Detection of toxic substances in water by means of a mammalian cell culture technique

The cloning efficiency of a mammalian cell culture was used as a sensitive technique for the detection of toxicants in water. The sensitivity of the technique was tested by exposing HeLa, mouse lymphoma and Buffalo green monkey cell lines to known amounts of six different toxicants. The cell culture system tested proved to be twice as sensitive as an electronic fish bioassay system. Biofilter humus tank, and activated sludge effluents as well as settled sewage showed toxic activity when tested by the cell culture system.     

Effect of chlorination on endotoxin activities in secondary sewage effluent and typical Gram-negative bacteria

Wastewater reuse is a viable and attractive method to address water shortage problems. However, wastewater can have high endotoxin activity. Endotoxins are toxic inflammatory agents and are considered a risk factor for wastewater reuse. In this study, the effect of chlorination on endotoxin activity in secondary sewage effluent was evaluated by Limulus (Tachypleus tridentatus) Amebocyte Lysate assay. It was found that chlorination could not decrease endotoxin activity of secondary effluent effectively under the conditions employed in this study. Chlorination of a pure cultured Gram-negative bacterium (Escherichia coli), and a Gram-negative bacterium isolated from secondary sewage effluent, resulted in a significant increase in endotoxin activity, suggesting that the presence of Gram-negative bacteria contributed substantially to endotoxin activity, masking any potential reduction that may be attributable to chlorination. Furthermore, the activities of both free and cell-bound endotoxins in pure culture increased significantly during chlorination due to cell wall damage induced by chlorination.     

Indirect effects of contaminants in aquatic ecosystems

Contaminants such as petroleum hydrocarbons, heavy metals and pesticides can cause direct toxic effects when released into aquatic environments. Sensitive species may be impaired by sublethal effects or decimated by lethality, and this ecological alteration may initiate a trophic cascade or a release from competition that secondarily leads to responses in tolerant species. Contaminants may exert direct effects on keystone facilitator and foundation species, and contaminant-induced changes in nutrient and oxygen dynamics may alter ecosystem function. Thus, populations and communities in nature may be directly and/or indirectly affected by exposure to pollutants. While the direct effects of toxicants usually reduce organism abundance, indirect effects may lead to increased or decreased abundance. Here we review 150 papers that reference indirect toxicant effects in aquatic environments. Studies of accidental contaminant release, chronic contamination and experimental manipulations have identified indirect contaminant effects in pelagic and benthic communities caused by many types of pollutants. Contaminant-induced changes in behavior, competition and predation/grazing rate can alter species abundances or community composition, and enhance, mask or spuriously indicate direct contaminant effects. Trophic cascades were found in 60% of the manipulative studies and, most commonly, primary producers increased in abundance when grazers were selectively eliminated by contaminants. Competitive release may also be common, but is difficult to distinguish from trophic cascades because few experiments are designed to isolate the mechanism(s) causing indirect effects. Indirect contaminant effects may have profound implications in environments with strong trophic cascades such as the freshwater pelagic. In spite of their undesirable environmental influence, contaminants can be useful manipulative tools for the study of trophic and competitive interactions in natural communities.     

Bioanalytical tools for the evaluation of organic micropollutants during sewage treatment, water recycling and drinking water generation

A bioanalytical test battery was used for monitoring organic micropollutants across an indirect potable reuse scheme testing sites across the complete water cycle from sewage to drinking water to assess the efficacy of different treatment barriers. The indirect potable reuse scheme consists of seven treatment barriers: (1) source control, (2) wastewater treatment plant, (3) microfiltration, (4) reverse osmosis, (5) advanced oxidation, (6) natural environment in a reservoir and (7) drinking water treatment plant. Bioanalytical results provide complementary information to chemical analysis on the sum of micropollutants acting together in mixtures. Six endpoints targeting the groups of chemicals with modes of toxic action of particular relevance for human and environmental health were included in the evaluation: genotoxicity, estrogenicity (endocrine disruption), neurotoxicity, phytotoxicity, dioxin-like activity and non-specific cell toxicity. The toxicity of water samples was expressed as toxic equivalent concentrations (TEQ), a measure that translates the effect of the mixtures of unknown and potentially unidentified chemicals in a water sample to the effect that a known reference compound would cause. For each bioassay a different representative reference compound was selected. In this study, the TEQ concept was applied for the first time to the umuC test indicative of genotoxicity using 4-nitroquinoline as the reference compound for direct genotoxicity and benzo[a]pyrene for genotoxicity after metabolic activation.The TEQ were observed to decrease across the seven treatment barriers in all six selected bioassays. Each bioassay showed a differentiated picture representative for a different group of chemicals and their mixture effect. The TEQ of the samples across the seven barriers were in the same order of magnitude as seen during previous individual studies in wastewater and advanced water treatment plants and reservoirs. For the first time a benchmarking was performed that allows direct comparison of different treatment technologies and covers several orders of magnitude of TEQ from highly contaminated sewage to drinking water with TEQ close or below the limit of detection. Detection limits of the bioassays were decreased in comparison to earlier studies by optimizing sample preparation and test protocols, and were comparable to or lower than the quantification limits of the routine chemical analysis, which allowed monitoring of the presence and removal of micropollutants post Barrier 2 and in drinking water. The results obtained by bioanalytical tools were reproducible, robust and consistent with previous studies assessing the effectiveness of the wastewater and advanced water treatment plants. The results of this study indicate that bioanalytical results expressed as TEQ are useful to assess removal efficiency of micropollutants throughout all treatment steps of water recycling.     

Impact of chloroanilines on hydrogenotrophic methanogenesis in ethanol-enriched cultures

The impact of toxic organic chemicals on the kinetics of hydrogen conversion in ethanol-enriched cultures has been investigated. The study first involved an assessment of the kinetics of gaseous hydrogen transformation in a hydrogen-enriched culture. Tests to quantify the distribution of biomass in an ethanol-enriched culture indicated that hydrogenotropic methanogens made up 25.4% of the total biomass. Various amounts of chlorinated anilines were then added to ethanol-enriched test cultures to determine their impact on the hydrogen conversion reaction. Test results indicated that hydrogen conversion was mass-transfer limited from the gas phase but not from solution. While chlorinated anilines had a major adverse impact on acetogenic conversion of ethanol to hydrogen, the associated hydrogenotrophic methanogenesis reaction was affected to lesser extent by the toxicants. Tests with 3-chlorophenol showed similar inhibition of acetogenesis and acetoclastic methanogenesis but not hydrogenotrophic methanogenesis. The principal significance in this study is thought to lie in the use of specific hydrogenotrophic biomass for assessing the kinetics of hydrogen conversion and in the observation that the kinetics of hydrogenotrophic methanogenesis was not adversely affected by chlorinated anilines and chlorinated phenols. The principal application lies in the observation that gas-phase hydrogen may not be a good measure of the impact of toxic organic chemicals on anaerobic digestion processes.     

A novel algal toxicity testing technique for assessing the toxicity of both metallic and organic toxicants

This study presents a closed-system algal toxicity test technique that is capable of detecting the effects of both organic and metallic toxicants. Toxicity testing was conducted by transferring adequate amounts of algal suspension, dilution water (with culture growth medium), and toxicants into 300-mL BOD bottles. The BOD bottles were completely filled up with no head-space left. The initial cell density and the exposure time were 15,000 cells/mL and 48 h, respectively. The performance of the above test method was evaluated using three heavy metals and six organic toxicants based on three different test endpoints, i.e., dissolved oxygen production, algal growth rate, and cell density. The proposed test revealed excellent test sensitivity and reproducibility. Currently, none of the existing algal toxicity test protocols is adequate for assessing the toxicity of organic chemicals. The closed-system algal toxicity tests developed by previous researchers also may not be ideal because the enlarged headspace and/or enriched bicarbonate buffer may result in either underestimations of the exposure concentrations or insensitive responses to both heavy metals and organic toxicants. Compared to the aforementioned algal toxicity test methods, the proposed technique in the present study has a more general applicability under conditions such as effluent samples containing both metals and organic toxicants or samples with unknown compositions.     

Criteria and methods for evaluation of toxic hazard

Evaluating the toxic hazard of fire gases has been an area of recent interest. Units of measure for toxic hazard based on the concept of dose are presented. The role of carbon monoxide and hydrogen cyanide as principal toxicants in combustion products is reviewed. The importance of the time-related aspects of smoke transport and exposure to the toxicants is discussed.     

柔性聚氨酯泡沫燃烧产物毒性评估

介绍了对弹性聚氨酯泡沫燃烧产物毒性的相关研究,并对其是否能用于毒性危害分析进行了评估。研究显示,暴露到聚氨酯燃烧产物中的人会导致中毒。窒息气体氮气有效剂量法能有效预测受害人因暴露于聚氨酯燃烧产物而导致的行为能力丧失情况。同时,也证明有效剂量法不能正确预测聚氨酯阴燃时情况。该方法不适用于热分解。研究发现,目前很多经验方法具有很大的局限性,对毒性危害研究造成了很多负面的影响。烟气粒子携带毒物的吸入和传播以及化学合成复杂的毒物,研究和考虑得很少。因此,很有必要对复杂毒性有机体的火灾燃烧产物进行更深入的物理和化学分析研究以及动物模型研究。     

The effect of applied direct current on biological phosphorus uptake

An increase in metabolic activity of bacteria capable of accumulating excess phosphorus may also improve the phosphorus removal efficiency of the biological phosphorus removal system.It has been observed that the amount of intracellular biochemical energy (ATP) in bacterial cells has increased when they were exposed to pH or electrical stress. The utilization of the surplus intracellular energy generated by the stress may be utilized to stimulate the metabolic activity of bacterial cells.It was hypothesized that the introduction of direct current into mixed liquor might also increase the metabolic activity of bacteria capable of accumulating excess phosphorus.A direct current with 6 V and 200 mA was applied intermittently for one minute in every five. The direct current was introduced at the anaerobic step of a Phoredox type laboratory scale continuous flow system. Graphite electrodes were used to introduce the current. To separate the effect of the electric current from that caused by other environmental factors a control system was run in parallel using the same operational parameters. Synthetic wastewater with the same composition was used as feed for both systems.As a result of the intermittent application of direct current, the rate of phosphorus accumulation in activated sludge increased by 30% during the start-up period of the biological system and by 11 % during steady state conditions. The applied direct current changed the movement and concentration of potassium ions. The pH of the mixed liquor in the anaerobic reactor dropped to 4 as soon as the direct current was introduced and returned to the initial pH 6 value when the current flow ceased. The number of viable cells in the activated sludge decreased by a factor of 2–5 as a result of the electrification. The reduction observed in viable cell count did not influence the overall organic carbon removal efficiency or the nitrification-denitrification processes.A hypothetical model was developed which combines known biological reactions with results observed during the research. The model summarizes the effects of direct current on biological phosphorus uptake.This work provides insight into a method of enhancing the biological removal of phosphorus from a wastewater stream without adding chemicals. It has many practical applications where electric power is available but chemical addition is unacceptable (water reuse), the cost is prohibitive or the appropriate chemicals are locally unavailable.     

Toxicity characterization of urban stormwater with bioanalytical tools

Stormwater harvesting has become an attractive alternative strategy to address the rising demand for urban water supply due to limited water sources and population growth. Nevertheless, urban stormwater is also a major source of surface water pollution. Runoff from different urban catchments with source contributions from anthropogenic activities and various land uses causes variable contaminant profiles, thus posing a challenging task for environmental monitoring and risk assessment. A thorough understanding of raw stormwater quality is essential to develop appropriate treatment facilities for potential indirect potable reuse of stormwater. While some of the key chemical components have previously been characterized, only scarce data are available on stormwater toxicity. We benchmarked stormwater samples from urban, residential and industrial sites across various Australian capital cities against samples from the entire water cycle, from sewage to drinking water. Six biological endpoints, targeting groups of chemicals with modes of toxic action of particular relevance for human and environmental health, were investigated: non-specific toxicity (Microtox and combined algae test), the specific modes of action of phytotoxicity (combined algae test), dioxin-like activity (AhR-CAFLUX), and estrogenicity (E-SCREEN), as well as reactive toxicity encompassing genotoxicity (umuC) and oxidative stress (AREc32). Non-specific toxicity was highly variable across sites. The baseline toxicity equivalent concentrations of the most polluted samples were similar to secondary treated effluent from wastewater treatment plants. Phytotoxicity results correlated well with the measured herbicide concentrations at all sites. High estrogenicity was found in two sampling events and could be related to sewage overflow. Genotoxicity, dioxin-like activity, and oxidative stress response were evident in only three of the samples where the stormwater drain was beside a heavy traffic road, confirming that road runoff is the potential source of contaminants, while the bioanalytical equivalent concentrations (BEQ) of these samples were similar to those of raw sewage. This study demonstrates the benefit of bioanalytical tools for screening-level stormwater quality assessment, forming the basis for the evaluation of future stormwater treatment and reuse schemes.     

An analysis of the combined effects of organic toxicants

This paper presents a basic database for the joint actions of 44 binary mixtures of various organic toxicants on Escherichia coli. The multiple toxicity behaviors observed from the E. coli organisms were analyzed and compared with previous works based on the Microtox tests. The two kinds of tests produced quite different responses, in terms of the joint action mode and the sum of toxic units, to various organic mixtures. However, detailed analyses with the considerations of the chemical's mechanisms of toxicity and the slope of toxicant's dose-response curve have revealed several general criteria for the prediction of combined effects of organic toxicants. First, for both reactive and non-reactive toxicants, either additive or less than additive (antagonistic) joint actions will be observed for chemicals of the same mechanism of toxicity. Second, the mixture of reactive toxicants with different mechanisms is the only category of organic mixtures associated with frequent observations of synergism. Third, greater-than-additive (synergistic) effects are inherently associated with toxicants having flat dose-response curves. Less than additive effects are, however, mainly related to a chemical's display steep dose-response curves. Model analyses indicate that the observed synergistic effects are due to response addition or response multiplication joint actions. Hence, most of the synergistic joint actions are non-interactive in nature and are governed by the dose-response relationships of individual toxicants.     

Health-based targets for greywater reuse in residential schools in Madhya Pradesh,India

In line with developments in the water reuse sector, this paper applied quantitative microbial risk assessment (QMRA) techniques to seven greywater reuse systems used for recycling shower water for toilet flushing. The objective of the study was to establish a scientific basis for health-based greywater targets for India. It involved qualitative risk assessments and quantitative microbiological analysis using Enterococci, thermotolerant coliforms (TTC) and coliphage indicator organisms. Four conclusions are drawn from this study. Firstly, the systems indicated a low risk and high quality, and secondly that low levels of risk were present in systems, resulting in the recommendation of guideline of 5000 cfu/100 mL rather than 10 000 cfu/100 mL for greywater utilised for direct toilet flushing. Thirdly, disability adjusted life years (DALYs) are a useful indicator of risk for evaluating the performance of a greywater reuse system in addition to chemical/microbiological indicators DALYs, and finally that TTC are a useful surrogate microbial indicator for analysis of greywater in developing countries with limited analytical facilities.     

Evaluation of water quality control by using test organisms and their cells

The use of biotesting for quality assessment of different waters. This method consists in determining the action of toxicants on specially selected organisms under standard conditions with registration of changes at behavioral, physiological, cellular and subcellular levels. A micronucleus test and Arneth’s formula as a biomarker were proposed for an optimal set used to determine specific structural and functional changes in the cell genome caused by toxic effects. A particular attention was paid to assessing the risk to human health of those factors and substances, the genotoxicity and cytotoxicity of which can be identified by using biomarkers of plant and animal cells.     

Water Management In Textile Industry: Technical And Economic Aspects

Textile industry is an intensive water use sector. Consequently, enormous quantities of wastewater are generated from different manufacturing processes. Numerous opportunities for water reuse could be exploited by rational management of water use and reuse via segregation and upgrading of specific effluents using membrane filtration systems comprising ultrafiltration (UF) and reverse osmosis (RO). This paper is an endeavor to develop technoeconomic indicators for extended water reuse applications in textile industry using UF and RO. Extensive data gathered from a typical Egyptian dye house have been utilized to rationalize water use and reuse management based on different levels of pollutants in waste streams. A developed rationale enabled the identification of four applicable intervention scenarios comprising three reuse scenarios ranging from limited reuse (about 25%) to maximum reuse (about 87%) and a total end of the pipe treatment scenario. The financial assessment for a typical wastewater load (about 3500 r m/day) reveals that the maximum reuse scenario comprising UF, two stage RO in addition to direct use and low press RO is the most preferred one. Further, sensitivity analysis indicates that the annual cost of the total end of pipe treatment scenario will equal the annual cost of the maximum reuse scenario when the unit cost are US$ 0.5, 0.43 and 0.3 per cubic meter for wastewater treatment, RO and UF respectively.     

Combination of in vitro bioassays encompassing different mechanisms to determine the endocrine-disrupting effects of river water

In this study, the total toxic effects of river water samples were assessed using a series of cell culture bioassays which encompassed different mechanisms, based on specific modes of action. River water samples were collected from three tributaries of the Youngsan River in the western portion of Korea. We confirmed that Youngsan River water was polluted with a complex mixture of estrogenic and dioxin-like compounds. The total toxic effects of the downstream water samples were found to be higher than that of the upstream water samples. In the upstream water samples, total estrogenic activity was measured to be between 0.005 and 0.049 ng-EEQ/l (17beta-estradiol-equivalent concentration) and no CYP1A activity was detected. In the downstream water samples, however, total estrogenic activity was measured to be between 0.021 ng-EEQ/l and 1.918 ng-EEQ/l, and total CYP1A activity was between 0.63 and 3.55 microg-MEQ/l (3-methylcholanthrene-equivalent concentration). When assessed according to a concentration-response curve, downstream water sample extracts exerted dual actions on estrogen receptors, depending on the concentration volume of the samples. The concentration volume range proximal to the original water sample exhibited estrogenic activity, whereas antiestrogenic activity was observed at high concentration volumes (more than 5 times concentration) in the extracts. This study involved a combination of in vitro bioassays, designed to encompass different mechanisms. The bioassays used included the estrogen receptor binding affinity test, E-screen assay, aromatase assay, and EROD assay. These tests provided a great deal of useful information regarding the potency and action modes of estrogenicity and antiestrogenicity inherent in the sampled river water. Although further study is necessary to determine the relationship between toxic responses in in vitro bioassay systems and chronic toxicity in aquatic organisms, our approach is expected to be fairly accurate with regard to the detection of endocrine-disrupting effects in an aquatic environment.     

A bacterial bioassay for assessment of wastewater toxicity

A bioassay using freeze-dried Nitrobacter as the test organism has been shown to successfully detect various toxicants in municipal and industrial wastewaters. The test is simple, sensitive, rapid and inexpensive; as a result, this test shows potential as a quantitative measurement technique for wastewater toxicity.The bioassay technique was applied in the analysis of a wastewater treatment system at a fiberboard plant near Portland, Oregon. The fiberboard is manufactured from a slurry of chipped wood, shavings and sawdust.Wastewater from the process is treated at an on-site facility consisting of settling ponds, activated sludge basin, clarifier and holding ponds. Treated water is recycled to the mill for reuse. Recently the removal of BOD by the activated sludge system dropped significantly and soluble and particulate organics began to accumulate. The influent and effluent flows for the treatment facility were tested with the Nitrobacter bioassay and both were determined to have significant toxicity. Further tests at points in the process showed that the toxicity was not associated with a single waste stream and was prevalent throughout the entire wastewater treatment system. In an effort to identify the toxicant, toxicity tests were conducted for known chemicals used in the process. The wastewater was treated with various physical and chemical unit processes to determine the most effective method for toxicity removal.Due to the complexity of the wastewater composition, no specific agents have been identified as solely responsible for the observed toxic response, however, several possible explanations for the apparent toxicity are discussed.     

Ozonation and activated carbon treatment of sewage effluents: removal of endocrine activity and cytotoxicity

Concerns about endocrine disrupting compounds in sewage treatment plant (STP) effluents give rise to the implementation of advanced treatment steps for the elimination of trace organic contaminants. The present study investigated the effects of ozonation (O₃) and activated carbon treatment (AC) on endocrine activities [estrogenicity, anti-estrogenicity, androgenicity, anti-androgenicity, aryl-hydrocarbon receptor (AhR) agonistic activity] with yeast-based bioassays. To evaluate the removal of non-specific toxicity, a cytotoxicity assay using a rat cell line was applied. Wastewater (WW) was sampled at two STPs after conventional activated sludge treatment following the secondary clarifier (SC) and after subsequent advanced treatments: O₃, O₃ + sand filtration (O_3-SF), and AC. Conventional treatment reduced estrogenicity, androgenicity, and AhR agonistic activity by 78-99% compared to the untreated influent WW. Anti-androgenicity and anti-estrogenicity were not detectable in the influent but appeared in SC, possibly due to the more effective removal of respective agonists during conventional treatment. Endocrine activities after SC ranged from 2.0 to 2.8 ng/L estradiol equivalents (estrogenicity), from 4 to 22 μg/L 4-hydroxytamoxifen equivalents (anti-estrogenicity), from 1.9 to 2.0 ng/L testosterone equivalents (androgenicity), from 302 to 614 μg/L flutamide equivalents (anti-androgenicity), and from 387 to 741 ng/L β-naphthoflavone equivalents (AhR agonistic activity). In particular, estrogenicity and anti-androgenicity occurred in environmentally relevant concentrations. O₃ and AC further reduced endocrine activities effectively (estrogenicity: 77-99%, anti-androgenicity: 63-96%, AhR agonistic activity: 79-82%). The cytotoxicity assay exhibited a 32% removal of non-specific toxicity after O₃ compared to SC. O₃ and sand filtration reduced cytotoxic effects by 49%, indicating that sand filtration contributes to the removal of toxicants. AC was the most effective technology for cytotoxicity removal (61%). Sample evaporation reduced cytotoxic effects by 52 (AC) to 73% (O₃), demonstrating that volatile substances contribute considerably to toxic effects, particularly after O₃. These results confirm an effective removal or transformation of toxicants with receptor-mediated mode of action and non-specific toxicants during O₃ and AC. However, due to the limited extractability, polar ozonation by-products were neglected for toxicity analysis, and hence non-specific toxicity after O₃ is underestimated.     

滤池反冲洗废水的直接回流利用研究

通过小试研究了滤池反冲洗废水直接回流利用对出水水质的影响.结果表明,反冲洗废水的直接回流利用对混凝过程有较为明显的强化作用,能够降低出水浊度和残余铝含量;当回流比例<10%时,对出水TOC没有影响;反冲洗废水的直接回流利用会导致出水细菌总数有少量增加,但不会影响消毒效果,同时对总余氯衰减的影响也很小.UFC试验的结果表明,反冲洗废水直接回流利用仅会造成消毒副产物前质含量轻微升高,其增幅为4.5%～7.6%.     

Application of an ecosystem model for aquatic ecological risk assessment of chemicals for a Japanese lake.

The Lake Suwa version of the comprehensive aquatic systems model (CASM-SUWA) was developed using field data from Lake Suwa and evaluated to examine the utility of CASM-SUWA for assessing the ecological risk of chemicals for aquatic ecosystems. The calibration of the parameters for the model provided that the established reference model simulation could reproduce complex seasonal biomass behavior of populations that were not significantly different from the general seasonal pattern for the Lake Suwa ecosystem. The sensitivity analyses revealed the potential importance of indirect effects and demonstrated that the parameter values of all the trophic levels were important in determining the biomass of each trophic level in the model. The risk estimation of linear alkylbenzene sulfonates (LAS) demonstrated that the model estimated the risks of direct toxic effects on each population and the indirect ecological effects that propagate through the food-web in the model ecosystem. The CASM-SUWA-derived benchmark levels were approximately one order of magnitude less than the field-derived NOECs in literature. The analyses of the comparison implied that the model could provide a good basis in determining an ecological protective level of a chemical of concern in aquatic ecosystem. This modeling study demonstrated that the model can be used to provide additional information for the decision-making process in the management of the aquatic ecological risk of chemicals.     

Growth response of the green algae Chlorella vulgaris to selective concentrations of zinc

A modification of a previously published fish cell culture technique (Rachlin and Perlmutter, 1968) has been successfully employed in this study to evaluate the effects of a metal toxicant on the growth response and therefore productivity of the green algae Chlorella vulgaris. The results of this study indicate that the concentration of zinc which reduces the growth rate of a test population of this alga, during a 96 h exposure, by 50 per cent, is 2.4±0.02 ppm. The test design has advantages in that it concerns itself with productivity reduction rather than standard survival (LC₅₀) of the test organisms. In addition the data generated is readily treated by the system of probit analysis for ease of evaluating the results. The use of this technique and test system, allows for the design of parallel studies in which the effects of toxicants on both animal and plant cell systems can be evaluated in terms of a more unified cellular approach.