ISO/TC 147/SC 5/WG 9 - Genotoxicity and endocrine effects

This document specifies a method for the determination of the estrogenic potential of water and waste water by means of a reporter gene assay with genetically modified yeast strains Saccharomyces cerevisiae. This reporter gene assay is based on the activation of the human estrogen receptor alpha. This method is applicable to: — fresh water; — waste water; — aqueous extracts and leachates; — eluates of sediments (fresh water); — pore water; — aqueous solutions of single substances or of chemical mixtures; — drinking water. The limit of quantification (LOQ) of this method for the direct analysis of water samples is between 8 ng/l and 15 ng/l 17β-estradiol equivalents (EEQ) based on the results of the international interlaboratory trial (see Annex F). The upper threshold of the dynamic range for this test is between 120 ng/l and 160 ng/l 17β-estradiol equivalents (EEQ). Samples showing estrogenic potencies above this threshold have to be diluted for a valid quantification. Extraction and pre-concentration of water samples can prove necessary, if their estrogenic potential is below the given LOQ.

This document specifies a method for the determination of the estrogenic potential of water and waste water by means of a reporter gene assay utilizing stably transfected human cells. This reporter gene assay is based on the activation of the human estrogen receptor alpha. This method is applicable to: — fresh water; — waste water; — aqueous extracts and leachates; — eluates of sediments (fresh water); — pore water; — aqueous solutions of single substances or of chemical mixtures; — drinking water; — the limit of quantification (LOQ) of this method for the direct analysis of water samples is between 0,3 ng/l and 1 ng/l 17β-estradiol equivalents (EEQ) based on the results of the international interlaboratory trial (see Annex F). The upper working range was evaluated [based on the results of the international interlaboratory trial (see Table F.3)] up to a level of 75 ng EEQ/l. Samples showing estrogenic potencies above this threshold have to be diluted for a valid quantification. Extraction and pre concentration of water samples can prove necessary if their estrogenic potential is below the given LOQ.

This document specifies a method for the determination of the estrogenic potential of water and waste water by means of a reporter gene assay with a genetically modified yeast strain Arxula adeninivorans. This reporter gene assay is based on the activation of the human estrogen receptor alpha. Arxula adeninivorans is a highly robust and salt- and temperature-tolerant test organism and is especially suitable for the analysis of samples with high salinity (conductivity up to 70 mS/cm). The test organism can be cultivated in medium with sodium chloride content up to 20 %. This method is applicable to: — fresh water; — waste water; — sea water; — brackish water; — aqueous extracts and leachates; — eluates of sediments (fresh water); — pore water; — aqueous solutions of single substances or of chemical mixtures; — drinking water. The limit of quantification (LOQ) of this method for the direct analysis of water samples is between 1,5 ng/l and 3 ng/l 17β-estradiol equivalents (EEQ). The upper threshold of the dynamic range for this test is between 25 ng/l and 40 ng/l 17β-estradiol equivalents (EEQ). Samples showing estrogenic potencies above this threshold have to be diluted for a valid quantification. Extraction and pre-concentration of water samples can prove necessary, if their estrogenic potential is below the given LOQ. An international interlaboratory trial for the validation of this document has been carried out. The results are summarized in Annex F. NOTE Extraction and pre-concentration of water samples can prove necessary.

This International Standard specifies a method for the determination of the genotoxic potential of water and waste water using the bacterial strains Salmonella enterica subsp. enterica serotype Typhimurium TA 98 and TA 100 in a fluctuation assay. This combination of strains is able to measure the genotoxicity of chemicals that induce point mutations (base pair substitutions and frameshift mutations) in genes coding for enzymes that are involved in the biosynthesis of the amino acid, histidine. NOTE 1 ISO 13829[8] applies for the measurement of genotoxicity of samples containing DNA-crosslinking agents. This method is applicable to:
— fresh water;
— waste water;
— aqueous extracts and leachates;
— eluates of sediments (fresh water);
— pore water;
— aqueous solutions of single substances or of chemical mixtures;
— drinking water.
NOTE 2 When testing drinking water, extraction and pre-concentration of water samples can prove necessary.

ISO 21427-2:2006 specifies a method for the determination of genotoxicity of water and waste water using a mammalian in vitro test which detects damage, induced by water-soluble substances, to the chromosomes or the mitotic apparatus of V79 cells from the Chinese hamster. The micronucleus test allows the identification of substances that cause cytogenetic damage which results in the formation of micronuclei containing lagging chromosome fragments and/or whole chromosomes. The assay is based on the increase in the frequency of micronucleated cells after incubation with and without metabolic activation.

ISO 16240:2005 specifies a method for the determination of the genotoxic potential of water and wastewater using the bacterial strains Salmonella typhimurium TA 100 and TA 98. This method includes sterile filtration of water and wastewater prior to the test. ISO 16240:2005 is applicable only to the detection of genotoxic substances which are in the filtered aqueous phase. It is not applicable to the detection of genotoxic substances adsorbed by the retained particles.