固氮紅細菌對模擬水體中無機三態(tài)氮去除特性的研究
發(fā)布時間：2018-07-25 18:50
【摘要】：目前,海水養(yǎng)殖水體污染問題日益嚴重,其中氮污染是影響海水養(yǎng)殖業(yè)發(fā)展的首要問題,因此消除養(yǎng)殖水體氮污染、恢復微生態(tài)平衡,是解決養(yǎng)殖水體污染問題的關鍵。不產氧光合細菌(Anoxygenic phototrophic bacteria,APB)已在去除水體中的無機三態(tài)氮,恢復水體微生態(tài)平衡及提高養(yǎng)殖動物免疫力等方面顯示出重要作用。然而,APB處理的實際水體環(huán)境復雜,探求復雜水體環(huán)境中APB的除氮效果,具有重要的實際應用價值。目前,關于APB去除亞硝氮的能力、以及對養(yǎng)殖水體中三氮的利用和轉化研究報道較少。APB的實際應用中會受到水體中多種小分子有機氮源、碳源及鹽度等的影響。針對以上問題,本文以固氮紅細菌(Rhodobacter azotoformans)R7為材料,首先通過條件優(yōu)化提高生物量,為其應用奠定基礎;在此基礎上,進一步探究了該菌株對三種無機態(tài)氮的去除特性、對無機三態(tài)氮的利用與轉化能力以及無機三態(tài)氮對菌株生長和光合色素合成的影響;為了提高菌株的實際應用效果,又探究了復雜的水體環(huán)境因素,包括無機和有機氮化合物、有機碳化合物和鹽度對菌株生長以及除氮效果的影響。主要結果如下:通過單因子輪換試驗法(One-factor-at-a-time,OFAT)優(yōu)化獲得提高菌體生物量及Car產量的最適條件為:裝樣量為80%、3 g/L酵母粉、5 g/L蛋白胨、2.5 g/L葡萄糖。在最適條件下,菌體生物量由3.34 g/L提高到17.59 g/L,提高了4.27倍;菌體類胡蘿卜素含量由優(yōu)化前2.43 mg/L提高到30.31 mg/L,提高了11.47倍,主要積累球形烯、羥基球形烯及球形烯酮3種類胡蘿卜素。Car光穩(wěn)定性研究表明,含酮基的球形烯酮穩(wěn)定性較高,在0 lux、500 lux及3000 lux光照條件下其半衰期依次為:568.53 h、405.56 h及112.64 h。R7菌株對無機三態(tài)氮的去除與轉化結果顯示,R7菌株能利用氨氮、硝氮及亞硝態(tài)氮良好的生長,對硝氮和亞硝氮具有較強的耐受和去除能力。培養(yǎng)8天內,R7菌株對氨氮、硝氮和亞硝氮最大去除量分別為:6.10 mmol/L、21.36mmol/L和16.86 mmol/L,最大耐受濃度分別為43.47 mmol/L、52.17 mmol/L和34.78 mmol/L;在4.54 mmol/L氨氮、34.35 mmol/L硝氮和34.64 mmol/L亞硝氮共存的體系中,該菌株對氨氮、硝氮及亞硝氮的去除率分別為54.38%、31.48%和99.95%;對無機三態(tài)氮的利用和轉化的結果分析顯示,分別以氨氮、硝氮和亞硝氮為唯一氮源培養(yǎng)的過程中,均出現了另外兩種無機氮的暫時性積累,隨著培養(yǎng)時間的延長,逐漸被去除。上述結果表明,R7菌株存在多種氮代謝作用,具有反硝化作用、厭氧硝化作用和亞硝酸鹽厭氧氧化作用。氨氮、硝氮及亞硝氮對菌株生長和菌體光合色素合成的影響,結果表明:水體中氨氮(≤43.48 mmol/L)或硝氮(≤52.17 mmol/L)對菌體生物量及Car含量影響不明顯,低濃度氨氮(≤8.70 mmol/L)或硝氮(≤8.66 mmol/L)對菌株生長具有促進作用;上述結果表明,水體中氨氮和硝氮對菌株色素含量的影響不明顯,水體中亞硝氮濃度越高,對R7菌體色素的合成抑制越明顯。環(huán)境中可能存在的小分子碳(糖類、有機酸或醇)、有機氮(酵母浸出粉、蛋白胨或尿素)和鹽度影響菌株對無機三態(tài)氮的去除,結果表明:環(huán)境中的不同碳源對R7去除硝氮和氨氮的影響較大;丙酸鈉能夠提高菌株去除無機三態(tài)氮的能力,去除率均在87%以上;含丙酸鈉或乙酸鈉的環(huán)境中,利于菌株對氨氮和亞硝氮的去除,去除率分別在85%及98%以上;含蔗糖、丙酸鈉或甘露醇的環(huán)境中,利于菌株對硝氮的去除,去除率均在80%以上;酵母粉(≤1.43mmol/L)的環(huán)境中,菌株對氨氮、硝氮和亞硝氮的最大去除率分別為97.61%、99.50%及99.08%;蛋白胨(≤3.21 mmol/L)的環(huán)境中,菌株對氨氮、硝氮和亞硝氮的最大去除率分別為90.57%、94.36%及99.54%;尿素(≤0.71 mmol/L)環(huán)境中,菌株對氨氮、硝氮和亞硝氮的最大去除率分別為98.26%、50.68%及99.69%;不同鹽濃度(≤2 g/L)環(huán)境中,氨氮、硝氮及亞硝氮的最大除率分別為55.03%、96.62%及83.53%以上。R7對實際養(yǎng)殖水體中三態(tài)氮的去除效果結果表明,微好氧條件下該菌株能夠有效地去除復雜的自然養(yǎng)殖水體中的三態(tài)氮。綜上所述,經過條件優(yōu)化,R7菌體生物量明顯增加,且積累抗氧化性強的含酮基的類胡蘿卜素,為該菌株在養(yǎng)殖水體中的實際應用奠定了基礎。R7菌株對氨氮、硝氮及亞硝氮均具有較強的去除能力,對高濃度無機三態(tài)氮共存水體中的三態(tài)氮也具有較高去除及耐受能力。在含有小分子有機糖類、有機酸或醇、有機氮和不同鹽度等復雜水體環(huán)境中,該菌株適應性強,能夠去除水體中的無機三態(tài)氮。R7菌株的營養(yǎng)功能及去除無機三態(tài)氮的高效性具有潛在應用價值,為進一步開發(fā)環(huán)境適應能力強、針對性強的新型高效APB微生物生態(tài)制劑提供了理論參考。
[Abstract]:At present, the pollution problem of aquaculture water body is becoming more and more serious, and nitrogen pollution is the primary problem affecting the development of marine aquaculture. Therefore, eliminating nitrogen pollution in aquaculture water and restoring microecological balance are the key to solve the problem of aquaculture water pollution. Anoxygenic phototrophic bacteria (APB) has been removed from the water of water. Three state nitrogen, restoring the microecological balance of water body and improving the immunity of cultured animals, showed an important role. However, the actual water environment of the APB treatment was complex. It was of great practical value to explore the nitrogen removal effect of APB in the complex water environment. At present, the ability of APB to remove nitrite and the benefit of three nitrogen in the aquaculture water. The application and transformation of the research reports that less.APB will be affected by a variety of organic nitrogen sources, carbon sources and salinity in the water body. Aiming at the above problems, this paper uses the nitrogen fixing red bacteria (Rhodobacter azotoformans) R7 as the material to optimize the biomass by optimizing the conditions and lay the foundation for its application. The effects of the strain on the removal of three inorganic nitrogen, the utilization and conversion of inorganic three state nitrogen and the effect of inorganic three nitrogen on the growth and synthesis of photosynthetic pigments were investigated. In order to improve the practical application effect of the strain, the complex water environmental factors were also explored, including inorganic and organic nitrogen compounds, organic carbon compounds and the organic nitrogen compounds. The main results were as follows: the optimum conditions for improving the biomass and Car production by single factor rotation test (One-factor-at-a-time, OFAT) were 80%, 3 g/L yeast, 5 g/L peptone, 2.5 g/L glucose. Under the optimum conditions, the biomass of the bacteria was 3.34 g/L The increase to 17.59 g/L, increased by 4.27 times, the content of carotenoids increased from 2.43 mg/L to 30.31 mg/L, increased 11.47 times, and the main accumulation of globose, hydroxyl and spherical carotene.Car photostability showed that the stability of ketone based spherical ketones was higher, 0 lux, 500 lux and 3000 lux illumination strips. The removal and transformation of 568.53 h, 405.56 h and 112.64 h.R7 strains to inorganic three nitrogen show that R7 strain can grow well with ammonia nitrogen, nitrate nitrogen and nitrite nitrogen, and have strong tolerance and removal ability to nitrate nitrogen and nitrite nitrogen. The maximum removal of ammonia nitrogen, nitrate nitrogen and nitrite nitrogen by R7 strain in 8 days The maximum tolerable concentrations of 6.10 mmol/L, 21.36mmol/L and 16.86 mmol/L were 43.47 mmol/L, 52.17 mmol/L and 34.78 mmol/L, respectively. In the system of 4.54 mmol/L ammonia nitrogen, 34.35 mmol/L nitrate nitrogen and 34.64 mmol/L nitrite nitrogen, the removal rate of ammonia nitrogen, nitrate nitrogen and nitrite nitrogen were 54.38%, 31.48% and 99.95%, respectively, and inorganic three nitrogen nitrogen, respectively. The results of utilization and transformation showed that in the process of cultivation of ammonia nitrogen, nitrate nitrogen and nitrite as the sole nitrogen source, two other kinds of inorganic nitrogen were accumulated temporarily and gradually removed with the prolongation of the incubation time. The results showed that the R7 strain had a variety of nitrogen metabolism, denitrification and anaerobic nitrification. The effect of ammonia nitrogen, nitrite and nitrite on the growth and photosynthetic pigment synthesis of strains showed that ammonia nitrogen (less than 43.48 mmol/L) or nitrate nitrogen (less than 52.17 mmol/L) had no obvious effect on the biomass and Car content in the water, and the low concentration of ammonia nitrogen (less than 8.70 mmol/L) or nitrate nitrogen (less than 8.66 mmol/L) grew to the strain growth. The results showed that the effect of ammonia nitrogen and nitrate nitrogen on the pigment content of the strain was not obvious. The higher the concentration of nitrite nitrogen in the water body, the more obvious inhibition of the synthesis of R7 bacterial pigment. The possible small molecular carbon (saccharide, organic acid or alcohol), organic nitrogen (yeast extract, peptone or urea) and salinity affected bacteria in the environment The removal of inorganic three state nitrogen shows that the different carbon sources in the environment have great influence on the removal of nitrogen and ammonia nitrogen by R7. Sodium propionate can improve the ability of removing three nitrogen of inorganic nitrogen by the strain, and the removal rate is above 87%. In the environment containing sodium propionate or sodium acetate, the removal rate of ammonia and nitrite is 85% and 98, respectively. In the environment containing sucrose, sodium propionate or mannitol, the removal rate of nitrite was more than 80%, and the maximum removal rate of ammonia nitrogen, nitrite and nitrite was 97.61%, 99.50% and 99.08% in the yeast (less than 1.43mmol/L) environment, and the strain to ammonia nitrogen, nitrite and nitrite in the environment of egg white peptone (less than 3.21 mmol/L) The maximum removal rates of nitrogen were 90.57%, 94.36% and 99.54%, respectively, and the maximum removal rates of ammonia nitrogen, nitrite and nitrite were 98.26%, 50.68% and 99.69% in urea (less than 0.71 mmol/L) environment, and the maximum removal rate of ammonia nitrogen, nitrite and nitrite in different salt concentrations (less than 2 g/L) was 55.03%, 96.62% and 83.53%.R7 were used for actual cultivation. The results of the removal of three state nitrogen in water show that the strain can effectively remove three states of nitrogen in the complex natural aquaculture water under the condition of microaerobic. In summary, the biomass of R7 bacteria increases obviously and accumulates the ketone like Hu Luo, which has strong antioxidant activity, and is the practical application of the strain in the aquaculture water. The base.R7 strain has strong removal ability to ammonia nitrogen, nitrate nitrogen and nitrite nitrogen. It also has high removal and tolerance to three state nitrogen in high concentration inorganic three state nitrogen. It has strong adaptability in the complex water environment containing small molecular organic sugar, organic acid or alcohol, organic nitrogen and different salinity. It is of potential application value to remove the nutritional function of the inorganic three state nitrogen.R7 strain in the water and the efficiency of removing the inorganic three nitrogen nitrogen. It provides a theoretical reference for the further development of the environmental adaptation ability of the new high efficiency APB microorganism ecologic agent.
【學位授予單位】：華僑大學
【學位級別】：碩士
【學位授予年份】：2015
【分類號】：X52;X714
本文編號：2144745
[Abstract]:At present, the pollution problem of aquaculture water body is becoming more and more serious, and nitrogen pollution is the primary problem affecting the development of marine aquaculture. Therefore, eliminating nitrogen pollution in aquaculture water and restoring microecological balance are the key to solve the problem of aquaculture water pollution. Anoxygenic phototrophic bacteria (APB) has been removed from the water of water. Three state nitrogen, restoring the microecological balance of water body and improving the immunity of cultured animals, showed an important role. However, the actual water environment of the APB treatment was complex. It was of great practical value to explore the nitrogen removal effect of APB in the complex water environment. At present, the ability of APB to remove nitrite and the benefit of three nitrogen in the aquaculture water. The application and transformation of the research reports that less.APB will be affected by a variety of organic nitrogen sources, carbon sources and salinity in the water body. Aiming at the above problems, this paper uses the nitrogen fixing red bacteria (Rhodobacter azotoformans) R7 as the material to optimize the biomass by optimizing the conditions and lay the foundation for its application. The effects of the strain on the removal of three inorganic nitrogen, the utilization and conversion of inorganic three state nitrogen and the effect of inorganic three nitrogen on the growth and synthesis of photosynthetic pigments were investigated. In order to improve the practical application effect of the strain, the complex water environmental factors were also explored, including inorganic and organic nitrogen compounds, organic carbon compounds and the organic nitrogen compounds. The main results were as follows: the optimum conditions for improving the biomass and Car production by single factor rotation test (One-factor-at-a-time, OFAT) were 80%, 3 g/L yeast, 5 g/L peptone, 2.5 g/L glucose. Under the optimum conditions, the biomass of the bacteria was 3.34 g/L The increase to 17.59 g/L, increased by 4.27 times, the content of carotenoids increased from 2.43 mg/L to 30.31 mg/L, increased 11.47 times, and the main accumulation of globose, hydroxyl and spherical carotene.Car photostability showed that the stability of ketone based spherical ketones was higher, 0 lux, 500 lux and 3000 lux illumination strips. The removal and transformation of 568.53 h, 405.56 h and 112.64 h.R7 strains to inorganic three nitrogen show that R7 strain can grow well with ammonia nitrogen, nitrate nitrogen and nitrite nitrogen, and have strong tolerance and removal ability to nitrate nitrogen and nitrite nitrogen. The maximum removal of ammonia nitrogen, nitrate nitrogen and nitrite nitrogen by R7 strain in 8 days The maximum tolerable concentrations of 6.10 mmol/L, 21.36mmol/L and 16.86 mmol/L were 43.47 mmol/L, 52.17 mmol/L and 34.78 mmol/L, respectively. In the system of 4.54 mmol/L ammonia nitrogen, 34.35 mmol/L nitrate nitrogen and 34.64 mmol/L nitrite nitrogen, the removal rate of ammonia nitrogen, nitrate nitrogen and nitrite nitrogen were 54.38%, 31.48% and 99.95%, respectively, and inorganic three nitrogen nitrogen, respectively. The results of utilization and transformation showed that in the process of cultivation of ammonia nitrogen, nitrate nitrogen and nitrite as the sole nitrogen source, two other kinds of inorganic nitrogen were accumulated temporarily and gradually removed with the prolongation of the incubation time. The results showed that the R7 strain had a variety of nitrogen metabolism, denitrification and anaerobic nitrification. The effect of ammonia nitrogen, nitrite and nitrite on the growth and photosynthetic pigment synthesis of strains showed that ammonia nitrogen (less than 43.48 mmol/L) or nitrate nitrogen (less than 52.17 mmol/L) had no obvious effect on the biomass and Car content in the water, and the low concentration of ammonia nitrogen (less than 8.70 mmol/L) or nitrate nitrogen (less than 8.66 mmol/L) grew to the strain growth. The results showed that the effect of ammonia nitrogen and nitrate nitrogen on the pigment content of the strain was not obvious. The higher the concentration of nitrite nitrogen in the water body, the more obvious inhibition of the synthesis of R7 bacterial pigment. The possible small molecular carbon (saccharide, organic acid or alcohol), organic nitrogen (yeast extract, peptone or urea) and salinity affected bacteria in the environment The removal of inorganic three state nitrogen shows that the different carbon sources in the environment have great influence on the removal of nitrogen and ammonia nitrogen by R7. Sodium propionate can improve the ability of removing three nitrogen of inorganic nitrogen by the strain, and the removal rate is above 87%. In the environment containing sodium propionate or sodium acetate, the removal rate of ammonia and nitrite is 85% and 98, respectively. In the environment containing sucrose, sodium propionate or mannitol, the removal rate of nitrite was more than 80%, and the maximum removal rate of ammonia nitrogen, nitrite and nitrite was 97.61%, 99.50% and 99.08% in the yeast (less than 1.43mmol/L) environment, and the strain to ammonia nitrogen, nitrite and nitrite in the environment of egg white peptone (less than 3.21 mmol/L) The maximum removal rates of nitrogen were 90.57%, 94.36% and 99.54%, respectively, and the maximum removal rates of ammonia nitrogen, nitrite and nitrite were 98.26%, 50.68% and 99.69% in urea (less than 0.71 mmol/L) environment, and the maximum removal rate of ammonia nitrogen, nitrite and nitrite in different salt concentrations (less than 2 g/L) was 55.03%, 96.62% and 83.53%.R7 were used for actual cultivation. The results of the removal of three state nitrogen in water show that the strain can effectively remove three states of nitrogen in the complex natural aquaculture water under the condition of microaerobic. In summary, the biomass of R7 bacteria increases obviously and accumulates the ketone like Hu Luo, which has strong antioxidant activity, and is the practical application of the strain in the aquaculture water. The base.R7 strain has strong removal ability to ammonia nitrogen, nitrate nitrogen and nitrite nitrogen. It also has high removal and tolerance to three state nitrogen in high concentration inorganic three state nitrogen. It has strong adaptability in the complex water environment containing small molecular organic sugar, organic acid or alcohol, organic nitrogen and different salinity. It is of potential application value to remove the nutritional function of the inorganic three state nitrogen.R7 strain in the water and the efficiency of removing the inorganic three nitrogen nitrogen. It provides a theoretical reference for the further development of the environmental adaptation ability of the new high efficiency APB microorganism ecologic agent.
【學位授予單位】：華僑大學
【學位級別】：碩士
【學位授予年份】：2015
【分類號】：X52;X714
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