2-hydroxy-5-octanoylbenzoic acid

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:

No data

Reliability:

2 (reliable with restrictions)

Rationale for reliability incl. deficiencies:

other: GLP study similar to OECD guideline 474 with minor deviations: data about test substance purity, no certificate of analysis, animal bodyweight not followed, standard deviations not reported in the results.

Cross-reference

Cross-reference 1

Reason / purpose for cross-reference:

reference to same study

Cross-reference 2

Reason / purpose for cross-reference:

reference to other study

Data source

Reference

Reference Type:

study report

Title:

Unnamed

Year:

1994

Report date:

1994

Materials and methods

Test guideline

Qualifier:

equivalent or similar to guideline

Guideline:

OECD Guideline 474 (Mammalian Erythrocyte Micronucleus Test)

Deviations:

yes

Remarks:

no data about test substance purity; no certificate of analysis; animal bodyweight not followed; standard deviations not reported in the results

Principles of method if other than guideline:

Not applicable

GLP compliance:

yes

Type of assay:

micronucleus assay

Test material

Test material information

Constituent 1

Reference substance name:

-

EC Number:

417-790-1

EC Name:

-

Cas Number:

78418-01-6

Molecular formula:

C15H20O4

IUPAC Name:

2-Hydroxy-5-(1-oxooctyl)benzoic acid

Constituent 2

Reference substance name:

MEXORYL SAB

IUPAC Name:

MEXORYL SAB

Details on test material:

- Name of test material (as cited in study report): Mexoryl SAB
- Physical state: white cristalline powder
- Analytical purity: no data
- Lot/batch No.: op T 4 E

Test animals

Species:

mouse

Strain:

CD-1

Sex:

male/female

Details on test animals or test system and environmental conditions:

TEST ANIMALS
- Source: Charles River Limited, Margate, U.K.
- Fasting period before study: no
- Housing: in groups of 5 per cage
- Diet (e.g. ad libitum): SQC rat and mouse maintenance diet no. 1 (Special Diets Services Limited, Witham, England), ad libitum
- Water (e.g. ad libitum): tap water ad libitum

ENVIRONMENTAL CONDITIONS
- Temperature (°C): 22 ± 3
- Humidity (%): 50 ± 20
- Photoperiod (hrs dark / hrs light): 12 h/12 h

Administration / exposure

Route of administration:

oral: gavage

Vehicle:

- Vehicle(s)/solvent(s) used: test substance: carboxymethylcellulose (CMC); positive control: 0.9% sterile saline
- Concentration of test material in vehicle: test substance: 0.5%; positive control: 0.4 mg/mL

Details on exposure:

No data

Duration of treatment / exposure:

Single administration

Frequency of treatment:

Single administration

Post exposure period:

Range finding study: 3 days
Main study: 24, 48 and 72 h

Doses / concentrations

Doses / concentrations 1

Remarks:

Doses / Concentrations:
250 mg/kg bw
Basis:
actual ingested

Doses / concentrations 2

Remarks:

Doses / Concentrations:
500 mg/kg bw
Basis:
actual ingested

Doses / concentrations 3

Remarks:

Doses / Concentrations:
1000 mg/kg bw
Basis:
actual ingested

No. of animals per sex per dose:

5

Control animals:

yes, concurrent vehicle

Positive control(s):

Mitomycin C
- Route of administration: intraperitoneal injection
- Doses / concentrations: 4 mg/kg

Examinations

Tissues and cell types examined:

Bone marrow

Details of tissue and slide preparation:

CRITERIA FOR DOSE SELECTION: based on the range finding study where animals exposed at 1000 mg/kg (highest dose tested) had no critical signs of toxicity

TREATMENT AND SAMPLING TIMES ( in addition to information in specific fields): for each dose group, 5 males and 5 females were sacrified at 24, 48 and 72 h after dosing

DETAILS OF SLIDE PREPARATION:
Marrow cells were centrifuged at 800 rpm for 5 min, the supernatant withdrawn, and the cells re-suspended in a minimal volume of foetal calf serum. One drop of cell suspension was placed on each of 2 slides and was left to air dry for 24 hours before staining based on the method of Gollapudi and Kamra. Cells were fixed for 5 min in methanol, rinsed twice in deionised water, stained for 10 min in Giemsa and rinsed several times in tap water and finally in deionised water.

METHOD OF ANALYSIS: A minimum of 1000 polychromatic erythrocytes (PCE), including micronucleated PCE, were counted for each animal.

Evaluation criteria:

Based on statistical analysis and the toxicity shown by PCE/NCE ratio.

Statistics:

Analysis of micronucleus counts to determine the significance of differences between control and Mexoryl SAB treated groups was carried out by the Mann-Whitney U test.

Results and discussion

Test results

Sex:

male/female

Genotoxicity:

negative

Toxicity:

yes

Remarks:

Decrease of PCE/NCE ratio for 1000 mg/kg at 24 h

Vehicle controls validity:

valid

Negative controls validity:

not applicable

Positive controls validity:

valid

Additional information on results:

RESULTS OF RANGE-FINDING STUDY
- Dose range: 50, 320 and 1000 mg/kg
- Clinical signs of toxicity in test animals: no severe toxic events
- Evidence of cytotoxicity in tissue analyzed: not examined

RESULTS OF DEFINITIVE STUDY
- Induction of micronuclei (for Micronucleus assay): after 24 h at 1000 mg/kg
- Ratio of PCE/NCE (for Micronucleus assay): 0.52 (males and females) at 24 h
- Appropriateness of dose levels and route: acceptable
- Statistical evaluation: Mann-Whitney U test

Any other information on results incl. tables

Table 1: group means for animals sacrificed at 24 h

		Mean	Mean	Mean	Ratio	Mean %
Group		PCE	MN-PCE	MN-NCE	PCE/NCE	MN-PCE
CONTROL	M	1031.4	0.0	0.0	0.83	0.00
CONTROL	F	1004.2	0.0	0.0	0.88	0.00
CONTROL	M+F	1017.8	0.0	0.0	0.86	0.00
250 mg/kg	M	1018.4	0.0	0.0	1.06	0.00
250 mg/kg	F	1047.2	0.2	0.0	1.26	0.02
250 mg/kg	M+F	1032.8	0.1	0.0	1.16	0.01
500 mg/kg	M	1027.8	0.0	0.0	1.08	0.00
500 mg/kg	F	1023.2	0.0	0.0	1.14	0.00
500 mg/kg	M+F	1025.5	0.0	0.0	1.11	0.00
1000 mg/kg	M	1018.2	0.0	0.2	0.60	0.00
1000 mg/kg	F	1036.0	0.2	0.0	0.44	0.02
1000 mg/kg	M+F	1027.1	0.1	0.1	0.52	0.01
Mitomycin C	M	1024.2	8.4	0.2	1.23	0.82
Mitomycin C	F	1064.8	12.6	0.6	1.22	1.19
Mitomycin C	M+F	1044.5	10.5	0.4	1.23	1.01

Table 2: group means for animals sacrificed at 48 h

		Mean	Mean	Mean	Ratio	Mean %
Group		PCE	MN-PCE	MN-NCE	PCE/NCE	MN-PCE
CONTROL	M	1006.2	0.0	0.0	1.26	0.00
CONTROL	F	1018.4	0.0	0.0	1.44	0.00
CONTROL	M+F	1012.3	0.0	0.0	1.35	0.00
250 mg/kg	M	1010.8	0.0	0.0	0.97	0.00
250 mg/kg	F	1034.0	0.0	0.0	0.90	0.00
250 mg/kg	M+F	1022.4	0.0	0.0	0.93	0.00
500 mg/kg	M	1011.4	0.0	0.0	1.05	0.00
500 mg/kg	F	1053.0	0.0	0.0	1.06	0.00
500 mg/kg	M+F	1032.2	0.0	0.0	1.06	0.00
1000 mg/kg	M	1041.8	0.0	0.0	1.14	0.00
1000 mg/kg	F	1046.6	0.0	0.0	1.27	0.00
1000 mg/kg	M+F	1044.4	0.0	0.0	1.21	0.00

Table 3: group means for animals sacrificed at 72 h

		Mean	Mean	Mean	Ratio	Mean %
Group		PCE	MN-PCE	MN-NCE	PCE/NCE	MN-PCE
CONTROL	M	1006.2	0.0	0.0	1.26	0.00
CONTROL	F	1018.4	0.0	0.0	1.44	0.00
CONTROL	M+F	1012.3	0.0	0.0	1.35	0.00
250 mg/kg	M	1010.8	0.0	0.0	0.97	0.00
250 mg/kg	F	1034.0	0.0	0.0	0.90	0.00
250 mg/kg	M+F	1022.4	0.0	0.0	0.93	0.00
500 mg/kg	M	1011.4	0.0	0.0	1.05	0.00
500 mg/kg	F	1053.0	0.0	0.0	1.06	0.00
500 mg/kg	M+F	1032.2	0.0	0.0	1.06	0.00
1000 mg/kg	M	1041.8	0.0	0.0	1.14	0.00
1000 mg/kg	F	1046.6	0.0	0.0	1.27	0.00
1000 mg/kg	M+F	1044.4	0.0	0.0	1.21	0.00

Applicant's summary and conclusion

Conclusions:

Interpretation of results (migrated information): negative
Under these test conditions, Mexoryl SAB was found not to be clastogenic in vivo on bone marrow cells of mice.

Executive summary:

In an in vivo micronucleus test similar to OECD guideline 474 and in compliance with GLP, mice were exposed to Mexoryl SAB. In a range finding assay, pairs of male mice were orally exposed to Mexoryl SAB at concentrations of 50, 320 and 1000 mg/kg without any critical sign of toxicity. Then, 5 animals/sex/dose/time of sacrifice were given single dose of 250, 500 and 1000 mg/kg of Mexoryl SAB by oral gavage and were sacrificed at 24, 48 and 72 h after dosing. Then bone marrow cells were harvested, stained with Giemsa and analysed for micronuclei.

Positive controls (mitomycin C at 4 mg/kg intraperitoneally) induced an appropriate increase in the number of polychromatic erythrocytes. Micronuclei were not induced at any tested concentrations and at any time of sacrifice in control and in Meroxyl SAB-treated mice despite a decrease in polychromatic erythrocytes/normochromatic erythrocytes for 1000 mg/kg at 24 h.

Under the test conditions, Mexoryl SAB was found not to be clastogenic in vivo in mice.