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Toxicological information

Genetic toxicity: in vivo

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Administrative data

Endpoint:
genetic toxicity in vivo
Remarks:
Type of genotoxicity: chromosome aberration
Type of information:
experimental study
Adequacy of study:
key study
Reliability:
2 (reliable with restrictions)
Rationale for reliability incl. deficiencies:
other: Data is from peer reviewed journal

Data source

Reference
Reference Type:
publication
Title:
Mutagenic Screening of Marker Grenade Dyes by the Salmonella Reversion Assay, L5178Y/TK+/- Mouse Lymphoma Assay, and In Vivo Sister Chromatid Exchange Analysis in Mice
Author:
Martha M. Moore, James W. Allen, Larry Claxton, Carolyn Doerr, Carolyn Gwaltney, John S. Dutcher, Michael Kohan, B. Kay Lawrence, Ruth Templeton, and Barbara Westbrook-Collins
Year:
1988
Bibliographic source:
Environmental and Molecular Mutagenesis 12:219-233 (1988)

Materials and methods

Test guideline
Qualifier:
according to guideline
Guideline:
other: Data is from Journal with permission
Principles of method if other than guideline:
In Vivo Sister Chromatid Exchange Assay was performed to evaluate the SCE frequencies and cell replication kinetics was analyzed in mouse bone marrow cells.
GLP compliance:
not specified
Type of assay:
sister chromatid exchange assay

Test material

Constituent 1
Chemical structure
Reference substance name:
1,3-isobenzofurandione, reaction products with methylquinoline and quinoline
EC Number:
232-318-2
EC Name:
1,3-isobenzofurandione, reaction products with methylquinoline and quinoline
Cas Number:
8003-22-3
Molecular formula:
C18H11NO2
IUPAC Name:
1,3-isobenzofurandione, reaction products with methylquinoline and quinoline
Test material form:
other: Solid
Details on test material:
- Name of test material (as cited in study report): Solvent Yellow 33 2-(2-Quinolyl)-1,3-indandione
- Molecular formula (if other than submission substance): C18H11NO2
- Molecular weight (if other than submission substance): 273.29
- Substance type: Organic
- Physical state: Solid
Purity :No data available
- Impurities (identity and concentrations): No data available

Test animals

Species:
mouse
Strain:
other: C57B1/6J mice
Sex:
male
Details on test animals or test system and environmental conditions:
TEST ANIMALS
- Source: Jackson Laboratory, Bar Harbor, ME
- Age at study initiation: 3-4 months old - Weight at study initiation: No data available
- Fasting period before study: No data available
- Housing: five per cage in a U.S. EPA animal facility in laminar-flow rooms
- Diet - ad libitum chow (noncertified Purina Water (e.g. ad libitum): ad libitum
- Acclimation period: at least 10 days

ENVIRONMENTAL CONDITIONS
- Temperature (°C): 20-21°C
- Humidity (%): 60-68% relative humidity
- Air changes (per hr): 15 cycles/hr of biocleaned air
- Photoperiod (hrs dark / hrs light): 12-hr light-dark cycle.

Administration / exposure

Route of administration:
intraperitoneal
Vehicle:
- Vehicle(s)/solvent(s) used: DMSO
- Justification for choice of solvent/vehicle: Higher volumes of DMSO were determined in preliminary experiments
to be toxic, as evidenced by animal death or inhibited marrow cell cycling
- Amount of vehicle (if gavage or dermal): 0.1mL
Details on exposure:
No details available.
Duration of treatment / exposure:
Duration of exposure: 23 hrs
Frequency of treatment:
Once
Post exposure period:
3-4 hrs
Doses / concentrations
Remarks:
Doses / Concentrations:
5, 15, 25, 35mg/kg
Basis:
no data
No. of animals per sex per dose:
4/sex/dose
Control animals:
yes, concurrent vehicle
Positive control(s):
Cyclophosphamide
- Route of administration: Intraperitoneal
- Doses / concentrations: 30 mg/Kg

Examinations

Tissues and cell types examined:
Marrow cells, second division metaphase cells
Details of tissue and slide preparation:
Details of tissue and slide preparation

DETAILS OF SLIDE PREPARATION: Marrow cells were harvested and processed through hypotonic (0.075 M KCl) and fixative (3:1 methano1:glacial acetic acid) steps, and slides were prepared in accordance with standard cytogenetic methodology. Chromatid differential staining was achieved with the fluorescence-plus-Giemsa (FPG) technique.

METHOD OF ANALYSIS: For each mouse, SCE frequencies were analyzed in 30 randomly selected, well-differentiated second-division metaphase cells that contained the diploid ±2 chromosomal complement.

Cell replication kinetics were also assessed in 200 marrow cell/animal, revealing frequencies of first- (M1), second- (M2), and third- (M3) division stain patterns. A replicative index (RI) was calculated for each animal with the formula RI = (1 X M1) + (2 X M2) + (3 x M3)
Evaluation criteria:
Bone-marrow-cell SCE frequencies and cell kinetics were analyzed. An increase in the SCE frequency was evaluated.

Additional studies were performed to determine
1) If the injected test dye was dispersing or remaining localized within the peritoneum and
2) If higher marrow cell SCE frequencies would result from giving injections over 3 consecutive days.
Statistics:
No details

Results and discussion

Test results
Sex:
male
Genotoxicity:
negative
Toxicity:
no effects
Remarks:
negative
Vehicle controls validity:
valid
Negative controls validity:
valid
Positive controls validity:
valid
Additional information on results:
Slight evidence of sister chromatid exchange but not mentioned in details.

Any other information on results incl. tables

Genotoxicity:

Negative

Treatment

No. of Animals

SCE/cell

Mean ± SD

RI

Negative Control

4

4.0 ± 2.01

1.91 ± 0.10

Solvent Control

4

4.5 ± 1.91

2.00 ± 0.14

Positive control

4

52.4 ± 10.92

1.58 ± 0.27

5 mg/Kg

4

4.5 ± 2.49

2.14 ± 0.13

15 mg/Kg

3

4.0 ± 2.14

2.31 ± 0.20

25 mg/Kg

3

3.2 ± 2.00

2.24 ± 0.22

35 mg/Kg

4

3.3 ± 2.13

2.54 ± 0.27

Applicant's summary and conclusion

Conclusions:
Interpretation of results (migrated information): negative
The given dye C.I. Solvent Yellow 33 failed to increase the SCE frequencies and hence if negative for gene mutation in vivo.
Executive summary:

In Vivo Sister Chromatid Exchange Analysis was performed using Male C57B1/6J mice bobe marrow cells. SCE induction was determined by administering the test chemical as a single i.p. injection (≤ 0.2 ml volume) over a 3-4 point dose range, four mice per dose.

The dye injections were given 1/2 hr after BrdUrd tablet implantation. Negative control animals received no injections or were injected with only the solvent. Positive control mice were injected with 30 mg/kg cyclophosphamide.

Approximately 23 hr later, all control mice were injected i.p. with 0.6 mg/kg of colchicine to collect metaphases. Treated mice were injected with colchicine after an additional 3-4 hr because preliminary chemical injection trials had indicated that cell-cycle delays were occurring. Two hours after colchicine injection, animals were sacrificed by cervical dislocation, marrow cells were harvested and processed through hypotonic (0.075 M KCl) and fixative (3:1 methanol: glacial acetic acid) steps, and slides were prepared in accordance with standard cytogenetic methodology.

SCE levels were not significantly different from control levels and higherRIsprobably were a reflection of later cell harvest times. The dye-treated animals generally showed no greater SCE frequencies or cytotoxicity than controls.

Dye coloration or evidence of dye crystals was not apparent in the peritoneum of animals dissected at the time of marrow cell harvest. There were also no crystals of dye evident in peritoneal cell pellets examined under a microscope. The percentage of cells represented by macrophages also was comparable between negative control and treated mice (83-84%). No traces of the dyes were observed in the marrow cell preparations.

The i.p. injected dyes are distributed to marrow cells and are inactive for SCE induction and hence is negative for gene mutation in vivo.

According to the CLP regulation, the given test substance is negative for gene mutation.