CN114062074A - Method for discriminating toxicity of organic composite polluted sediment - Google Patents

Method for discriminating toxicity of organic composite polluted sediment Download PDF

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CN114062074A
CN114062074A CN202111243975.3A CN202111243975A CN114062074A CN 114062074 A CN114062074 A CN 114062074A CN 202111243975 A CN202111243975 A CN 202111243975A CN 114062074 A CN114062074 A CN 114062074A
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toxicity
organic composite
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可欣
冯书娜
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Shenyang Aerospace University
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    • GPHYSICS
    • G01MEASURING; TESTING
    • G01NINVESTIGATING OR ANALYSING MATERIALS BY DETERMINING THEIR CHEMICAL OR PHYSICAL PROPERTIES
    • G01N1/00Sampling; Preparing specimens for investigation
    • G01N1/28Preparing specimens for investigation including physical details of (bio-)chemical methods covered elsewhere, e.g. G01N33/50, C12Q
    • GPHYSICS
    • G01MEASURING; TESTING
    • G01NINVESTIGATING OR ANALYSING MATERIALS BY DETERMINING THEIR CHEMICAL OR PHYSICAL PROPERTIES
    • G01N33/00Investigating or analysing materials by specific methods not covered by groups G01N1/00 - G01N31/00
    • G01N33/48Biological material, e.g. blood, urine; Haemocytometers
    • G01N33/50Chemical analysis of biological material, e.g. blood, urine; Testing involving biospecific ligand binding methods; Immunological testing
    • G01N33/5005Chemical analysis of biological material, e.g. blood, urine; Testing involving biospecific ligand binding methods; Immunological testing involving human or animal cells
    • G01N33/5008Chemical analysis of biological material, e.g. blood, urine; Testing involving biospecific ligand binding methods; Immunological testing involving human or animal cells for testing or evaluating the effect of chemical or biological compounds, e.g. drugs, cosmetics
    • G01N33/5014Chemical analysis of biological material, e.g. blood, urine; Testing involving biospecific ligand binding methods; Immunological testing involving human or animal cells for testing or evaluating the effect of chemical or biological compounds, e.g. drugs, cosmetics for testing toxicity

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Abstract

The invention discloses a method for screening toxicity of organic composite polluted sediments, which comprises the following steps: 1) domesticating chironomid larvae; 2) pretreating organic composite pollution sediments; 3) extracting sediment extracting solution; 4) and (3) testing: separating interference components of the sediment extracting solution to obtain components for biological toxicity test, finding out toxic components by combining with the midge larva in-vitro toxicity test, then carrying out scanning analysis and spectrum library comparison on the toxic components, and tracking specific toxic pollutant. The method for screening the toxicity of the organic composite polluted sediments adopts common biological tests, can economically and visually reflect the toxicity of the sediments, meets the test requirements of most laboratories, can be popularized and used, can narrow the detection range, defines specific toxic organic pollutants, and has low cost in the whole process.

Description

Method for discriminating toxicity of organic composite polluted sediment
Technical Field
The invention relates to a method for screening toxicity of organic composite polluted sediments.
Background
Since the 20 th century, with the rapid development of industries such as smelting, chemical engineering, pesticides and medicines, a large amount of industrial, agricultural and domestic sewage is discharged into water, so that the water is seriously polluted. After entering the water body, the pollutants are gradually enriched in the water body sediments, and the pollutants in the sediments are released into the water body again through the chemical and physical process to become a secondary pollution source, so that the water quality and the ecological system are influenced. Thus, sediment is a "source" and "sink" of water pollution. Furthermore, even if the source of pollution ceases to contaminate the water, the contaminated sediment may cause the water quality to deteriorate. High concentrations of contaminants accumulated in the sediment can be harmful to benthic organisms. For every body of water, contaminated sediments pose a potential risk to fish and humans, as well as to wild animals eating fish. Contaminated sediments also cause ecological and economic losses to aquatic resources, such as deterioration of habitats and expensive remediation costs.
In recent years, a large amount of pollutants including three types of organic pollutants, heavy metals and ammonia nitrogen are detected in sediments. Organic pollution is more serious. The toxicity of organic pollutants in water sediments is often the result of the combined action of multiple pollutants and various complex intermediates thereof, and the complexity and complexity of the pollution make the traditional characteristic pollutant-based toxicity evaluation method inapplicable. The extensive extension of the chemical analysis object, even the full scan of known organic pollutants, may partially compensate this drawback, however, the large amount of time, financial consumption and the lack of toxicity data limit the further application of the method. Therefore, in order to accurately evaluate and control the pollution of the water body sediments, on one hand, the toxicity of the water body sediments needs to be known, and on the other hand, the types of direct or indirect pollutants which generate toxic effects or specific pollutants need to be known.
When the toxicity of organic pollutants in the sediment is screened, some unknown pollutants cannot be tracked due to the coexistence of multiple organic pollutants. Therefore, it is an urgent need to solve the problem of determining the current biological toxicity of organic pollutants in sediments, tracing toxic pollutants, and finally identifying toxic pollutants.
Disclosure of Invention
In view of the above, the invention aims to provide a method for screening the toxicity of organic composite polluted sediments.
The technical scheme provided by the invention is as follows: a method for screening toxicity of organic composite polluted sediments comprises the following steps:
1) domesticating chironomid larvae
Collecting chironomus larvae as a test organism, and then domesticating the chironomus larvae for not less than 7 days;
2) pretreatment of organic composite contaminated deposits
Storing the organic composite pollution sediments to be treated in a refrigerator, and uniformly stirring;
3) extraction of sediment extract
Taking 200-500 g of wet sediment, 200-500 ml of medium-hardness water and 10-20g of XAD resin, stirring for 24 hours at 23 ℃ in a dark place, then recovering the resin, washing with deionized water, and then respectively extracting with mixed liquid of acetone, n-hexane and acetone to obtain extract liquid, namely: a sediment extract;
4) testing
Separating interference components of the sediment extracting solution to obtain components for biological toxicity test, finding out toxic components by combining with the midge larva in-vitro toxicity test, then carrying out scanning analysis and spectrum library comparison on the toxic components, and tracking specific toxic pollutant.
Preferably, in the step 1), feeding 1-2 times every day and changing water every 3 days in the process of domesticating the chironomid larvae.
Further preferably, in step 4), the interfering components of the sediment extract are separated by normal phase chromatography and reverse phase chromatography.
Further preferably, in the step 4), the method for testing the toxicity of the chironomus larvas in vitro is as follows:
first, 12-well plates were taken for testing, and then the components for biotoxicity testing were dissolved in dimethyl sulfoxide and poured into the wells of 12-well plates, respectively, and then 5 chironomid larvae, 4-5 ml of medium hardness water and a small amount of clean sand were added per well for testing.
Further preferably, the test time of the toxicity of the chironomus larvas in vitro is 72 hours, the test temperature is set to be 23-25 ℃, the illumination is 16-18 hours per day, the darkness is 8-6 hours, and the whole process is not changed with water and is not fed.
Further preferably, a glass lining adapted to the hole of the 12-hole plate is arranged in the hole of the 12-hole plate to prevent organic compounds from being adsorbed on the hole plate.
The method for screening the toxicity of the organic composite polluted sediment provided by the invention adopts common biological tests, can economically and visually reflect the toxicity of the sediment, meets the test requirements of most laboratories, can be popularized and used, can narrow the detection range, defines specific toxic organic pollutants, and has low cost in the whole process.
Detailed Description
The invention will be further explained with reference to specific embodiments, without limiting the invention.
In order to solve the problem that the conventional method for screening the toxicity of the organic composite polluted sediment is complex, the invention provides a method for screening the toxicity of the organic composite polluted sediment, which comprises the following steps:
1) domesticating chironomid larvae
Collecting chironomus larvae as a test organism, and then domesticating the chironomus larvae for not less than 7 days;
2) pretreatment of organic composite contaminated deposits
Storing the organic composite pollution sediments to be treated in a refrigerator, and uniformly stirring;
3) extraction of sediment extract
Taking 200 g-500 g of wet sediment, 200-500 ml of medium-hardness water and 10-20g of XAD resin, stirring for 24 h at 23 ℃ in a dark place, then recovering the resin, washing with deionized water, and then respectively extracting with mixed liquid of acetone, n-hexane and acetone to obtain extract liquid, namely: a sediment extract;
4) testing
Separating interference components of the sediment extracting solution to obtain components for biological toxicity test, finding out toxic components by combining with the midge larva in-vitro toxicity test, then carrying out scanning analysis and spectrum library comparison on the toxic components, and tracking specific toxic pollutant.
As an improvement of the technical scheme, in the step 1), feeding 1-2 times every day and changing water every 3 days in the process of domesticating the chironomid larvae.
As an improvement of the technical scheme, in the step 4), the interfering components of the sediment extracting solution are separated through normal phase chromatography and reverse phase chromatography.
As an improvement of the technical scheme, in the step 4), the method for testing the toxicity of the chironomus larvas in vitro comprises the following steps:
first, 12-well plates were taken for testing, and then the components for biotoxicity testing were dissolved in dimethyl sulfoxide and poured into the wells of 12-well plates, respectively, and then 5 chironomid larvae, 4-5 ml of medium hardness water and a small amount of clean sand were added per well for testing.
As an improvement of the technical scheme, the test time of the toxicity of the chironomus larvas in vitro is 72 hours, the test temperature is set to be 23-25 ℃, the light is 16-18 hours per day, the darkness is 8-6 hours, and the whole process does not need to change water or feed.
As an improvement of the technical scheme, a glass lining adapted to the hole of the 12-hole plate is arranged in the hole of the 12-hole plate so as to prevent organic compounds from being adsorbed on the hole plate.
Example 1
The method for screening the toxicity of the organic composite polluted sediments comprises the following steps:
1) domesticating chironomid larvae
Taking Chironomus larvae collected in Liaohe river as test organisms, then domesticating the test organisms for 7 days, feeding the test organisms for 1 time every day, changing water every 3 days, wherein the water temperature is 25 ℃, and the pH value is 8 so as to ensure the optimal domesticating environment;
2) pretreatment of organic composite contaminated deposits
Storing the organic composite pollution sediments to be treated, which are taken from the area A, in a refrigerator, and uniformly stirring;
3) extraction of sediment extract
Taking 200g of wet sediment, 200ml of medium-hardness water and 10g of XAD resin, putting the wet sediment, 200ml of medium-hardness water and 10g of XAD resin into a 500ml conical flask, stirring the mixture for 24 hours at the temperature of 23 ℃ in a dark place at the speed of 500 revolutions per minute, then recovering the resin, washing the resin with deionized water for 3 times, and then respectively extracting the resin once with acetone and extracting the resin for 2 times with a mixed solution of normal hexane and acetone (the volume ratio is 1: 1) to obtain extract liquid, namely: a sediment extract;
4) testing
Separating interference components of the sediment extracting solution through a normal phase chromatography and a reverse phase chromatography to obtain components for biological toxicity test, finding out toxic components by combining with the midge larva in-vitro toxicity test, then, carrying out scanning analysis and spectrum library comparison on the toxic components, and tracking specific toxic pollutant;
the method for testing the in vitro toxicity of the chironomus larvas comprises the following steps:
firstly, taking a 12-hole plate for testing, wherein a glass lining adapted to the 12-hole plate is arranged in the 12-hole plate, then dissolving components for biological toxicity testing in dimethyl sulfoxide and pouring the components into the 12-hole plate, then adding 5 chironomus larvae, 4 ml of medium-hardness water and a small amount of clean sand into each hole to serve as attachments of organisms, testing, wherein the testing time is 72 hours, the testing temperature is set to be 23 ℃, the lighting is 16 hours every day, the darkness is 8 hours, and the whole process is not changed with water and is not fed.
5) Analysis of results
The sediment extracting solution is subjected to normal phase and reverse phase chromatographic purification combined with chironomus larvas in-vitro toxicity test, the toxicity effect of specific components is determined by calculating the death rate of chironomus larvas, then scanning analysis and spectrum library comparison are carried out on the toxicity components, the similarity of galaxolide is more than 80%, the fact that galaxolide has obvious toxicity effect on the chironomus larvas is proved, and LC50=7.5 ng/ml.
Example 2
The method for screening the toxicity of the organic composite polluted sediments comprises the following steps:
1) domesticating chironomid larvae
Chironomus larvae collected in Liaohe river are taken as test organisms, and then domesticated for 10 days, the test organisms are fed for 2 times every day, water is changed every 3 days, the water temperature is controlled at 23 ℃, the proper pH value is 7, and the dissolved oxygen is controlled to be more than 2 mg/L. Culturing with a light-to-dark ratio of 16: 8;
2) pretreatment of organic composite contaminated deposits
Storing the organic composite pollution sediments to be treated, which are taken from the area B, in a refrigerator, and uniformly stirring;
3) extraction of sediment extract
Taking 500g of wet sediment, 500ml of medium-hardness water and 20g of XAD resin, putting the wet sediment, 500ml of medium-hardness water and 20g of XAD resin into a 1000ml conical flask, stirring the mixture for 24 hours at the temperature of 23 ℃ in a dark place at the speed of 800 revolutions per minute, then recovering the resin, washing the resin with deionized water for 3 times, and then respectively extracting the resin once with acetone and extracting the resin for 3 times with a mixed solution of normal hexane and acetone (the volume ratio is 1: 1) to obtain extract liquid, namely: a sediment extract;
4) testing
Separating interference components of the sediment extracting solution through a normal phase chromatography and a reverse phase chromatography to obtain components for biological toxicity test, finding out toxic components by combining with the midge larva in-vitro toxicity test, then, carrying out scanning analysis and spectrum library comparison on the toxic components, and tracking specific toxic pollutant;
the method for testing the in vitro toxicity of the chironomus larvas comprises the following steps:
firstly, taking a 12-hole plate for testing, wherein a glass lining adapted to the 12-hole plate is arranged in the 12-hole plate, then dissolving components for biological toxicity testing in dimethyl sulfoxide and pouring the dimethyl sulfoxide into the 12-hole plate holes respectively, then adding 5 chironomus larvae, 5ml of medium-hardness water and a small amount of clean sand into each hole to serve as attachments of organisms, testing, wherein the testing time is 72 hours, the testing temperature is set to be 25 ℃, the illumination is 16 hours every day, the darkness is 8 hours, and the whole process is not changed with water or fed.
5) Analysis of results
The sediment extracting solution is subjected to normal phase and reverse phase chromatographic purification combined with chironomid larva in-vitro toxicity test, the toxicity effect of specific components is determined by calculating the death rate of chironomid larvae, then, scanning analysis and spectrum library comparison are carried out on toxic components, and the similarity of tonalid musk and galaxolide musk is more than 80%. Galaxolide and tonalid demonstrated significant toxic effects on chironomid larvae with LC50=9 ng/ml and 8.5 ng/ml.
Example 3
The method for screening the toxicity of the organic composite polluted sediments comprises the following steps:
1) domesticating chironomid larvae
Chironomus larvas collected in Liaohe river are used as test organisms and identified as local brook Chironomus, then the test organisms are domesticated for 7 days, the test organisms are fed for 1 time every day, water is changed every 3 days, the water temperature is controlled at 24 ℃, the proper pH value is 7, and the dissolved oxygen is controlled to be more than 2 mg/L. Culturing with a light-to-dark ratio of 18: 6;
2) pretreatment of organic composite contaminated deposits
Storing the organic composite pollution sediments to be treated from the region C in a refrigerator, and uniformly stirring;
3) extraction of sediment extract
Taking 300g of wet sediment, 300ml of medium-hardness water and 15g of XAD resin, putting the wet sediment, 300ml of medium-hardness water and 15g of XAD resin into a 1000ml conical flask, stirring the mixture for 24 hours at the temperature of 23 ℃ in a dark place at the speed of 700 revolutions per minute, then recovering the resin, washing the resin with deionized water for 3 times, and then respectively extracting the resin once with acetone and extracting the resin for 3 times with a mixed solution of normal hexane and acetone (the volume ratio is 1: 1) to obtain extract liquid, namely: a sediment extract;
4) testing
Separating interference components of the sediment extracting solution through a normal phase chromatography and a reverse phase chromatography to obtain components for biological toxicity test, finding out toxic components by combining with the midge larva in-vitro toxicity test, then, carrying out scanning analysis and spectrum library comparison on the toxic components, and tracking specific toxic pollutant;
the method for testing the in vitro toxicity of the chironomus larvas comprises the following steps:
firstly, taking a 12-hole plate for testing, wherein a glass lining adapted to the 12-hole plate is arranged in the 12-hole plate, then dissolving components for biological toxicity testing in dimethyl sulfoxide and pouring the dimethyl sulfoxide into the 12-hole plate holes respectively, then adding 5 chironomus larvae, 4 ml of medium-hardness water and a small amount of clean sand into each hole to serve as attachments of organisms, testing, wherein the testing time is 72 hours, the testing temperature is set to be 25 ℃, the light is set for 18 hours every day, the dark is set for 6 hours, and the whole process is not changed with water and fed.
5) Analysis of results
The sediment extracting solution is subjected to normal phase and reverse phase chromatographic purification combined with chironomus larvas in-vitro toxicity test, the toxicity effect of specific components is determined by calculating the death rate of chironomus larvas, then scanning analysis and spectrum library comparison are carried out on the toxicity components, the similarity of galaxolide is more than 80%, the fact that galaxolide has obvious toxicity effect on the chironomus larvas is proved, and LC50=8 ng/ml.

Claims (6)

1. A method for screening toxicity of organic composite polluted sediments is characterized by comprising the following steps:
1) domesticating chironomid larvae
Collecting chironomus larvae as a test organism, and then domesticating the chironomus larvae for not less than 7 days;
2) pretreatment of organic composite contaminated deposits
Storing the organic composite pollution sediments to be treated in a refrigerator, and uniformly stirring;
3) extraction of sediment extract
Taking 200-500 g of wet sediment, 200-500 ml of medium-hardness water and 10-20g of XAD resin, stirring for 24 hours at 23 ℃ in a dark place, then recovering the resin, washing with deionized water, and then respectively extracting with mixed liquid of acetone, n-hexane and acetone to obtain extract liquid, namely: a sediment extract;
4) testing
Separating interference components of the sediment extracting solution to obtain components for biological toxicity test, finding out toxic components by combining with the midge larva in-vitro toxicity test, then carrying out scanning analysis and spectrum library comparison on the toxic components, and tracking specific toxic pollutant.
2. The method for screening the toxicity of the organic composite polluted sediments according to claim 1, which is characterized by comprising the following steps: in the step 1), feeding 1-2 times every day and changing water every 3 days in the process of domesticating chironomus larvae.
3. The method for screening the toxicity of the organic composite polluted sediments according to claim 1, which is characterized by comprising the following steps: in step 4), the interfering components of the sediment extract are separated by normal phase chromatography and reverse phase chromatography.
4. The method for screening the toxicity of the organic composite polluted sediments according to claim 1, which is characterized by comprising the following steps: in the step 4), the test method for the in vitro toxicity of the chironomus larvas comprises the following steps:
first, 12-well plates were taken for testing, and then the components for biotoxicity testing were dissolved in dimethyl sulfoxide and poured into the wells of 12-well plates, respectively, and then 5 chironomid larvae, 4-5 ml of medium hardness water and a small amount of clean sand were added per well for testing.
5. The method for screening the toxicity of the organic composite polluted sediments according to claim 4, which is characterized by comprising the following steps: the test time of the toxicity of chironomus larvas in vitro is 72 hours, the test temperature is set to be 23-25 ℃, the illumination is 16-18 hours per day, the darkness is 8-6 hours, and the whole process does not change water or feed.
6. The method for screening the toxicity of the organic composite polluted sediments according to claim 4, which is characterized by comprising the following steps: and a glass lining adaptive to the hole of the 12-hole plate is arranged in the hole of the 12-hole plate so as to prevent organic compounds from being adsorbed on the hole plate.
CN202111243975.3A 2021-10-26 2021-10-26 Method for discriminating toxicity of organic composite polluted sediment Pending CN114062074A (en)

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Citations (4)

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CN103364481A (en) * 2013-07-12 2013-10-23 沈阳航空航天大学 Discriminating method for biological toxicity of sediment
CN103940961A (en) * 2014-04-13 2014-07-23 浙江大学 Method for evaluating effectiveness of organic pollution adsorbent in multiple pollution modes
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Patent Citations (4)

* Cited by examiner, † Cited by third party
Publication number Priority date Publication date Assignee Title
CN102279231A (en) * 2011-06-28 2011-12-14 中国科学院生态环境研究中心 Quick qualitative detection method for polychlorinated biphenyl pollutants
CN103364481A (en) * 2013-07-12 2013-10-23 沈阳航空航天大学 Discriminating method for biological toxicity of sediment
CN103940961A (en) * 2014-04-13 2014-07-23 浙江大学 Method for evaluating effectiveness of organic pollution adsorbent in multiple pollution modes
CN104237436A (en) * 2014-09-03 2014-12-24 上海大学 Qualitative screening method of semi-volatile halogenated organic pollutants in sediments

Non-Patent Citations (1)

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Title
易小意: "毒性鉴别评价和效应导向分析的联用:广州河涌沉积物中致毒物的筛查", 《中国博士学位论文全文数据库 工程科技I辑》, no. 1, 15 January 2018 (2018-01-15), pages 37 - 44 *

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