不是一直增加的,而是在前四天里,体系中的氨氮含量从0增加到35%,此后,氨氮含量一直稳定在33-36%之间。
2.反应体系的初始pH值对纳米铁-反硝化细菌复合体系还原硝酸盐的反应速率产生明显的影响,随初始pH值的升高反应速率呈现出减慢的趋势,但对脱氮产物中氨氮的生成量并无太大影响。此外,随复合体系温度的升高,硝酸盐降解速率加快,但脱氮产物中氨氮的产量也有所上升。而溶解氧的含量高低对复合体系中硝酸盐还原速率的影响很小,但溶解氧含量过高或过低均会增加脱氮产物中氨氮的比例,适宜的DO水平在0.4 mg·L-1左右。
3.SO42-和ClO4离子的存在都会对纳米铁-反硝化细菌复合体系的脱氮效果产生干扰作用。随着体系中存在的SO42-和ClO4离子浓度的增加,复合体系脱氮速率出现降低,而且脱氮产物中氨氮的比例也会随之降低。
4.Ni和Cu这两种催化剂的引入均可以提高纳米铁-反硝化细菌复合体系的脱氮反应速率,且催化效率相差不大。但是,对于由纳米Fe/Ni双金属所构建的复合体系,反应结束后氨氮比例高达69%。另一方面,纳米Fe/Cu双金属复合体系中仅有39%的硝酸盐氮转变为氨氮。
5.在纳米铁-反硝化细菌复合体系中,当反应进行到第四天时,体系中硝酸盐降解反应的准一级速率常数(kobs)从0.0969d-1突然增大到0.4213 d-1,复合体系的脱氮反应过程很难用一个准一级反应方程式来表示,整个硝酸盐降解过程应该分段使用两个不同的准一级反应式来表示,其中第一个准一级反应式主要描述反应初期纳米铁对硝酸盐的化学还原作用,第二个准一级反应式主要描述反应后期微生物的反硝化脱氮作用。
6.反硝化细菌的引入对于脱氮体系中纳米铁的形貌和成分均产生较大的影响。未加菌时,纳米铁在脱氮反应之后,其表面为一层致密的簇状氧化层所覆盖,此时纳米铁材料的主要成分为Fe、Fe2O3、Fe3O4的混合体。然而,加入细菌之后再进行脱氮反应时,纳米铁的表面变为一层云状的疏松结构,其主要成分为非晶态的FeOOH。
7.脱氮反应中,复合体系总RNA含量测定及平板菌落计数的结果均显示,在复合体系中的细菌数量出现了先降低后升高的趋势。这说明在反应初期纳米铁对反硝化细菌产生明显的毒害作用,导致细菌的大量死亡；而后,随着纳米铁的消耗和氧化,毒性降低,细菌得以逐渐适应新环境开始进行生长。TEM的结果显示,纳米铁可以破坏细菌的细胞膜从而进入到细胞内部,造成细菌死亡。
8.从纳米铁-反硝化细菌复合体系中分离出来的耦合体可以在5天的时间里将硝酸盐完全去除,同时,随着体系中硝酸盐的降低,反硝化细菌的数量逐渐增多。此外,脱氮反应结束之后,溶液中NO3--N、NO2--N、NH4+-N含量均低于检出限,而反应器顶部的气体中N20-N的含量达到21.60 mg·L-1,占到初始NO3-N含量的95.65%,这就是说,利用纳米铁-反硝化细菌耦合体去除硝酸盐时,脱氮产物几乎全部为气态氮,水体中不含NO2-、NH4+等有毒脱氮副产物。
9.油酸钠包覆型纳米铁-反硝化细菌耦合体可以在6天的时间里将约50 mg-N·L-1的硝酸盐完全去除,其中仅有7%左右转化为氨氮,剩余93%的氮素很可能通过生物反硝化作用以气态氮的形式从水相中分离出去。而且,当硝酸盐完全去除时,反应体系中的总RNA含量由初始的0ng·μL-1增加到551ng·μL-1这就是说随着脱氮反应的进行,反硝化细菌的数量在不断地增加,反硝化活性得以不断地增强。
With great use of nitrogen-enriched fertilizers and abundant discharge of domestic sewage and waste water, nitrate pollution of groundwater and surface water has become a serious environment problem in many parts of the world. At elevated concentrations, nitrate intake causes methemoglobinemia in infants, and may also react with secondary or tertiary amines to form carcinogenic nitrosamines.
Popular methods used for nitrate removal include physical chemical method, chemical reductive method and biological denitrifying method. However, physical chemical methods can only help to remove the nitrate from the groundwater with no help to reduce it. As to the chemical reductive methods, they may also lead to the generation of large amounts of ammonium, which may result in greater degree of toxicity than the nitrate itself. Biological denitrification, however, is also likely to lead to excessive biomass and soluble microbial products that make it necessary to do secondary treatment before using it, especially, when heterotrophic bacteria are involved. That is to say, no methods can be properly used for nitrate removal without other treatment.
Based on these problems, an integrated nitrate treatment using nanoscale zero-valent iron (NZVI) and Alcaligenes eutrophus, which is a kind of hydrogenotrophic denitrifying bacteria, was conducted to reduce nitrate and decrease ammonium generation. When NZVI particles are immersed in water, their oxidation is coupled with the reduction of water derived protons to form cathodic hydrogen, which can be utilized by denitrifying populations to remove nitrate from groundwater.
The objectives of this work were to study the denitrifying performance in NZVI-bacteria system, and also to evaluate the influence factors, improving methods, kinetics and mechanism of this integrated system. Then, development of NZVI-bacteria compound was also carried out. The conclusions from these experiments were as follows:
1. Within 8 d, nitrate was removed completely in the reactors containing NZVI particles and bacteria while the proportion of ammonium generated was only 33%. That is a lower reduction rate but a smaller proportion of ammonium relative to that in abiotic reactors. It was also found that ammonium generation experienced a two-step process, involving an increasing period and a stable period.
2. Increasing pH decreased the reduction rate slightly, but did not have a distinguishable effect on the generation of ammonium in the range of pH 7-10. In addition, with the temperature increasing, both the rate of the nitrate removal and the generation of ammonium increased. Moreover, the rate of the nitrate reduction had no change with increasing DO level in the solution. However, too high or too low DO level enhanced the generation of ammonium, so the proper DO may maintain around 0.4mg·L-1.
3. The presence of sulfate or perchlorate could both inhibit the nitrate reduction in the combined system. As the initial concentration of sulfate or perchlorate elevated, both rate of nitrate removal and ammonium proportion in final products declined.
4. Adding Ni or Cu into the NZVI-bacteria composed system could improve the denitrification efficiency, and the rates of nitrate removal in these two systems were very nearly the same. However, about 69% of nitrate was converted into ammonium in the Ni-catalyzed composed system, while there was only 39% of ammonium generated in the Cu-catalyzed composed system.
5. During the first 4 d, nitrate reduction can be in very good agreement with a pseudo first-order reaction (kobs=0.0969 d-1). Afterwards, kobs of the denitrification increased into 0.4213 d-1 abruptly. Based on the results obtained, two sequential pseudo first-order expressions should be used to describe the nitrate removal rates in this integrated system.
6. Unlike the Fe3O4 and Fe2O3 coating of NZVI particles generated in the abiotic reactors, an incompact wave structure was found on the surface of nanoparticles in the biological reactors, and the major component was testified to be non-crystal FeOOH, which was the corrosion product in the reaction between iron and water.
7. The results of total RNA content and enumeration by plate count method showed that the amount of the bacteria in the NZVI-bacteria system had experienced a first decreased and then increased process, which indicated NZVI might have toxicity on bacteria growth. Also, TEM image showed the plasmamembrane of bacteria could be destroyed by NZVI when the bacteria contacted with NZVI particles.
8. Complete removal of nitrate was observed within 5 d when the compound was utilized, which was separated from NZVI-bacteria composed system, and 95.65% of the nitrate was converted into N2O without ammonium and nitrite remained. That is to say, all the nitrate could be reduced via a biological process without chemical reduction when the compound was used.
9. Using the compound including sodium oleate coated NZVI particles and the autotrophic bacteria could remove all the nitrate within 6 d, while only 7% of nitrate was converted into ammonium. Also, the total RNA content increased from 0 ng·μL-1 to 551 ng·μL-1 when the nitrate was removed completely.
引文
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