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Thousands different chemicals are discharged into the environment from agriculture, industry, medical facilities, house-holds. Currently, there is an increasing concern for the environmental impact of mixture of compounds since the additive and eventual synergistic effects are unknown and could produce serious adverse effects. Recently, a document from the European Commission on combination effects of chemicals highlighted the need to ensure that risks associated with chemical mixtures are properly understood and assessed. To address this issue, a joint-effort of 16 European and associated research groups participated to an exercise to test a synthetic reference chemical mixture on the own routine bioassays to investigate the chemical mixtures effects. The reference material included class of pesticides, pharmaceuticals, industrial products, heavy metals and polyaromatic hydrocarbons. The mixures were prepared, each compound at Equivalent Quality Standard (EQS) value, the safety limit concentration allowed by the European Water Framework Directive, (WFD).The bioassays proposed by the groups could cover the entire ecosystem from bacteria to fish as well in vitro assays providing an unique scenario from ecological risk assessment perspective. The results showed that effects were observed at very low concentration on algal-bacteria composition in a marine microcosm, immobilization in crustacean, fish embryo toxicity and frog embryo development. We conclude that some precaution on the chemical mixture assessment should be taken even in case the individual compounds are present at EQS, the safety limit concentration under European legislation.

Thousands different chemicals are discharged into the environment from agriculture, industry, medical facilities, house-holds. Currently, there is an increasing concern for the environmental impact of mixture of compounds since the additive and eventual synergistic effects are unknown and could produce serious adverse effects. Recently, a document from the European Commission on combination effects of chemicals highlighted the need to ensure that risks associated with chemical mixtures are properly understood and assessed. To address this issue, a joint-effort of 16 European and associated research groups participated to an exercise to test a synthetic reference chemical mixture on the own routine bioassays to investigate the chemical mixtures effects. The reference material included class of pesticides, pharmaceuticals, industrial products, heavy metals and polyaromatic hydrocarbons. The mixures were prepared, each compound at Equivalent Quality Standard (EQS) value, the safety limit concentration allowed by the European Water Framework Directive, (WFD).The bioassays proposed by the groups could cover the entire ecosystem from bacteria to fish as well in vitro assays providing an unique scenario from ecological risk assessment perspective. The results showed that effects were observed at very low concentration on algal-bacteria composition in a marine microcosm, immobilization in crustacean, fish embryo toxicity and frog embryo development. We conclude that some precaution on the chemical mixture assessment should be taken even in case the individual compounds are present at EQS, the safety limit concentration under European legislation.

Thousands different chemicals are discharged into the environment from agriculture, industry, medical facilities, house-holds. Currently, there is an increasing concern for the environmental impact of mixture of compounds since the additive and eventual synergistic effects are unknown and could produce serious adverse effects. Recently, a document from the European Commission on combination effects of chemicals highlighted the need to ensure that risks associated with chemical mixtures are properly understood and assessed. To address this issue, a joint-effort of 16 European and associated research groups participated to an exercise to test a synthetic reference chemical mixture on the own routine bioassays to investigate the chemical mixtures effects. The reference material included class of pesticides, pharmaceuticals, industrial products, heavy metals and polyaromatic hydrocarbons. The mixures were prepared, each compound at Equivalent Quality Standard (EQS) value, the safety limit concentration allowed by the European Water Framework Directive, (WFD).The bioassays proposed by the groups could cover the entire ecosystem from bacteria to fish as well in vitro assays providing an unique scenario from ecological risk assessment perspective. The results showed that effects were observed at very low concentration on algal-bacteria composition in a marine microcosm, immobilization in crustacean, fish embryo toxicity and frog embryo development. We conclude that some precaution on the chemical mixture assessment should be taken even in case the individual compounds are present at EQS, the safety limit concentration under European legislation.

Thousands different chemicals are discharged into the environment from agriculture, industry, medical facilities, house-holds. Currently, there is an increasing concern for the environmental impact of mixture of compounds since the additive and eventual synergistic effects are unknown and could produce serious adverse effects. Recently, a document from the European Commission on combination effects of chemicals highlighted the need to ensure that risks associated with chemical mixtures are properly understood and assessed. To address this issue, a joint-effort of 16 European and associated research groups participated to an exercise to test a synthetic reference chemical mixture on the own routine bioassays to investigate the chemical mixtures effects. The reference material included class of pesticides, pharmaceuticals, industrial products, heavy metals and polyaromatic hydrocarbons. The mixures were prepared, each compound at Equivalent Quality Standard (EQS) value, the safety limit concentration allowed by the European Water Framework Directive, (WFD).The bioassays proposed by the groups could cover the entire ecosystem from bacteria to fish as well in vitro assays providing an unique scenario from ecological risk assessment perspective. The results showed that effects were observed at very low concentration on algal-bacteria composition in a marine microcosm, immobilization in crustacean, fish embryo toxicity and frog embryo development. We conclude that some precaution on the chemical mixture assessment should be taken even in case the individual compounds are present at EQS, the safety limit concentration under European legislation.

Thousands different chemicals are discharged into the environment from agriculture, industry, medical facilities, house-holds. Currently, there is an increasing concern for the environmental impact of mixture of compounds since the additive and eventual synergistic effects are unknown and could produce serious adverse effects. Recently, a document from the European Commission on combination effects of chemicals highlighted the need to ensure that risks associated with chemical mixtures are properly understood and assessed. To address this issue, a joint-effort of 16 European and associated research groups participated to an exercise to test a synthetic reference chemical mixture on the own routine bioassays to investigate the chemical mixtures effects. The reference material included class of pesticides, pharmaceuticals, industrial products, heavy metals and polyaromatic hydrocarbons. The mixures were prepared, each compound at Equivalent Quality Standard (EQS) value, the safety limit concentration allowed by the European Water Framework Directive, (WFD).The bioassays proposed by the groups could cover the entire ecosystem from bacteria to fish as well in vitro assays providing an unique scenario from ecological risk assessment perspective. The results showed that effects were observed at very low concentration on algal-bacteria composition in a marine microcosm, immobilization in crustacean, fish embryo toxicity and frog embryo development. We conclude that some precaution on the chemical mixture assessment should be taken even in case the individual compounds are present at EQS, the safety limit concentration under European legislation.

International audience ; Estrogenic compounds are widely released to surface waters and may cause adverse effects to sensitive aquatic species. Three hormones, estrone, 17β-estradiol and 17α-ethinylestradiol, are of particular concern as they are bioactive at very low concentrations. Current analytical methods are not all sensitive enough for monitoring these substances in water and do not cover mixture effects. Bioassays could complement chemical analysis since they detect the overall effect of complex mixtures. Here, four chemical mixtures and two hormone mixtures were prepared and tested as reference materials together with two environmental water samples by eight laboratories employing nine in vitro and in vivo bioassays covering different steps involved in the estrogenic response. The reference materials included priority substances under the European Water Framework Directive, hormones and other emerging pollutants. Each substance in the mixture was present at its proposed safety limit concentration (EQS) in the European legislation. The in vitro bioassays detected the estrogenic effect of chemical mixtures even when 17β-estradiol was not present but differences in responsiveness were observed. LiBERA was the most responsive, followed by LYES. The additive effect of the hormones was captured by ERα-CALUX, MELN, LYES and LiBERA. Particularly, all in vitro bioassays detected the estrogenic effects in environmental water samples (EEQ values in the range of 0.75–304 × EQS), although the concentrations of hormones were below the limit of quantification in analytical measurements. The present study confirms the applicability of reference materials for estrogenic effects' detection through bioassays and indicates possible methodological drawbacks of some of them that may lead to false negative/positive outcomes. The observed difference in responsiveness among bioassays – based on mixture composition - is probably due to biological differences between them, suggesting that panels of bioassays with different characteristics should be applied according to specific environmental pollution conditions.

Estrogenic compounds are widely released to surface waters and may cause adverse effects to sensitive aquatic species. Three hormones, estrone, 17β-estradiol and 17α-ethinylestradiol, are of particular concern as they are bioactive at very low concentrations. Current analytical methods are not all sensitive enough for monitoring these substances in water and do not cover mixture effects. Bioassays could complement chemical analysis since they detect the overall effect of complex mixtures. Here, four chemical mixtures and two hormone mixtures were prepared and tested as reference materials together with two environmental water samples by eight laboratories employing nine in vitro and in vivo bioassays covering different steps involved in the estrogenic response. The reference materials included priority substances under the European Water Framework Directive, hormones and other emerging pollutants. Each substance in the mixture was present at its proposed safety limit concentration (EQS) in the European legislation. The in vitro bioassays detected the estrogenic effect of chemical mixtures even when 17β-estradiol was not present but differences in responsiveness were observed. LiBERA was the most responsive, followed by LYES. The additive effect of the hormones was captured by ERα-CALUX, MELN, LYES and LiBERA. Particularly, all in vitro bioassays detected the estrogenic effects in environmental water samples (EEQ values in the range of 0.75–304 × EQS), although the concentrations of hormones were below the limit of quantification in analytical measurements. The present study confirms the applicability of reference materials for estrogenic effects' detection through bioassays and indicates possible methodological drawbacks of some of them that may lead to false negative/positive outcomes. The observed difference in responsiveness among bioassays – based on mixture composition - is probably due to biological differences between them, suggesting that panels of bioassays with different characteristics should be ...

International audience ; Estrogenic compounds are widely released to surface waters and may cause adverse effects to sensitive aquatic species. Three hormones, estrone, 17β-estradiol and 17α-ethinylestradiol, are of particular concern as they are bioactive at very low concentrations. Current analytical methods are not all sensitive enough for monitoring these substances in water and do not cover mixture effects. Bioassays could complement chemical analysis since they detect the overall effect of complex mixtures. Here, four chemical mixtures and two hormone mixtures were prepared and tested as reference materials together with two environmental water samples by eight laboratories employing nine in vitro and in vivo bioassays covering different steps involved in the estrogenic response. The reference materials included priority substances under the European Water Framework Directive, hormones and other emerging pollutants. Each substance in the mixture was present at its proposed safety limit concentration (EQS) in the European legislation. The in vitro bioassays detected the estrogenic effect of chemical mixtures even when 17β-estradiol was not present but differences in responsiveness were observed. LiBERA was the most responsive, followed by LYES. The additive effect of the hormones was captured by ERα-CALUX, MELN, LYES and LiBERA. Particularly, all in vitro bioassays detected the estrogenic effects in environmental water samples (EEQ values in the range of 0.75–304 × EQS), although the concentrations of hormones were below the limit of quantification in analytical measurements. The present study confirms the applicability of reference materials for estrogenic effects' detection through bioassays and indicates possible methodological drawbacks of some of them that may lead to false negative/positive outcomes. The observed difference in responsiveness among bioassays – based on mixture composition - is probably due to biological differences between them, suggesting that panels of bioassays with different ...

International audience ; Estrogenic compounds are widely released to surface waters and may cause adverse effects to sensitive aquatic species. Three hormones, estrone, 17β-estradiol and 17α-ethinylestradiol, are of particular concern as they are bioactive at very low concentrations. Current analytical methods are not all sensitive enough for monitoring these substances in water and do not cover mixture effects. Bioassays could complement chemical analysis since they detect the overall effect of complex mixtures. Here, four chemical mixtures and two hormone mixtures were prepared and tested as reference materials together with two environmental water samples by eight laboratories employing nine in vitro and in vivo bioassays covering different steps involved in the estrogenic response. The reference materials included priority substances under the European Water Framework Directive, hormones and other emerging pollutants. Each substance in the mixture was present at its proposed safety limit concentration (EQS) in the European legislation. The in vitro bioassays detected the estrogenic effect of chemical mixtures even when 17β-estradiol was not present but differences in responsiveness were observed. LiBERA was the most responsive, followed by LYES. The additive effect of the hormones was captured by ERα-CALUX, MELN, LYES and LiBERA. Particularly, all in vitro bioassays detected the estrogenic effects in environmental water samples (EEQ values in the range of 0.75–304 × EQS), although the concentrations of hormones were below the limit of quantification in analytical measurements. The present study confirms the applicability of reference materials for estrogenic effects' detection through bioassays and indicates possible methodological drawbacks of some of them that may lead to false negative/positive outcomes. The observed difference in responsiveness among bioassays – based on mixture composition - is probably due to biological differences between them, suggesting that panels of bioassays with different characteristics should be applied according to specific environmental pollution conditions.

International audience ; Estrogenic compounds are widely released to surface waters and may cause adverse effects to sensitive aquatic species. Three hormones, estrone, 17β-estradiol and 17α-ethinylestradiol, are of particular concern as they are bioactive at very low concentrations. Current analytical methods are not all sensitive enough for monitoring these substances in water and do not cover mixture effects. Bioassays could complement chemical analysis since they detect the overall effect of complex mixtures. Here, four chemical mixtures and two hormone mixtures were prepared and tested as reference materials together with two environmental water samples by eight laboratories employing nine in vitro and in vivo bioassays covering different steps involved in the estrogenic response. The reference materials included priority substances under the European Water Framework Directive, hormones and other emerging pollutants. Each substance in the mixture was present at its proposed safety limit concentration (EQS) in the European legislation. The in vitro bioassays detected the estrogenic effect of chemical mixtures even when 17β-estradiol was not present but differences in responsiveness were observed. LiBERA was the most responsive, followed by LYES. The additive effect of the hormones was captured by ERα-CALUX, MELN, LYES and LiBERA. Particularly, all in vitro bioassays detected the estrogenic effects in environmental water samples (EEQ values in the range of 0.75–304 × EQS), although the concentrations of hormones were below the limit of quantification in analytical measurements. The present study confirms the applicability of reference materials for estrogenic effects' detection through bioassays and indicates possible methodological drawbacks of some of them that may lead to false negative/positive outcomes. The observed difference in responsiveness among bioassays – based on mixture composition - is probably due to biological differences between them, suggesting that panels of bioassays with different characteristics should be applied according to specific environmental pollution conditions.

The risk posed by complex chemical mixtures in the environment to wildlife and humans is increasingly debated, but has been rarely tested under environmentally relevant scenarios. To address this issue, two mixtures of 14 or 19 substances of concern (pesticides, pharmaceuticals, heavy metals, polyaromatic hydrocarbons, a surfactant, and a plasticizer), each present at its safety limit concentration imposed by the European legislation, were prepared and tested for their toxic effects. The effects of the mixtures were assessed in 35 bioassays, based on 11 organisms representing different trophic levels. A consortium of 16 laboratories was involved in performing the bioassays. The mixtures elicited quantifiable toxic effects on some of the test systems employed, including i) changes in marine microbial composition, ii) microalgae toxicity, iii) immobilization in the crustacean Daphnia magna, iv) fish embryo toxicity, v) impaired frog embryo development, and vi) increased expression on oxidative stress-linked reporter genes. Estrogenic activity close to regulatory safety limit concentrations was uncovered by receptor-binding assays. The results highlight the need of precautionary actions on the assessment of chemical mixtures even in cases where individual toxicants are present at seemingly harmless concentrations.

The risk posed by complex chemical mixtures in the environment to wildlife and humans is increasingly debated, but has been rarely tested under environmentally relevant scenarios. To address this issue, two mixtures of 14 or 19 substances of concern (pesticides, pharmaceuticals, heavy metals, polyaromatic hydrocarbons, a surfactant, and a plasticizer), each present at its safety limit concentration imposed by the European legislation, were prepared and tested for their toxic effects. The effects of the mixtures were assessed in 35 bioassays, based on 11 organisms representing different trophic levels. A consortium of 16 laboratories was involved in performing the bioassays. The mixtures elicited quantifiable toxic effects on some of the test systems employed, including i) changes in marine microbial composition, ii) microalgae toxicity, iii) immobilization in the crustacean Daphnia magna, iv) fish embryo toxicity, v) impaired frog embryo development, and vi) increased expression on oxidative stress-linked reporter genes. Estrogenic activity close to regulatory safety limit concentrations was uncovered by receptor-binding assays. The results highlight the need of precautionary actions on the assessment of chemical mixtures even in cases where individual toxicants are present at seemingly harmless concentrations.

The risk posed by complex chemical mixtures in the environment to wildlife and humans is increasingly debated, but has been rarely tested under environmentally relevant scenarios. To address this issue, two mixtures of 14 or 19 substances of concern (pesticides, pharmaceuticals, heavy metals, polyaromatic hydrocarbons, a surfactant and a plasticizer), each present at its safety limit concentration imposed by the European legislation, were prepared and tested for their toxic effects. The effects of the mixtures were assessed in 35 bioassays, based on eleven organisms representing different trophic levels. A consortium of 16 laboratories was involved in performing the bioassays. The mixtures elicited quantifiable toxic effects on some of the test systems employed, including i) changes in marine microbial composition, ii) microalgae toxicity iii) immobilization in the crustacean Daphnia magna, iii) fish embryo toxicity, iv) impaired frog embryo development and v) increased expression on oxidative stress-linked reporter genes. Estrogenic activity close to regulatory safety limit concentrations was uncovered by receptor-binding assays. The results highlight the need of precautionary actions on the assessment of chemical mixtures even in cases where individual toxicants are present at seemingly harmless concentrations.