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Additional information

In vitro data

ZPS was tested in a GLP compliant Ames reverse mutation assay according to OECD guideline 471 using Salmonella typhimurium strains TA 1535, TA 1537, TA 98, and TA 100 and Escherichia coli WP2 uvr A at 33 to 5000 µg/plate (two independent experiments (standard plate incorporation test and pre-incubation test) and each concentration was tested in triplicate) with and without metabolic activation (RCC Ltd., 2001). The plates incubated with ZPS showed normal background growth up to 5000 µg/plate with and without metabolic activation in both independent experiments. No relevant toxic effects, evident as a reduction in the number of revertants, occurred in the test groups with and without metabolic activation. No substantial increase in revertant colony numbers of any of the five tester strains was observed following treatment with ZPS at any dose level, neither in the presence nor absence of metabolic activation (S9 mix). There was also no tendency of higher mutation rates with increasing concentrations in the range below the generally acknowledged border of biological relevance. Appropriate reference mutagens were used as positive controls and showed a distinct increase of induced revertant colonies. In conclusion, under the experimental conditions reported, ZPS did not induce gene mutations by base pair changes or frameshifts in the genome of the strains used.

 

In a second GLP compliant reverse mutation assay according to OECD 471 using Salmonella typhimurium strains TA98, TA100, TA1535, TA1537 and TA1538, ZPS was tested at concentrations of 0.2 to 20 µL/plate (Microbiological Associates, Inc., 1986). Under the conditions of this test, the substance did not cause a positive response on any of the tester strains with or without metabolic activation.

ZPS was tested in the Unscheduled DNA Synthesis Test (GLP compliant and performed according to a protocol similar to OECD guideline 482) using rat primary hepatocytes. The test article was tested at nine dose levels ranging from 0.003 µL/mL to 3.0 µL/mL (Microbiological Associates, Inc., 1987). Dose levels 0.01 to 0.6 µL/mL were fully evaluated, dose levels 1.0 - 3.0 µL/mL caused too much toxicity to evaluate the nuclei for UDS. The results of the UDS assay indicate that under the test conditions, the test article did not cause a significant increase in the mean number of net nuclear grain counts (i.e., an increase of at least 5 counts over the control), at any dose level. Therefore, the test article is considered negative in this study.

 

ZPS was also tested in a GLP-compliant mouse lymphoma L5178Y cells (TK assay tested according to OECD guideline 476) (Sitek Research Laboratories, 1987).Cultures were treated at concentrations of 1.0, 8.0, 15, 22, 29, 36, 43 and 50 µL/mL in the absence of exogenous activation and at concentrations of 0.02,0.16, 0.30,0.44,0.58, 0.72, 0.86 and 1.0 µL/mL in the presence of S-9 mix. The results indicated that in the presence and absence of exogenous activation, the test article induced a significant increase in the mutant frequency of treated cultures as compared to the mutant frequency of the corresponding solvent control cultures. The response was dose dependent.

 

 

In vivo data

A GLP compliant bone marrow chromosome aberration assay according to a protocol similar to OECD guideline 475 was performed with rats (Microbiological Associates, Inc., 1989). Male and female Sprague-Dawley rats were treated with the substance at dose volumes of 1.6, 5.3 or 16 mL/kg body weight which was given as a single gavage administration. All animals receiving 16 mL/kg and one male receiving 5.3 mL/kg exhibited dark blue-green faeces, approximately 5 hours after dosing for the 8 hour harvest. All animals receiving the substance exhibited dark blue-green faeces approximately 10 hours after dosing for the 12 hour harvest. All animals receiving distilled water appeared normal throughout the study. No significant reduction in the rate of body weight gain was observed among test article-treated animals.

There was no apparent change in ploidy in any of the treated animals. ZPS had no apparent effect on the mitotic index. The percentage of cells containing one or more aberrations was not significantly increased relative to the vehicle control in any of the test article treated animals. The mean aberrations per cell per animal were also not significantly increased regardless of dose or sacrifice time. The negative and positive controls fulfilled the requirements for determination of a valid test.

Under the conditions of this assay, ZPS, was negative in the acute cytogenetics assay using male and female Sprague-Dawley rats.

 

A sex-linked recessive lethal (SLRL) assay was performed in Drosophila melanogaster (University of Wisconsin, 1991). Adult Canton-S males were subjected to a 3 day feeding exposure to 250 µL/mL ZPS. Other Canton-S males were injected with ZPS at a concentration of 0.2 µL/mL. Exposure via feeding induced approximately 8% mortality and no male sterility was induced. Exposure via injection induced approximately 9% mortaility and also no male sterility was induced. The Canton-S males were mated to Basc females and germ cells which were post-mitotic at the time of exposure were tested for lethal mutations. ZPS did not induce mutations in the post-meiotic germ cells of Drosophila melanogaster males when administered by feeding or injection.

Conclusion

From the toxicokinetic studies with ZPS, it is known that zinc as measured by 65Zn is absorbed to a small extent after oral absorption and that the organic part of the ZPS molecule is most likely excreted without absorption. The very small amount of absorption as measured by 14C (organic part of the molecule) after oral and dermal absorption could be accounted for by absorption of radiochemical impurities in the dosing preparation.

Zinc is known to give (false) positive results in in vitro genetic toxicity studies. In the EU Risk Assessment Report of zinc (http://esis.jrc.ec.europa.eu/doc/existing-chemicals/risk_assessment/DRAFT/R072_0609_env_hh.pdf), a negative (Amacher and Paillet, 1980) and two positive results (Cameron, 1991; Adams and Kirkpatrick, 1994) were reported in mouse lymphoma assays.

However, the EU Risk Assessment Report concluded that there is no evidence for genotoxicity in vivo for zinc. Furthermore, zinc is an essential trace element.

Therefore, the positive result in the mouse lymphoma assay for ZPS is not considered relevant for the in vivo situation. Overall, based on the available other in vitro and in vivo genotoxicity studies with ZPS which all gave negative results and the knowledge on the genotoxicity profile of zinc, ZPS is not considered genotoxic.

Short description of key information:
Based on the available genotoxicity studies with ZPS and the knowledge on the genotoxicity profile of zinc, ZPS is not considered genotoxic.

Endpoint Conclusion: No adverse effect observed (negative)

Justification for classification or non-classification

Based on the results of the in vitro and in vivo tests, ZPS does not need to be classified according to Directive 67/548/EEC and according to the EU Classification, Labelling and Packaging of Substances and Mixtures (CLP) Regulation (EC) No. 1272/2008.