Chloramide

Endpoint summary

Key value for chemical safety assessment

Genetic toxicity in vivo

Description of key information

In vitro:

Mammalian gene mutagen: not mutagenic

Mutagenic activity using the S. typhimurium TA 100 tester strain: not mutagenic

Reversion of TrpC to trp+ in B. subtilis: weak mutagen

Mutagenic activity using S. typhimurium TA102, TA100, TA 98, TA97: positive with TA100, TA98 and TA97; negative with TA102

In vivo:

- Chromosomal aberrations, CD-1 mouse bone-marrow cells: negative.

- Aneuploidy, bone-marrow cells of CD-1 mouse: negative.

- Sperm morphology, B6C3F1 mice: negative.

Link to relevant study records

Reference

Endpoint:

in vivo mammalian somatic cell study: cytogenicity / bone marrow chromosome aberration

Remarks:

Type of genotoxicity: chromosome aberration

Type of information:

experimental study

Adequacy of study:

key study

Study period:

no data

Reliability:

2 (reliable with restrictions)

Rationale for reliability incl. deficiencies:

other: Comparable to guideline study with acceptable restrictions.

Qualifier:

equivalent or similar to guideline

Guideline:

OECD Guideline 475 (Mammalian Bone Marrow Chromosome Aberration Test)

Deviations:

yes

Remarks:

four animals per group instead of five. First prelevement at 6 hours after the treatment instead of 12-18h.

GLP compliance:

no

Type of assay:

chromosome aberration assay

Species:

mouse

Strain:

Swiss

Sex:

male/female

Details on test animals or test system and environmental conditions:

TEST ANIMALS
- Source: Charles River Breeding Laboratories, Inc.
- Age at study initiation: 8-11 week
- Weight at study initiation: no data
- Assigned to test groups randomly: yes
- Fasting period before study: no data
- Housing: animals, were group housed, separated by sex and by treatment group
- Diet: ad libitum
- Water : ad libitum
- Acclimation period: no data

ENVIRONMENTAL CONDITIONS
- Temperature (°C): no data
- Humidity (%): no data
- Air changes (per hr): no data
- Photoperiod (hrs dark / hrs light): no data

IN-LIFE DATES: no data

Route of administration:

oral: gavage

Vehicle:

- Vehicle(s)/solvent(s) used: deionized water

Details on exposure:

Animals were dosed by oral gavage with 1 ml of test solution on a subchronic regimen (five daily administration approximately 24hr apart). Concurrent negative controls received deionized water, the diluent of the test solutions. Positive controls were also included to ensure that the assay were working properly. Generally these were also concurrent. The intraperitoneal (IP) route was used for administration of the positive control chemicals since this route was found to be suitable for eliciting consistent, effective responses in all assays employed. In addition to the subchronic dosing regimen an acute administration was also employed in this assay.

Duration of treatment / exposure:

Five daily administrations approximately 24hr appart or an acute administration.

Frequency of treatment:

daily for the subchronic assay or one-time administration for the acute assy

Post exposure period:

Animals dosed acutely were sacrificed 6, 24, and 48 hr after exposure; animals dosed subchronically were sacrificed 6 hr after the last exposure.

Remarks:

Doses / Concentrations:
0, 50, 100 and 200 mg/l
Basis:
nominal in water

No. of animals per sex per dose:

4

Control animals:

yes, concurrent no treatment

Positive control(s):

triethylenemelamine (1 mg/kg in 0.9 % saline)

Tissues and cell types examined:

Bone marrow cells

Details of tissue and slide preparation:

DETAILS OF SLIDE PREPARATION:
After the last dose administration, animals were killed with CO2 or by cervical dislocation and the tibiae excised. The marrow was flushed from the bone into centrifuge tubes containing 3 ml of fetal calf serum. The marrow obtained in Hanks' balanced salt solution was treated, after centrifugation, successively with hypotonic (0.075 M) KCl and with fixative (3:1 methanol:acetic acid). After storage of the fixed material at 4°C for at least overnight, slides were prepared and stained with 5-10 % Giemsa at pH 6.8.

Evaluation criteria:

A mitotic index was determined by scoring the number of cells in mitosis based on at least 500 cells. Fifty metaphase spreads were scored for each animal where possible for structural and numerical aberrations. Numerical aberrations included cells showing either hyperploidy or polyploidy. Strutural aberrations included chromosome and chromatid breaks, chromatid deletions, fragments, translocations, triradials, quadriradials, pulverized chromosomes, pulverized cells, complex rearrangements, ring chromosomes, dicentric chromosomes, and minute chromosomes. Four endpoints were evaluated for statistically significant differences between the response at each treatment level and the concurrent negative control. The endpoints were (1) number of structural aberrations present per animal, (2) number of numerical aberrations present per animal, (3) percentage of cells with at least one structural aberration present per animal, and (4) percentage of cells with two or more structural aberrations per animal.Data for male anf female animals were analyzed both separately and combined.

Statistics:

A Student's t-test was used to test for differences between each treatment level and the current negative control. A significant level of 0.01 was used to indicate a positive response.

Sex:

male/female

Genotoxicity:

negative

Toxicity:

not specified

Vehicle controls validity:

valid

Negative controls validity:

not examined

Positive controls validity:

valid

Additional information on results:

RESULTS OF DEFINITIVE STUDY
-- Types of structural aberrations for significant dose levels (for Cytogenetic or SCE assay): Negative

Conclusions:

Interpretation of results (migrated information): negative
Under the conditions of this test, Monochloramine gave no significant increases in the number of strutural or numerical abberations for either pooled or individual sex data.

Executive summary:

The present study was carried out to determine whether monochloramine induce genotoxic effects following oral administration to mice. Then, a mouse bone marrow cytogenetics assay was conducted. Four males and four females were used for each treatment group (3 dose levels of the test solution and controls). Doses of 0, 50, 100 and 200 mg/l (nominal in water). Animals were dosed by oral gavage with 1 ml of test solution on a subchronic regimen (five daily administration approximately 24 hr apart.) or on a single acute dose administration. Concurrent negative controls received deionized water, the diluent of the test solutions. Positive controls were also included to ensure that the assays were working properly. The positive control was 1 mg/kg triethylenemelamine (TEM) in 0.9 % saline, administered via the intraperitoneal route as a one-time (acute) administration concurrently with the 24 hr acute component of the study.Three hours prior to sacrifice, animals were injected IP with 4.0 mg/kg colchicine to collect mataphases. Six hours after the last (fifth) dose administration (subchronic dosing) or 6, 24, and 48 hr (acute dosing), animals were killed and bone marrow were harvested. A mitotic index was determined by scoring the number of cells in mitosis based on at least 500 cells. Fifty metaphase spreads were scored for each animal where possible for structural and numerical aberrations. Numerical aberrations included cells showing either hyperploidy or polyploidy. Strutural aberrations included chromosome and chromatid breaks, chromatid deletions, fragments, translocations, triradials, quadriradials, pulverized chromosomes, pulverized cells, complex rearrangements, ring chromosomes, dicentric chromosomes, and minute chromosomes. Four endpoints were evaluated for statistically significant differences between the response at each treatment level and the concurrent negative control. The endpoints were (1) number of structural aberrations present per animal, (2) number of numerical aberrations present per animal, (3) percentage of cells with at least one structural aberration present per animal, and (4) percentage of cells with two or more structural aberrations per animal. Data for male anf female animals were analyzed both separately and combined. Under the conditions of this test, Monochloramine gave no significant increases in the number of strutural or numerical abberations for either pooled or individual sex data. The positive control chemical, TEM, induced a significant increases in both structural and numerical chromosomal aberrations relative to control.

Endpoint conclusion

Endpoint conclusion:

no adverse effect observed (negative)

Additional information

Additional information from genetic toxicity in vivo:

There are in vitro studies available addressing mutagenicity in bacteria and an in vitro Mutation Test using Mouse Lymphoma L5178Y cells was conducted in compliance with the good laboratory practice standards. In addition, in vivo studies from the open scientific literature are available addressing micronucleus formation, chromosomal aberrations or aneuploidy in the bone marrow of CD-1 mice or sperm abnormalities in B6C3F1 mice.

Monochloramine did not induce a mutagenic effect in Salmonella typhimurium strain TA100 (Thomas et al., 1987) and was a weak mutagen in the reversion of trpC to trp+ in B. subtilis (Shih and Lederberg, 1976). Concentrated extracts of water samples treated with monochloramine showed mutagenic activity in the Ames/Salmonella assay with TA100, TA98 and TA97 strains and showed negative result with TA102 (Schenck et al., 1980). In the in vitro mammalian cell mutation assay designed to comply with the OECD Guidelines No. 476, Monochloramine did not demonstrate mutagenic potential.

In a single in vivo study, monochloramine did not induce micronucleus formation, chromosomal aberrations or aneuploidy in the bone marrow of CD-1 mice or sperm abnormalities in B6C3F1 mice. While another in vivo study reported positive results for rat bone marrow chromosomal aberrations after treatment with monochloramine, this study was disregarded as sufficient detail was not provided to allow a critical evaluation (Fujie et Aoki, 1998).

Based on the existing studies, monochloramine appears to have some mutagenic potential in vitro in bacterial reverse mutation test, but no mutagenic potential was detected in a reliable mouse lymphoma L5178Y TK+/- assay. Moreover, no significant in vivo

mutagenic properties were observed.

Justification for selection of genetic toxicity endpoint

Only one study with several in vivo data is available (Chromosomal aberrations, CD-1 mouse bone-marrow cells/ Aneuploidy, bone-marrow cells of CD-1 mouse/ Sperm morphology, B6C3F1 mice).

Justification for classification or non-classification

The test material appears to have some mutagenic potential in vitro, but no significant mutagenic properties in vivo. Therefore, Monochloramine do not have to be classified for genetic toxicity according to the criteria of the DSD or the CLP regulation.