Reaction mass of copper oxide and manganese dioxide

Acute Toxicity: oral

Administrative data

Endpoint:

acute toxicity: oral

Type of information:

experimental study

Adequacy of study:

key study

Study period:

year of publication: 2013

Reliability:

2 (reliable with restrictions)

Rationale for reliability incl. deficiencies:

guideline study without detailed documentation

Remarks:

no detailed information on observation periods, preparation of test item, and study design

Cross-reference

Reason / purpose for cross-reference:

reference to same study

Reference

Endpoint:

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

Type of information:

experimental study

Adequacy of study:

supporting study

Study period:

year of publication: 2013

Reliability:

2 (reliable with restrictions)

Rationale for reliability incl. deficiencies:

comparable to guideline study with acceptable restrictions

Remarks:

not fully conducted according to current guideline (older version), documentation lacking

Qualifier:

equivalent or similar to guideline

Guideline:

OECD Guideline 475 (Mammalian Bone Marrow Chromosome Aberration Test)

Version / remarks:

21 JUL 1997

Deviations:

no

Principles of method if other than guideline:

- Principle of test: bone marrow chromosome aberration assay
- Short description of test conditions: analysis was performed after rinsing the bone marrow cells (femur and tibia) from the rats as described by Adler (1984) Mutagenicity Testing: A Practical Approach Venitt S, Parry J (eds). IRL Press: Oxford; 273–306
- Parameters analysed / observed: determination of chromosome aberrations in metaphases, and mitotic index

GLP compliance:

not specified

Type of assay:

other: chromosome aberration assay

Specific details on test material used for the study:

The study investigated test item in form of nano- and micron-sized particles. The mean size distribution of test item particles were 45 ± 17 nm for nano-particles and 2.74 ± 29 µm for micron-particles. Both particles acted comparable in the CA assay, therefore this record focuses on micron-sized particles only.

SOURCE OF TEST MATERIAL
- Source and lot/batch No.of test material: Sigma Chemical Co., St. Louis, USA
- Expiration date of the lot/batch: not specified
- Purity: ≥99%

STABILITY AND STORAGE CONDITIONS OF TEST MATERIAL
- Solubility and stability of the test substance in the solvent/vehicle: suspended in Milli Q water


TREATMENT OF TEST MATERIAL PRIOR TO TESTING
- Treatment of test material prior to testing: suspended various doses of test item in Milli Q water after sonication using a probe sonicator (UPH 100, Germany) for 10 min at 90% amplitude


OTHER SPECIFICS:
- specific surface area analysis was determined by using the Brunauer–Emmett–Teller technique = 7.95 (m2 /g)

Species:

rat

Strain:

Wistar

Remarks:

albino

Sex:

female

Details on test animals or test system and environmental conditions:

TEST ANIMALS
- Source: National Institute of Nutrition, Hyderabad, India
- Females (if applicable) nulliparous and non-pregnant: not specified
- Age at delivery: 6 to 8 weeks
- Weight at delivery: 80–120 g
- Fasting period before study: not specified
- Housing: in groups of five in polypropylene cages
- Diet: commercial pellet diet
- Water: ad libitum
- Acclimation period: one week

ENVIRONMENTAL CONDITIONS
- Temperature (°C): 22 ± 3
- Humidity (%): 55–65
- Air changes (per hr): not specified
- Photoperiod (hrs dark / hrs light): 12 /12

IN-LIFE DATES: From: To: not specified

Route of administration:

oral: gavage

Vehicle:

- Vehicle(s)/solvent(s) used: Milli Q water

Details on exposure:

- suspended in Milli Q water and then applied
- No information on vehicle volume for dose groups available, but control group received 5 ml/kg bw of Milli Q water.

Duration of treatment / exposure:

single administration

Frequency of treatment:

single administration

Post exposure period:

24 h

Dose / conc.:

0 mg/kg bw/day

Dose / conc.:

100 mg/kg bw/day

Dose / conc.:

500 mg/kg bw/day

Dose / conc.:

1 000 mg/kg bw/day

No. of animals per sex per dose:

5 animals

Control animals:

yes, concurrent vehicle

Positive control(s):

cyclophosphamide
- Justification for choice of positive control(s): known mutagen
- Route of administration: intraperitoneally (i.p.)
- Doses / concentrations: 40 mg/kg bw in a 0.01 ml/kg bw volume

Tissues and cell types examined:

Determination of chromosome aberrations in metaphases and mitotic index in cells.

Details of tissue and slide preparation:

CRITERIA FOR DOSE SELECTION: based on acute toxicity test

TREATMENT AND SAMPLING TIMES ( in addition to information in specific fields):
- analysis was performed at 18 and 24 h


DETAILS OF SLIDE PREPARATION:
- 3 slides for each animal were generated using the flame-dried technique

METHOD OF ANALYSIS:
- 500 well spread metaphases were selected to assess the presence of CAs, and 1000 or more cells were examined at both sampling times to determine the mitotic index (MI)

Evaluation criteria:

CAs were identified using the established criteria of OECD guideline 475 (version 21 JUL 1997).

Statistics:

- results were expressed as means ± standard deviation (S.D.)
- statistically significant changes were analyzed using one-way ANOVA
- Multiple comparisons were performed using Dunnett’s test
- statistical significance for all tests was set at a p-value below 0.05
- for calculations Graph Pad Instat 3 software for Windows was used

Sex:

female

Genotoxicity:

negative

Toxicity:

no effects

Vehicle controls validity:

valid

Negative controls validity:

not specified

Positive controls validity:

valid

Additional information on results:

Treatment with manganese dioxide did not show an increase (p > 0.05) in the total cytogenetic and structural (gaps, breaks, minute, acentric fragment and reciprocal translocation) changes, numerical (aneuploidy and polyploidy) CAs and percentage (%) of aberrant cells at all doses and treatment times. None of the treatment groups of test item showed significant differences in the mitotic index compared with control groups.

Conclusions:

The chromosome aberration assay of manganese dioxide did not show a statistically significant increase in total cytogenetic and structural changes, numerical CAs and percentage of aberrant cells at all doses and treatment times. Additionally, the mitotic index was not increased by manganese dioxide compared to control groups, indicating no bone marrow toxicity and genotoxicity.

Executive summary:

In a bone marrow chromosome aberration assay comparable to OECD guideline 475, the test item was administered to female rats (5 per dose) orally by gavage at various doses (0, 100, 500, 1000 mg/kg bw), bone marrow cells extracted from the femurs and tibia were collected 18 and 24 h after treatment. For each animal three slides were prepared and assessed for the presence of CAs (500 metaphases investigated) and the mitotic index was determined after analysing more than 1000 cells. Both controls, negative vehicle and positive (cyclophosphamide), were valid. No statistically significant increase in CAs or the mitotic index compared to control groups was observed, resulting in the absence of genotoxicity and bone marrow toxicity.

Data source

Reference

Reference Type:

publication

Title:

Unnamed

Year:

2013

Report date:

2013

Materials and methods

Test guideline

Qualifier:

according to guideline

Guideline:

OECD Guideline 420 (Acute Oral Toxicity - Fixed Dose Method)

Version / remarks:

17 DEC 2001

Deviations:

no

GLP compliance:

not specified

Remarks:

no further information provided in the publication

Test type:

fixed dose procedure

Limit test:

yes

Test material

Test material information

Constituent 1

Reference substance name:

Manganese dioxide

EC Number:

215-202-6

EC Name:

Manganese dioxide

Cas Number:

1313-13-9

Molecular formula:

MnO2

IUPAC Name:

Manganese dioxide

Test material form:

solid

Specific details on test material used for the study:

SOURCE OF TEST MATERIAL
- Source and lot/batch No.of test material: Sigma Chemical Co., St. Louis, USA
- Expiration date of the lot/batch: not specified
- Purity: ≥99%

STABILITY AND STORAGE CONDITIONS OF TEST MATERIAL
- Storage condition of test material: not specified
- Stability under test conditions: not specified
- Solubility and stability of the test substance in the solvent/vehicle: suspended in Milli Q water
- Reactivity of the test substance with the solvent/vehicle of the cell culture medium: not specified

TREATMENT OF TEST MATERIAL PRIOR TO TESTING
- Treatment of test material prior to testing: not specified, but a more detailed description was found for the genotoxicity test assay and was as followed: suspended in Milli Q water and properly ultrasonicated (UPH 100, Germany) for 10 min at 90% amplitude
- Preliminary purification step (if any): no
- Final dilution of a dissolved solid, stock liquid or gel: not specified
- Final preparation of a solid: no

FORM AS APPLIED IN THE TEST (if different from that of starting material): as a suspension

OTHER SPECIFICS:
- Size: mean= 2.74 ± 29 µm
- specific surface area analysis was determined by using the Brunauer–Emmett–Teller technique = 7.95 (m2 /g)

Test animals

Species:

rat

Strain:

Wistar

Remarks:

Albino

Sex:

female

Details on test animals or test system and environmental conditions:

TEST ANIMALS
- Source: National Institute of Nutrition, Hyderabad, India
- Females (if applicable) nulliparous and non-pregnant: not specified
- Age at delivery: 6 to 8 weeks
- Weight at delivery: 80–120 g
- Fasting period before study: not specified
- Housing: in groups of five in polypropylene cages
- Diet: commercial pellet diet
- Water: ad libitum
- Acclimation period: one week

ENVIRONMENTAL CONDITIONS
- Temperature (°C): 22 ± 3
- Humidity (%): 55–65
- Air changes (per hr): not specified
- Photoperiod (hrs dark / hrs light): 12 /12

Administration / exposure

Route of administration:

oral: gavage

Vehicle:

water

Details on oral exposure:

- suspended in Milli Q water and then applied
- No information on vehicle volume for dose groups, but control group received 5 ml/kg bw of Milli Q water.

Doses:

fixed dose: 2000 mg/kg bw

No. of animals per sex per dose:

5 female rats

Control animals:

yes

Details on study design:

- Duration of observation period following administration: 14 days
- Frequency of observations and weighing: not specified, as shown in a graphic for food intake: 1, 3, 7, 10, and 14 d, for body weight: 0, 3, 7, 10, and 14 d after oral administration
- Necropsy of survivors performed: yes
- Other examinations performed: clinical signs: yes
body weight: yes
organ weights: yes
histopathology: yes

Statistics:

The results were expressed as means ± standard deviation (S.D.). Statistically significant changes were analyzed using one-way ANOVA. Multiple comparisons were performed using Dunnett’s test and statistical significance for all tests was set at p < 0.05. All calculations were performed using Graph Pad Instat 3 software for Windows.

Results and discussion

Preliminary study:

In an observational study, no mortality or toxic symptoms were seen at any dose levels (5, 50, 300, and 2000 mg/kg bw) administered.

Effect levels

Key result

Sex:

female

Dose descriptor:

LD50

Effect level:

> 2 000 mg/kg bw

Based on:

test mat.

Mortality:

There was no case of mortality observed.

Clinical signs:

No adverse signs or symptoms were observed.

Body weight:

No significant changes were observed in food consumption or body weight.

Gross pathology:

The test item did not induce treatment-related effects.

Other findings:

- Organ weights: relative weights of liver, kidney, brain, heart and spleen did not significantly change.
- Histopathology: At the highest dose (2000 mg/kg bw) spleen, brain, and liver tissue of rats revealed lesions: spleen showed congestion, brain displayed inflammation, and the liver depicted a dilated central vein. However, rats exposed to 5, 50, 300 and 2000 mg/kg bw of test item showed normal architecture of the spleen, brain, liver, heart and kidney tissues.

Applicant's summary and conclusion

Interpretation of results:

GHS criteria not met

Conclusions:

Single oral administration of the limit dose of 2000 mg test item per kg bw did not cause lethality in 5 female albino Wistar rats during the 14 day observation period, resulting in a LD50 > 2000 mg/kg bw.

Executive summary:

To evaluate the acute toxicity after a single oral administration according to OECD Guideline 420 (acute oral toxicity-fixed dose method), the test item was administered to female albino Wistar rats (5 animals/group) at doses of 0 and 2000 mg/kg bw. Mortality, clinical signs, body weight changes, food consumption, gross findings and relative organ weights (liver, kidney, brain, heart, and spleen) were screened for 14 days following the single dose. No treatment-related effects on mortality, clinical signs, body weight changes, food consumption, gross findings, and relative organ weights in Wistar rats treated with a single oral dose of test item were observed. Therefore, the LD50 value of the test item was considered to be over 2000 mg/kg bw for female rats.