用于高盐度再生水富营养化控制的蓖齿眼子菜克藻效应与机理研究
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摘要
本文针对天津开发区再生水景观河道夏秋季节的富营养化问题,选用该河道中的优势沉水植物——蓖齿眼子菜为植物材料,以引起该河道富营养化的两种优势藻种——蛋白核小球藻、铜绿微囊藻为研究对象,主要研究了蓖齿眼子菜对高盐度再生水景观河道的富营养化藻类的克藻效应及其机理。主要成果结论如下:实验室光照培养箱中,在蓖齿眼子菜与两种藻的共培养、培植水培养、水浸提液培养的条件下,蓖齿眼子菜对两种藻的生长均产生了明显的抑制作用。其中共培养条件下的效果较好,藻液吸光值OD650的相对抑制率在实验3天后可以达到90%;藻细胞个数降至1.0×105个/mL以下。蓖齿眼子菜所分泌的克藻化学物质是产生克藻效应的主要原因。
通过响应曲面法Box-Behnken实验和模型分析,得出蓖齿眼子菜对小球藻的藻液吸光值OD650、藻细胞个数的相对抑制率在温度、光照强度、全盐量分别位于(43℃,2649lx,3787mg/L)、(42℃,2683lx,3706mg/L)时达到最佳点。铜绿微囊藻的相应指标位于(23℃,2700lx,4415mg/L)、(24℃,2891lx,4407mg/L)时抑制率达到最大值。模型经过方差分析具有高度显著性,相关系数较高,模型预测值与真实值之间拟合度较好。
对气-质联机鉴定出的克藻物质进行单一克藻效应实验,琥珀酸对小球藻抑制效果最好,棕榈酸对铜绿微囊藻效果最好;组合效应实验中,棕榈酸+琥珀酸对于铜绿微囊藻的组合协同作用最大,两种组合对小球藻出现拮抗作用,其余为协同作用。
在克藻化学物质的作用下,两种藻叶绿素a和细胞可溶性蛋白的生物合成受明显抑制,其中小球藻有低浓度促进、高浓度抑制的规律。小球藻SOD酶活性有抗逆性反应,表现为Fe-SOD前三天有明显升高,但是仍造成了不可逆伤害。铜绿微囊藻总SOD酶活性随克藻物质浓度加大而逐步降低,但是Mn-SOD酶表现出了抗逆性反应。总的来说,两种藻的正常生理功能受到了克藻物质的破坏,藻体最终凝聚沉淀,直至死亡。
连续流小试反应器中,蓖齿眼子菜对真实再生水景观河道中藻类的抑制作用明显,所有处理组叶绿素a的含量均低于对照。水力停留时间越长,再生水景观水体富营养化的可能性越大。
The allelopathic effects of submerged macrophytes against algae and its mechanism were investigated in this study based on the Hi-tech Research and Development Program of China——Water Environmental Quality Improvement Technology and Comprehensive Demonstration in Binhai District of Tianjin. Salt-proof submerged macrophytes——Potamogeton pectinatus,which is one of the advantageous species in the TEDA reclaimed water landscape water body, was the plant material in this paper. Two kinds of eutrophicated alga——Chlorella vulgaris and Microcystis aeruginosa, were selected as the objects of the allelopathic effects.
The results showed that Potamogeton pectinatus exerted obvirously inhibitory effect on the two alga growth under coexistence culture, solution culture and aqueous leachate culture conditions. The relatively inhibitory rate of OD650 was 90% and algal cell number was less than 1.0×105cells/mL under coexistence culture. Not the nutrients competition but allelopathic chemicals that secreted by Potamogeton pectinatus was the primary cause of algal growth inhibition.
Response surface methodology (RSM) based on three-level three-factor Box-Behnken design of experiment was used to optimize the environmental factors of allelopathy. The critical factors selected for this investigation were temperature, light intensity and total dissolved solids. By analyzing the response surface plots, the optimum ranges of the relatively inhibitory rate of OD650 and algal cell number lied in (43℃,2649lx,3787mg/L), (42℃,2683lx,3706mg/L) respectively for Chlorella vulgaris. The optimal conditions to gain were (23℃,2700lx,4415mg/L), (24℃,2891lx,4407mg/L) respectively for Microcystis aeruginosa. The results of RSM indicated that experimental values were in good agreement with the predicted ones and the models were satisfactory and accurate.
According to the methods of the Organization for Economic Cooperation and Development (OECD) for algal growth inhibition test, in the single-effect tests of the three allelopathic chemicals identified by gas chromatography-mass (GC-Mass) spectrometry, succinic acid had the best effect on Chlorella vulgaris and palmitic acid had the best effect on Microcystis aeruginosa. In the combined-effect tests, the group of palmitic acid + succinic acid had the best additive effect on Microcystis aeruginosa and two groups had opposite effect on Chlorella vulgaris.
The chlorophlla-a content and protein in Chlorella vulgaris and Microcystis aeruginosa were obviously inhibited under the stress of allelopathic chemicals. Fe-SOD enzyme activity of Chlorella vulgaris increased firstly, but decreased as treatment time went on. The total-SOD enzyme activity of Microcystis aeruginosa decreased with allelopathic compound concentrations. But the Mn-SOD enzyme activity of Microcystis aeruginosa exerted resistance response. In short, the physiological founction of the two algae were damaged by allelopathic compounds and finally algae cell flocculates and subsides to death.
In the continuous flow reator test, salt-proof Potamogeton pectinatus inhibited the growth of alga of real reclaimed scenic water with high salinity. The chlorophlla-a contents of all treatment tests were lower compared with the reactor without plants. The possibility of eutrophication in reclaimed scenic water was in direct proportion to the hydraulic residence time (HRT).
The three main innovation points of this study as follows:①the allelopathic effects of salt-proof Potamogeton pectinatus on Chlorella vulgaris and Microcystis aeruginosa, which are the main cause of eutrophication of TEDA reclaimed scenic water with high salinity;②the effects of three environmental factors on allelopathic effects by response surface methodology at the same time;③the mechanism of the allelopathy against algae.
通过响应曲面法Box-Behnken实验和模型分析,得出蓖齿眼子菜对小球藻的藻液吸光值OD650、藻细胞个数的相对抑制率在温度、光照强度、全盐量分别位于(43℃,2649lx,3787mg/L)、(42℃,2683lx,3706mg/L)时达到最佳点。铜绿微囊藻的相应指标位于(23℃,2700lx,4415mg/L)、(24℃,2891lx,4407mg/L)时抑制率达到最大值。模型经过方差分析具有高度显著性,相关系数较高,模型预测值与真实值之间拟合度较好。
对气-质联机鉴定出的克藻物质进行单一克藻效应实验,琥珀酸对小球藻抑制效果最好,棕榈酸对铜绿微囊藻效果最好;组合效应实验中,棕榈酸+琥珀酸对于铜绿微囊藻的组合协同作用最大,两种组合对小球藻出现拮抗作用,其余为协同作用。
在克藻化学物质的作用下,两种藻叶绿素a和细胞可溶性蛋白的生物合成受明显抑制,其中小球藻有低浓度促进、高浓度抑制的规律。小球藻SOD酶活性有抗逆性反应,表现为Fe-SOD前三天有明显升高,但是仍造成了不可逆伤害。铜绿微囊藻总SOD酶活性随克藻物质浓度加大而逐步降低,但是Mn-SOD酶表现出了抗逆性反应。总的来说,两种藻的正常生理功能受到了克藻物质的破坏,藻体最终凝聚沉淀,直至死亡。
连续流小试反应器中,蓖齿眼子菜对真实再生水景观河道中藻类的抑制作用明显,所有处理组叶绿素a的含量均低于对照。水力停留时间越长,再生水景观水体富营养化的可能性越大。
The allelopathic effects of submerged macrophytes against algae and its mechanism were investigated in this study based on the Hi-tech Research and Development Program of China——Water Environmental Quality Improvement Technology and Comprehensive Demonstration in Binhai District of Tianjin. Salt-proof submerged macrophytes——Potamogeton pectinatus,which is one of the advantageous species in the TEDA reclaimed water landscape water body, was the plant material in this paper. Two kinds of eutrophicated alga——Chlorella vulgaris and Microcystis aeruginosa, were selected as the objects of the allelopathic effects.
The results showed that Potamogeton pectinatus exerted obvirously inhibitory effect on the two alga growth under coexistence culture, solution culture and aqueous leachate culture conditions. The relatively inhibitory rate of OD650 was 90% and algal cell number was less than 1.0×105cells/mL under coexistence culture. Not the nutrients competition but allelopathic chemicals that secreted by Potamogeton pectinatus was the primary cause of algal growth inhibition.
Response surface methodology (RSM) based on three-level three-factor Box-Behnken design of experiment was used to optimize the environmental factors of allelopathy. The critical factors selected for this investigation were temperature, light intensity and total dissolved solids. By analyzing the response surface plots, the optimum ranges of the relatively inhibitory rate of OD650 and algal cell number lied in (43℃,2649lx,3787mg/L), (42℃,2683lx,3706mg/L) respectively for Chlorella vulgaris. The optimal conditions to gain were (23℃,2700lx,4415mg/L), (24℃,2891lx,4407mg/L) respectively for Microcystis aeruginosa. The results of RSM indicated that experimental values were in good agreement with the predicted ones and the models were satisfactory and accurate.
According to the methods of the Organization for Economic Cooperation and Development (OECD) for algal growth inhibition test, in the single-effect tests of the three allelopathic chemicals identified by gas chromatography-mass (GC-Mass) spectrometry, succinic acid had the best effect on Chlorella vulgaris and palmitic acid had the best effect on Microcystis aeruginosa. In the combined-effect tests, the group of palmitic acid + succinic acid had the best additive effect on Microcystis aeruginosa and two groups had opposite effect on Chlorella vulgaris.
The chlorophlla-a content and protein in Chlorella vulgaris and Microcystis aeruginosa were obviously inhibited under the stress of allelopathic chemicals. Fe-SOD enzyme activity of Chlorella vulgaris increased firstly, but decreased as treatment time went on. The total-SOD enzyme activity of Microcystis aeruginosa decreased with allelopathic compound concentrations. But the Mn-SOD enzyme activity of Microcystis aeruginosa exerted resistance response. In short, the physiological founction of the two algae were damaged by allelopathic compounds and finally algae cell flocculates and subsides to death.
In the continuous flow reator test, salt-proof Potamogeton pectinatus inhibited the growth of alga of real reclaimed scenic water with high salinity. The chlorophlla-a contents of all treatment tests were lower compared with the reactor without plants. The possibility of eutrophication in reclaimed scenic water was in direct proportion to the hydraulic residence time (HRT).
The three main innovation points of this study as follows:①the allelopathic effects of salt-proof Potamogeton pectinatus on Chlorella vulgaris and Microcystis aeruginosa, which are the main cause of eutrophication of TEDA reclaimed scenic water with high salinity;②the effects of three environmental factors on allelopathic effects by response surface methodology at the same time;③the mechanism of the allelopathy against algae.
引文
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