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Measures for protecting water quality: current approaches and future developments.

An important component in protecting the ecological quality of watercourses is the regulation of point source discharges. These are usually expressed in terms of "end of pipe" limits on certain physical parameters or on the concentrations of specific chemicals. Sometimes discharge licences take the form of Emission Limit Values (ELVs) which are based on Best Available Technology with respect to emissions and apply to all discharges (usually for a specified industry sector) irrespective of the dilution capacity of the receiving watercourse. Alternatively, they may be based on water quality standards (e.g., EQSs) for specific chemicals which describe thresholds below which no adverse impact on the receiving water is predicted and which take explicit account of available dilution at different discharge locations. The strengths and limitations of these approaches are reviewed, along with a consideration of new approaches that address some of the limitations associated with control measures based on ELVs or EQSs. These include approaches for controlling well-defined mixtures by a "Toxic Equivalent" approach, exemplified here for discharges containing alkylphenol ethoxylate surfactants, and a fundamentally different approach for regulating complex effluents based on Direct Toxicity Assessment of whole effluents. Circumstances under which these different approaches to controlling point source emissions to surface waters might be appropriate are discussed.

Conservation of Natural Resources↗

Chemical-activated luciferase gene expression (CALUX): a novel in vitro bioassay for Ah receptor active compounds in sediments and pore water.

This study demonstrates that the novel in vitro CALUX (chemical-activated luciferase expression) assay is a rapid, sensitive assay for assessing the toxic potency of (mixtures of) aryl hydrocarbon receptor (AhR)-active compounds in sediments and pore waters. A rat hepatoma (H4IIE) cell line, stably transfected with a construct containing the dioxin-responsive element (DRE) sequence and the luciferase reporter gene, was used to determine the relative potency or the total activities of AhR-active compounds in sediment and pore water extracts. This novel CALUX assay had a detection limit of 0.5 fmol of 2,3,7,8-tetrachlorodibenzo-p-dioxin (TCDD). The sensitivity and linear working range was slightly better than for the ethoxyresorufin O-deethylase (EROD) assay in H4IIE wild type cells. The primary improvement of the CALUX assay compared to the EROD assay, however, is that the CALUX assay is insensitive to substrate inhibition. The CALUX activity induced by organic extracts from 450-mg aliquots of sediment or 250-microl aliquots of pore water corresponded with the instrumentally analyzed degree of pollution of the sediment. Using pore water, only a simple and rapid extraction procedure was needed, without additional clean-up to prevent cell death. The response from pore water samples in an 8-day early life stage test with zebra fish (Branchydanio rerio) corresponded with the CALUX induction, although the correlation was sometimes disturbed by heavy metals. Two polychlorinated terphenyl mixtures, the PCB-substitute Ugilec 141, polybrominated diphenylethers, and the PCB-mixture Clophen A50 were tested in the CALUX assay and had induction potencies that were 10(-4)-10(-7) compared to TCDD.

Animals↗

Effects of heroin, alone or in combination with other drugs, on the locomotor activity in two inbred strains of mice.

The locomotor activity of C57Bl/6J and DBA/2J mice was studied, under the influences of heroin, amphetamine, strychnine, or ethanol, and of combinations of the opiate with each one of the other drugs. Heroin treatment was followed by the typical "running fit" in the C57 mice, while the DBA strain was unaffected. Amphetamine enhanced the activity in the C57 strain only. The combination of heroin with amphetamine or ethanol increased the locomotor activity only in the DBA strain, while heroin + strychnine exerted a clear stimulating effect on the activity of the C57 mice. The strychnine + heroin mixture was more toxic than heroin alone when the lethal doses (LD50) were determined in the 2 strains.

Amphetamine↗

Dietary therapy in two patients with vitamin B12-unresponsive methylmalonic acidemia.

The biochemical and therapeutic responses to dietary therapy were studied in a 25-month-old girl and a 1-month-old girl with methylmalonic acidemia (MMA-emia), which was unresponsive to vitamin B12. The minimum daily intake of protein which patients could tolerate and display a good development was between 1.0 and 1.2 g per kg body weight. Supplementation with amino acid mixture devoid of toxic amino acids was required to prevent protein malnutrition when daily protein intake was restricted to 0.6 g per kg body weight. Caloric intake should be sufficient, not only to promote growth but also to prevent a rise in MMA level, especially when a patient has ketoacidosis. It was found that MMA excretion per mg creatinine in random urine specimens correlated significantly with serum MMA and twenty four-hour output of MMA per kg body weight. Therefore measurement of MMA in a single urine specimen is useful for evaluating the in vivo accumulation of MMA.

Acidosis↗

Isolation of a lethal toxin from Cortinarius orellanus Fr.

The fungal toxin 'orellanine', isolated by Grzymala (10) from Cortinarius orellanus, seems to be a heterogeneous mixture of several toxic and nontoxic substances. We succeeded in isolating a lethal toxin which is sensitive to light and which is striking because of its long latency period. We consider this the main toxin of C. orellanus. Even in extremely high doses of ten times LD 100 mice do not die earlier than 48 hours after application. The paper gives details about the isolation and several properties of this slow acting toxin.

2,2'-Dipyridyl↗

Toxicity equivalents and EPA's risk assessment of 2,3,7,8-TCDD.

Toxicity equivalent factors (TEFs) have proved useful in estimating the toxicity of complex mixtures of chlorinated dibenzo-p-dioxins and dibenzofurans (CDDs/CDFs). An international consensus has formed around a specific set of TEF values as interim solution for addressing environmental contamination by CDDs/CDFs. This procedure capitalizes on the congener-specific analytical results that are more routinely available in recent years. The TEF approach should be updated as necessary and replaced by more definitive bioassay approaches as soon as practicable. In an independent activity, the USEPA considered a proposal to change (reduce) the cancer potency ascribed to 2,3,7,8-TCDD by a factor of 16. The recommendation was based upon an analysis of the literature and the Agency's earlier risk assessment. The proposal was reviewed by the Science Advisory Board, a group of outside scientific advisors. Subsequently, the Agency decided against making any changes in its assessment at this time. However, it is likely that a reassessment will be conducted shortly that will incorporate new data and a new approach to estimating of cancer risks posed by 2,3,7,8-TCDD.

Animals↗

Influence of combined alpha, beta and gamma radionuclide contamination on the frequency of waxy-reversions in barley pollen.

The mutagenic effectiveness of chronic, combined alpha-, beta-, and gamma-irradiation caused by radionuclide contamination of the biogeocenosis was studied in comparison with chronic gamma-irradiation from an external 60Co source. The waxy-test on barley pollen proved to be sufficiently sensitive to evaluate the increase in waxy-reversion frequencies in plants exposed to a 3 mSv dose. A dose of 50-200 mSv from external irradiation was needed to double the mutation rate in barley pollen. Moreover, the smaller doses (dose rates) were more harmful than the higher doses if measured as the increase in mutation frequency per unit of irradiation. Higher genotoxicity was observed from combined irradiation due to radionuclide contamination than from similar doses from an external gamma source. We concluded that mutagenic effects in plant cells, in the case of radionuclide contamination of the biogeocenosis, could not be extrapolated only on the basis of measured dose, but separate investigations should be carried out to establish the genotoxicity of specific radionuclide mixtures and possible toxic chemical synergisms.

Accidents↗

The mutagenic potencies of plant extracts containing quercetin in Salmonella typhimurium TA98 and TA100.

Four commercial ethanolic plant extracts, Tinctura Alchemillae, Extractum Crataegi, Extractum Myrtilli and Tinctura Hyperici, were tested for their mutagenicity in Salmonella typhimurium TA98 and TA100 with and without S9 mix obtained from rats pretreated with phenobarbital. The extracts studied differed greatly in their mutagenic potencies but exhibited a very similar mutation pattern in which the strongest effect was always seen in tester strain TA98 with S9 mix. Simultaneously we investigated the extracts for the presence of quercetin and kaempferol. Only quercetin was detected in small amounts by thin-layer chromatography (TLC). The fractions containing quercetin were separated and collected using a Sephadex LH-20 column. Two different methods were employed to estimate the amount of quercetin in the extracts: a colorimetric assay developed by Christ and Müller, and a complexometric method by Belikov. The quercetin concentrations ranged between 2 mg (Tinctura Alchemilla) and 89 mg (Tinctura Hyperici) per 100 g of extract. We suggest that the mutagenicity of the 4 plant extracts is mainly due to the presence of quercetin for the following reasons: (1) all the plant extracts exhibit a mutation pattern which is very similar to that of quercetin, (2) the mutagenic potential of the extracts correlates well with their quercetin content, considering the fact that plant extracts are very complex mixtures often containing toxic or antimutagenic compounds.

Animals↗

Hepatic metallothionein production and resistance to heavy metals by rainbow trout (Salmo gairdneri)--II. Held in a series of contaminated lakes.

Rainbow trout held in net pens for 4 weeks in three locations contaminated by heavy metals developed elevated levels of hepatic metallothionein but no acclimation to a combination of zinc, copper and cadmium. Toxicity of the mixture of zinc, copper and cadmium in a ratio of 400:20:1 increased with temperature in the range of 6-21 degrees C. Hepatic metallothionein concentrations after 4 weeks of exposure in situ was correlated with the degree of contamination as measured by zinc concentrations (P less than 0.01).

Adaptation, Physiological↗

Phytotoxicity of surface waters of the Thames and Brisbane River estuaries: a combined chemical analysis and bioassay approach for the comparison of two systems.

The Thames Estuary, UK, and the Brisbane River, Australia, are comparable in size and catchment area. Both are representative of the large and growing number of the world's estuaries associated with major cities. Principle differences between the two systems relate to climate and human population pressures. In order to assess the potential phytotoxic impact of herbicide residues in the estuaries, surface waters were analysed with a PAM fluorometry-based bioassay that employs the photosynthetic efficiency (photosystem II quantum yield) of laboratory cultured microalgae, as an endpoint measure of phytotoxicity. In addition, surface waters were chemically analysed for a limited number of herbicides. Diuron, atrazine and simazine were detected in both systems at comparable concentrations. In contrast, bioassay results revealed that whilst detected herbicides accounted for the observed phytotoxicity of Brisbane River extracts with great accuracy, they consistently explained only around 50% of the phytotoxicity induced by Thames Estuary extracts. Unaccounted for phytotoxicity in Thames surface waters is indicative of unidentified phytotoxins. The greatest phytotoxic response was measured at Charing Cross, Thames Estuary, and corresponded to a diuron equivalent concentration of 180 ng L(-1). The study employs relative potencies (REP) of PSII impacting herbicides and demonstrates that chemical analysis alone is prone to omission of valuable information. Results of the study provide support for the incorporation of bioassays into routine monitoring programs where bioassay data may be used to predict and verify chemical contamination data, alert to unidentified compounds and provide the user with information regarding cumulative toxicity of complex mixtures.

Australia↗

Inactivation of antineoplastics in clinical wastewater by electrolysis.

Wastewater from clinical institutions contains a considerable amount of toxic substances. Among the toxic substances, antineoplastics may induce carcinogenesis, teratogenesis, and the emergence of mutant microorganisms in the environment. Although the incineration or chemical treatments of disposed antineoplastics are recommended, a high energy during incineration and a careful quality control during chemical treatment are required. In this study, we determined the conditions for the electrolytic treatment of an antineoplastic, epirubicin hydrochloride (EH), using two platinum electrodes with a constant current of 100 mA. We analyzed the cytotoxicity, mutagenicity and antibacterial activity of electrolyzed EH and compared them with those of unelectrolyzed EH. Nearly 100% cytotoxicity, mutagenicity and antibacterial activity were eliminated and HPLC did not detect an EH molecule, in the case of electrolysis for 6 h. We also examined the biological cytotoxicities of electrolyzed irinotecan hydrochloride, vincristine sulfate, mitomycin C, paclitaxel, methotrexate and cisplatin, and found that 72.1-99.999% toxicity was eliminated by electrolysis under the same conditions. The biological toxicity of a mixture of these drugs was determined to be decreased by approximately 99% by electrolysis under the same conditions.

Antibiotics, Antineoplastic↗

Dynamic ultrasound-assisted extraction of oleuropein and related biophenols from olive leaves.

A continuous approach for the ultrasound-assisted extraction of olive biophenols (OBPs) from olive leaves is proposed. Multivariate methodology was used to carry out a detailed optimisation of extraction. Under the optimal working conditions, complete extraction of the target analytes (namely, oleuropein, verbacoside, apigenin-7-glucoside and luteolin-7-glucoside with LODs 11.04, 2.68, 1.49 and 3.91 mg/kg, respectively) was achieved in 25 min. The extract was injected into a chromatograph-photodiode array detector assembly (HPLC-DAD) for individual separation-quantification. No clean-up or preconcentration steps were required. Gas chromatography-mass spectrometry (without derivatization of the analytes) was used to identify OBPs at concentrations below the LODs obtained by HPLC-DAD. The efficacy of ethanol-water mixtures to extract OBPs from olive leaves has been demonstrated and compared with that of a conventional method which requires 24h for complete extraction; so these mixtures can substitute toxic extractants used to date.

Chromatography, High Pressure Liquid↗

Emerging chemicals of concern: pharmaceuticals and personal care products (PPCPs) in Asia, with particular reference to Southern China.

In many western nations, pharmaceuticals and personal care products (PPCPs) are present in aquatic environments, raising concerns amongst chemists and toxicologists regarding their potential environmental fates and effects. However, there are few published reports of PPCPs in environmental samples from Southeast Asia. Whilst the environmental toxicology of PPCPs is not well understood, several effects cause concern, such as feminisation or masculinisation by hormones and xenoestrogens, synergistic toxicity from complex mixtures at low concentrations, potential creation of resistant strains in natural bacterial populations, and other potential concerns for human health. Whilst both the presence and distributions of PPCPs in Southeast Asia and China are not well known, observations elsewhere suggest that they may be important contaminants in the aquatic environment. This is particularly emphasised by the enormous production and widespread use of many PPCPs in China, particularly antibiotics utilised in human and veterinary medicine applications. This Viewpoint presents a general description of the issue, characterises the current status of PPCP analyses and reporting in the Southeast Asian region, and proposes a recommended approach for monitoring and chemical assessment of one group of PPCPs, antibiotics, in the aquatic environments of Hong Kong and the Pearl River Delta.

China↗

A practical ranking system to compare toxicity of anti-fouling paints.

The toxicity of a number of new anti-fouling paints, claimed to function by physical means and not by leakage of toxic substances, have been tested on two common organisms in the Baltic Sea, i.e., the red macro alga Ceramium tenuicorne and the copepod Nitocra spinipes. In order to compare the toxicity between the paints a ranking system was developed based on the EC(50)- and LC(50)-values. The results showed a wide span in toxicity with the most toxic paints ranked 160 times more toxic than the ones ranked least toxic. Also, TBT, irgarol and diuron, which have been used as active ingredients in traditional anti-fouling paints, were used to evaluate the sensitivity of the two test organisms. The results showed that the test organisms were equally sensitive to the substances as similar organisms in earlier studies. In conclusion, the ranking system presented in this study permits ranking and comparison of total toxicity of complex mixtures.

Animals↗

An integrated biomarker-based strategy for ecotoxicological evaluation of risk in environmental management.

Environmental impacts by both natural events and man-made interventions are a fact of life; and developing the capacity to minimise these impacts and their harmful consequences for biological resources, ecosystems and human health is a daunting task for environmental legislators and regulators. A major challenge in impact and risk assessment, as part of integrated environmental management (IEM), is to link harmful effects of pollution (including toxic chemicals) in individual sentinel animals to their ecological consequences. This obstacle has resulted in a knowledge-gap for those seeking to develop effective policies for sustainable use of resources and environmental protection. Part of the solution to this problem may lie with the use of diagnostic clinical-type laboratory-based ecotoxicological tests or biomarkers, utilising sentinel animals as integrators of pollution, coupled with direct immunochemical tests for contaminants. These rapid and cost-effective ecotoxicological tools can provide information on the health status of individuals and populations based on relatively small samples of individuals. In the context of ecosystem status or health of the environment, biomarkers are also being used to link processes of molecular and cellular damage through to higher levels (i.e., prognostic capability), where they can result in pathology with reduced physiological performance and reproductive success. Complex issues are involved in evaluating environmental risk, such as the effects of the physico-chemical environment on the speciation and uptake of pollutant chemicals and inherent inter-individual and inter-species differences in vulnerability to toxicity; and the toxicity of complex mixtures. Effectively linking the impact of pollutants through the various hierarchical levels of biological organisation to ecosystem and human health requires a pragmatic integrated approach based on existing information that either links or correlates processes of pollutant uptake, detoxication and pathology with each other and higher level effects. It is further proposed here that this process will be facilitated by pursuing a holistic or whole systems approach with the development of computational simulation models of cells, organs and animals in tandem with empirical data (i.e., the middle-out approach). In conclusion, an effective integrated environmental management strategy to secure resource sustainability requires an integrated capability for risk assessment and prediction. Furthermore, if such a strategy is to influence and help in the formulation of environmental policy decisions, then it is crucial to demonstrate scientific robustness of predictions concerning the long-term consequences of pollution to politicians, industrialists and environmental managers; and also increase stakeholder awareness of environmental problems.

Animals↗

Genotoxicity evaluation of individuals working with photocopying machines.

Photocopying machines are a common sight in the cities of India. There is ample evidence showing that the components of toners individually or in the form of a complex mixture are genotoxic. Toxic components of the photocopiers are from their emissions, toners and extremely low frequency electromagnetic fields (ELF-EMFs). In the present study micronucleus test (MNT) on buccal epithelial cells, cytokinesis block micronucleus (CBMN) assay and chromosomal aberration analysis on peripheral blood mononuclear cells was performed on 98 workers occupationally involved in photocopying and 90 age and sex matched controls. The results showed a significant increase in the frequency of MN in buccal epithelial cells and peripheral blood lymphocytes, as well as chromosomal aberrations in the exposed as compared to the control subjects.

Adolescent↗

Environmental monitoring using acetylcholinesterase inhibition in vitro. A case study in two Mexican lagoons.

Cholinesterase inhibition is considered a specific biomarker of exposure and effect for organophosphorous pesticides. Its use for monitoring has been hindered, particularly in tropical countries where organophosphates are widely used for malaria and dengue control, because of the frequent lack of suitable controls. An in vitro technique is proposed as a biochemical method for monitoring pollutant mixtures in sediment toxicity tests. Brain homogenate from the fish Oreochromis niloticus is used as the enzyme source. Optimum incubation time, extraction solvent and effect of crude oil on acetylcholinesterase (AChE) are reported. The method described was used in sediments from two Mexican lagoons, located in an oil extraction area where pesticides are used in agriculture and vector control campaigns. AChE inhibitions from 3 to 21% were found in these lagoons, even in the presence of high concentrations of petroleum.

Acetylcholinesterase↗

Biocomplexity: the post-genome challenge in ecotoxicology.

There are four crucial challenges for the environmental toxicologists in the next decade: (1) understanding the mechanisms of molecular and subcellular interactions with pollutant chemicals, including genomic and proteomic aspects; (2) the development of predictive simulation models of toxic effects on complex cellular and physiological processes; (3) linking molecular, cellular and patho-physiological 'endpoints' with higher level ecological consequences; and (4) precautionary anticipation of possible harmful impacts of novel developments in industrial processes, including biotechnology and nanotechnology. One of the major difficulties in ecotoxicology is to link harmful effects of chemical pollutants in individual animals and plants with the ecological consequences. Consequently, this obstacle has resulted in a 'knowledge-gap' for those seeking to develop policies for sustainable use of resources and environmental protection. The overall problem is: how to develop effective procedures for environmental/ecological impact and risk assessment? However, the use of diagnostic 'clinical-type' tests or 'biomarkers' has started to provide information on the health-status of populations based on relatively small samples of individuals. Also, biomarkers can now be used to begin to link processes of molecular and cellular damage through to the higher levels (i.e. prognostic capability), where they can result in reduced performance and reproductive success. Research effort to meet this challenge must be inter-disciplinary in character, since the key questions mainly involve complex interfacial problems. These include effects of physico-chemical speciation on uptake and toxicity, the toxicity of complex mixtures; and linking the impact of pollutants through the various hierarchical levels of biological organisation to ecosystem and human health. Finally, the development and use of process-based computational simulation models (i.e. 'virtual' cells, organs and animals), illustrated using an endosomal/lysosomal uptake and cell injury model, will facilitate the development of a predictive capacity for estimating risk associated with the possibility of future environmental events.

Animals↗