1-Octadecanaminium,N,N-Dimethyl-N-Octadecyl-,(Sp-4-2)-[29h,31h-Phthalocyanine2-Sulfonato(3-)-N 29,N 30,N 31,N 32]Cuprate(1-)

Genetic toxicity: in vivo

Administrative data

Endpoint:

in vivo mammalian somatic cell study: cytogenicity / erythrocyte micronucleus

Remarks:

Type of genotoxicity: chromosome aberration

Type of information:

experimental study

Adequacy of study:

key study

Study period:

1989

Reliability:

1 (reliable without restriction)

Rationale for reliability incl. deficiencies:

comparable to guideline study

Data source

Reference

Reference Type:

study report

Title:

Unnamed

Year:

1989

Report date:

1989

Materials and methods

Test guideline

Test guideline 1

Qualifier:

according to guideline

Guideline:

OECD Guideline 474 (Mammalian Erythrocyte Micronucleus Test)

Deviations:

no

Test guideline 2

Qualifier:

according to guideline

Guideline:

EU Method B.12 (Mutagenicity - In Vivo Mammalian Erythrocyte Micronucleus Test)

Deviations:

no

GLP compliance:

yes

Type of assay:

micronucleus assay

Test material

Test material information

Constituent 1

Details on test material:

- Lot/batch No.: Mix 51785
- Stability under test conditions: Stable under storage conditions
- Storage condition of test material: At room temperature in the dark

Test animals

Species:

mouse

Strain:

Swiss

Sex:

male/female

Details on test animals or test system and environmental conditions:

TEST ANIMALS
- Source: Charles River Wiga, Sulzfeld, F.R.G.
- Age at study initiation: approximately 8 weeks
- Weight at study initiation: 27-36 g (males) and 20-29g (females)
- Assigned to test groups randomly: yes
-Fasting period before study: Withheld overnight before dosing until about 4 hours after administration of the test substance
- Housing: polycarbonate cages
- Diet (e.g. ad libitum): free access to standard laboratory animal diet
- Water (e.g. ad libitum):free access to tap-water
- Acclimation period: 13 days
ENVIRONMENTAL CONDITIONS
- Temperature (°C): 21 +/- 2C
- Humidity (%): 63-65%
- Photoperiod (hrs dark / hrs light): 12 hours light/ 12 hours dark
IN-LIFE DATES: From: January 30, 1989 To: March 15, 1989

Administration / exposure

Route of administration:

oral: gavage

Vehicle:

- Vehicle(s)/solvent(s) used: corn oil
- Concentration of test material in vehicle: 3000 mg/kg

Duration of treatment / exposure:

Bone marrow was sampled 24, 48, and 72 hours after dosing

Frequency of treatment:

once

Post exposure period:

Bone marrow was sampled 24, 48, and 72 hours after dosing

Doses / concentrations

Remarks:

Doses / Concentrations:

Basis:
nominal conc.

No. of animals per sex per dose:

3000 mg/kg -Five males and five males at 24 hour sacrifice time
3000 mg/kg -Five males and five males at 48 hour sacrifice time
3000 mg/kg -Five males and five males at 72 hour sacrifice time

Control animals:

yes, concurrent vehicle

Positive control(s):

cyclophosphamide
-Route of administration: oral
-Doses / concentrations: 50 mg/kg body weight

Examinations

Tissues and cell types examined:

Bone marrow

Details of tissue and slide preparation:

CRITERIA FOR DOSE SELECTION: 3000 mg/kg was chosen as the appropriate dose for this assay as it was the highest dose that could be achieved because of aggregation of the test substance in suspension and it did not show adverse clinical.
CRITERIA FOR DOSE SELECTION: 3000 mg/kg was chosen as the appropriate dose for this assay as it was the highest dose that could be achieved because of aggregation of the test substance in suspension and it did not show adverse clinical.
TREATMENT AND SAMPLING TIMES ( in addition to information in specific fields): Animals were dosed once and the bone marrow was sampled at either 24, 48 or 72 hours.
DETAILS OF SLIDE PREPARATION: A drop of the bone marrow suspension was added to the slides. The drop was spread by moving a clean slide with the drop of bone marrow suspension, The preparations were air dried and fixed for 5 minutes in 100% methanol and air-dried overnight. The slides were stained for 3 min. in undiluted May-Gruwald solution following 2 min in May-Gruwald solution diluted 1:1 with Sorensen buffer pH 6.8. The slides were then rinsed in this buffer and stained for 25 min. in 5% (v/v) Giemsa solution in Sorensen buffer pH 6.8. The preparations were rinsed for 1 min in running tap-water and blotted dry between filter paper. The dry slides were cleared by dipping them in xylol before they were embedded in DePeX and mounted with a coverslip.
METHOD OF ANALYSIS: The slides were first screened at a magnification of 1000x for regions of suitable technical quality. The slides were then scored at a magnification of 1000x. The number of micronuclei was counted in 1000 polychromatic erythrocytes. The ratio polychromatic to normochromatic erythrocytes was determined by counting and differentiating the first 1000 erythrocytes at the same time. Micronuclei were counted in polychromatic erythrocytes only.


Wilcoxon rank-sum to assess the significant differences between the numbers of micronuclei in the treatment and control groups.

Evaluation criteria:

Wilcoxon rank-sum to assess the significant differences between the numbers of micronuclei in the treatment and control groups.

Results and discussion

Test results

Sex:

male/female

Genotoxicity:

negative

Toxicity:

no effects

Vehicle controls validity:

valid

Negative controls validity:

not applicable

Positive controls validity:

valid

Additional information on results:

RESULTS OF DEFINITIVE STUDY
- Ratio of PCE/NCE (for Micronucleus assay): The ratios for males was 1.13, 0.97, and 1.19 for 24, 48, and 72 hours respectively and for females was 1.11, 0.92, 1.02 for 24, 48, and 72 hours respectively.
- Appropriateness of dose levels and route: The oral route was selected taking into account the possible route of human exposure during manufacture, handling and use. 3000 mg/kg was chosen as the appropriate dose for this assay as it was the highest dose that could be achieved because of aggregation of the test substance in suspension and it did not show adverse clinical.

Any other information on results incl. tables

The test material caused no increases in the frequency of micronuclei. The incidence of micronuclei in the control animals were within the historical control ranges. The groups treated with the positive control showed a decrease in the ratio of polychromatic to normochromatic erythrocytes, which reflects a toxic effect of this compound on the erythrocytes. The positive control substance induced in both sexes a statistically significant increase in the number of micronuclei

Sex (M/F)	Group	Treatment	Dose (mg/kg body weight)	Sampling time (hours)	Number of micronuclei per 1000 polychromatic erythrocytes (mean +/- S.D.)a	Ratio polychromatic/normochromatice erythrocytes (mean +/- S.D.)
M	A	Vehicle b)	--	24	0.4 +/- 0.5	1.10 +/- 0.16
M	B	Vehicle	--	48	1.0 +/- 1.0	0.97 +/- 0.08
M	C	Vehicle	--	72	0.2 +/- 0.4	1.15 +/- 0.09
M	D	Test material	3000	24	0.6 +/- 0.5	1.13 +/- 0.18
M	E	Test material	3000	48	0.8 +/- 1.3	0.97 +/- 0.16
M	F	Test material	3000	72	1.2 +/- 1.1	1.19 +/- 0.17
M	G	CP	50	48	14.2 +/- 8.5*	0.33 +/- 0.14
F	A	Vehicle b)	--	24	0.2 +/- 0.4	1.06 +/- 0.17
F	B	Vehicle	--	48	0.4 +/- 0.5	1.08 +/- 0.12
F	C	Vehicle	--	72	0.2 +/- 0.4	1.15 +/- 0.12
F	D	Test material	3000	24	0.0 +/- 0	1.11 +/- 0.10
F	E	Test material	3000	48	0.0 +/- 0	0.92 +/- 0.21
F	F	Test material	3000	72	1.16 +/- 1.1	1.02 +/- 0.10
F	G	CP	50	48	8.6 +/- 2.1*	0.37 +/-0.10

a) Five animals per treatment group

b) Corn oil

c) Significantly different from corresponding control group (Wilcoxon rank sum test, P <=0.05)

    

Applicant's summary and conclusion

Conclusions:

Interpretation of results (migrated information): negative
The test material was non-mutagenic under the conditins of the assay.

Executive summary:

The test material did not show any evidence of causing chromosomal damage or bone marrow toxicity when administered orally by gavage in this in vivo assay.