Octane-1-thiol

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:

migrated information: read-across from supporting substance (structural analogue or surrogate)

Adequacy of study:

key study

Study period:

1995-10-05

Reliability:

1 (reliable without restriction)

Rationale for reliability incl. deficiencies:

other: This study is classified as reliable without restriction because it is well-documented and followed OECD Guideline 474.

Data source

Reference

Reference Type:

study report

Title:

Unnamed

Year:

1995

Report date:

1995

Materials and methods

Test guideline

Qualifier:

according to guideline

Guideline:

OECD Guideline 474 (Mammalian Erythrocyte Micronucleus Test)

Deviations:

yes

Remarks:

animal housing temperature was higher than the recommended range for over 4 hours but the study director determined this not to have an adverse affect on study results

GLP compliance:

yes (incl. QA statement)

Remarks:

U.S. Food and Drug Administration, U.S. Environmental Protection Agency, UK GLP Compliance Programme, Japanese GLP Standard, and OECD Principles of GLP

Type of assay:

micronucleus assay

Test material

Test material information

Constituent 1

Reference substance name:

Dodecane-1-thiol (CAS # 112-55-0)

IUPAC Name:

Dodecane-1-thiol (CAS # 112-55-0)

Details on test material:

- Name of test material (as cited in study report): n-dodecyl mercaptan (dodecane-1-thiol)
- Substance type: clear, colorless
- Physical state: liquid
- Analytical purity: 99.5% (provided by sponsor)
- Composition of test material, percentage of components:
- Lot/batch No.: 3000NDM95
- Expiration Date: 1996-05-01
- Storage condition of test material: room temperature, protected from exposure to light
- Test Article Receipt Date: 1995-05-12
- MA Study No.: G95AT23.122

Test animals

Species:

mouse

Strain:

ICR

Sex:

male/female

Details on test animals or test system and environmental conditions:

TEST ANIMALS
- Source: Harlan Sprague Dawley, Inc., Frederick, MD
- Age at study initiation: 6 to 8 weeks
- Weight at study initiation: Pilot study: Male 29.0 to 38.1 grams Female 26.4 to 30.1 grams; Micronucleus Assay: Male 27.4 to 35.8 grams Female 24.5 to 30.6 grams
- Assigned to test groups randomly: yes
- Fasting period before study: Not reported
- Housing: AAALAC-accredited facility, same sex five per cage in plastic autoclavable cages with filter tops; heat-treated hardwood chips were used for bedding.
- Diet (e.g. ad libitum): certified laboratory rodent chow which had be analyzed for environmental contaminants (Purina Certified Rodent Chow 5002) ad libitum
- Water (e.g. ad libitum): tap water, Washington Suburban Sanitary Commission, Potomac Plant ad libitum
- Acclimation period: no less than 5 days


ENVIRONMENTAL CONDITIONS
- Temperature (°C): 20 to 26.7°C
- Humidity (%): 30 to 70 %
- Photoperiod (hrs dark / hrs light): 12/12

Administration / exposure

Route of administration:

oral: gavage

Vehicle:

- Vehicle(s)/solvent(s) used: corn oil
- Justification for choice of solvent/vehicle: the vehicle was selected to be the solvent of choice by the sponsor and the test material was soluble in corn oil at a concentration of 250 mg/mL, the maximum concentration tested.

Details on exposure:

PREPARATION OF DOSING SOLUTIONS:


DIET PREPARATION
- Rate of preparation of diet (frequency): 20 mL/kg bw

Duration of treatment / exposure:

single oral dose

Frequency of treatment:

single oral dose

Post exposure period:

24, 48, and 72 hours

Doses / concentrations

Remarks:

Doses / Concentrations:
1250, 2500, or 5000 mg/kg
Basis:
actual ingested
oral gavage

No. of animals per sex per dose:

5

Control animals:

other: yes, concurrent vehicle and a positive control

Positive control(s):

cyclophosphamide (CP CAS: 6055-19-2)
- Route of administration: oral gavage
- Doses / concentrations: 60 mg/kg

Examinations

Tissues and cell types examined:

bone marrow from the femur, polychromatic erythrocytes

Details of tissue and slide preparation:

CRITERIA FOR DOSE SELECTION: Conducted using established and validated procedures (Heddle, 1973; Heddle et al., 1983; Matter and Grauwiler, 1974)


TREATMENT AND SAMPLING TIMES ( in addition to information in specific fields): The test article-vehicle mixture, the vehicle alone, or the positive control was administered by oral gavage at a constant volume of 20 ml/kg body weight. All mice in the experimental and control groups were weighed immediately prior to dose administration and the dose volume was based on individual body weights. Mice were observed after dose administration for clinical signs of chemical effect.


DETAILS OF SLIDE PREPARATION: Five mice per treatment were sacrificed by CO2 asphyxiation. Immediately following sacrifice, the femurs were exposed, cut just above the knee, and the bone marrow was aspirated into a syringe containing fetal bovine serum. The bone marrow cells were transferred to a capped centrifuge tube containing approximately 1mL fetal bovine serum. The bone marrow cells were pelleted by centrifugation at approximately 100 x g for five minutes and the supernatant was drawn off, leaving a small amount of serum with the remaining cell pellet. The cells were resuspended by aspiration with a capillary pipet and a small drop of bone marrow suspension was spread onto a clean glass slide. Two to four slides were prepared from each mouse. The slides were fixed in methanol, stained with May-Gruenwald-Giemsa and permanently mounted.


METHOD OF ANALYSIS: Slides were coded using a random number table by an individual not involved with the scoring process. Using medium magnification, an area of acceptable quality was selected such that the cells were well spread and stained. Using oil immersion, 1000 polychromatic erythrocytes were scored for the presence of micronuclei which are defined as round, darkly staining nuclear fragments, having a sharp contour with diameters usually from 1/20 to 1/5 of the erythrocyte. The number of micronucleated normochromatic erythrocytes in the field of 1000 polychromatic erythrocytes was enumerated. The proportion of polychromatic erythrocytes to total erythrocytes was also recorded per 1000 erythrocytes.

Evaluation criteria:

In order to quantify the test article effect on erythropoiesis, as an indicator of bone marrow toxicity, the proportion of polychromatic erythrocytes to total erythrocytes was determined for each animal and treatment group.

The test article was considered to induce a positive response if a treatment-related increase in micronucleated polychromatic erythrocytes was observed and one or more doses were statistically elevated relative to the vehicle control (~0.05, Kastenbaum-Bowman Tables) at any sampling time. If a single treatment group was significantly elevated at one sacrifice time with no evidence of a dose-response, the assay was considered a suspect or unconfirmed positive and a repeat assay recommended. The test article was considered negative if no statistically significant increase in micronucleated polychromatic erythrocytes above the concurrent vehicle control was observed at any sampling time.

To confirm a valid test the mean incidence of micronucleated polychromatic erythrocytes must not exceed 5/1000 polychromatic erythrocytes (0.5%) in the vehicle control. The incidence of micronucleated polychromatic erythrocytes in the positive control group must be significantly increased relative to the vehicle control group (p≤0.05, Kastenbaum-Bowman Tables).

Statistics:

Statistical significance was determined for the incidence of micronucleated polychromatic erythrocytes per 1000 polychromatic erythrocytes using the Kastenbaum-Bowman tables which are based on the binomial distribution (Kastenbaum and Bowman, 1970; Mackey and MacGregor, 1979). All analyses were performed separately for each sex and sampling time.

Results and discussion

Test results

Sex:

male/female

Genotoxicity:

negative

Toxicity:

yes

Remarks:

clinical signs; 23% reduction in PCE/total erythrocyte ratio

Vehicle controls validity:

valid

Negative controls validity:

not applicable

Positive controls validity:

valid

Additional information on results:

RESULTS OF PILOT STUDY
- Dose range: 1- 5000 mg/kg bw
- Solubility: test article was soluble in corn oil at highest dose tests of 250 mg/mL
- Clinical signs of toxicity in test animals: lethargy and diarrhea in male and female mice at 5000 mg/kg. All other animals appeared normal.
- Other: Absence of mortality meant no toxicity study was required. Highest dose level for the micronucleus study was set at 5000 mg/kg.


RESULTS OF DEFINITIVE STUDY
- Induction of micronuclei (for Micronucleus assay):
- Ratio of PCE/NCE (for Micronucleus assay): see tables below. Reductions of up to 23% in the ratio of polychromatic erythrocytes to total erythrocytes were observed in some of test article-treated groups relative to their respective vehicle controls.
- Appropriateness of dose levels and route: dose level and route were appropriate based on the guideline and pilot study. The positive control produced a significant increase in micronucleated polychromatic erythrocytes in both male and female mice (p≤0.05)
- Statistical evaluation: not statistically increased relative to their respective vehicle control in either male or female mice, regardless of dose level or bone marrow collection time (p> 0.05, Kastenbaum-Bowman Tables)

Any other information on results incl. tables

Summary of Bone Marrow Micronucleus Study using n-Dodecyl Mercaptan

Treatment	Sex	Time (HR)	Number of Mice	PCE/Total Erythrocytes	Micronucleated Polychromatic Erythrocytes
					Number per 1000 PCE’s (MEAN ± S.D.)	Number Per PCE’s Scored’
Corm Oil 20 mL/kg	M	24	5	0.53	0.8 ± 0.84	4/5000
		48	5	0.58	1.4 ± 1.14	7/5000
		72	5	0.57	0.8 ± 0.84	4/5000
	F	24	5	0.70	2.0 ± 1.00	10/5000
		48	5	0.59	1.0 ± 0.71	5/5000
		72	5	0.61	1.2 ± 0.84	6/5000
n-Dodecyl mercaptan 1250 mg/kg	M	24	5	0.54	1.0 ± 0.71	5/5000
		48	5	0.52	1.6 ± 1.14	8/5000
		72	5	0.64	0.8 ± 0.84	4/5000
	F	24	5	0.69	1.2 ± 1.30	6/5000
		48	5	0.46	0.8 ± 1.30	4/5000
		72	5	0.67	1.6 ± 1.14	8/5000
2500 mg/kg	M	24	5	0.53	1.0 ± 0.71	5/5000
		48	5	0.51	1.0 ± 1.00	5/5000
		72	5	0.61	0.8 ± 0.84	4/5000
	F	24	5	0.63	1.4 ± 0.89	7/5000
		48	5	0.45	0.6 ± 0.55	3/5000
		72	5	0.67	0.8 ± 1.30	4/5000
5000 mg/kg	M	24	5	0.54	0.6 ± 0.55	3/5000
		48	5	0.49	0.4 ± 0.55	2/5000
		72	5	0.51	2.0 ± 1.22	10/5000
	F	24	5	0.66	0.6 ± 0.89	3/5000
		48	5	0.43	1.0 ± 1.00	5/5000
		72	5	0.52	0.8 ± 1.30	4/5000
CP ± 60 mg/kg	M	24	5	0.54	17.4 ± 3.21	87/5000*
	F	24	5	0.46	34.4 ± 2.41	172/5000

* p≤0.05 (Kastenbaum-Bowman Tables)

 

Applicant's summary and conclusion

Conclusions:

Interpretation of results (migrated information): negative
In an ICR mouse bone marrow micronucleus assay, 5 animals per sex per dose were treated orally with dodecane-1-thiol at doses of 0, 1250, 2500, or 5000 mg/kg bw. Bone marrow cells were harvested at 24, 48, and 72 hours post-treatment. The vehicle was corn oil, recommended by the sponsor. The test material was administered by oral gavage as a single dose of 20 mL of the test material per kg body weight.  There was not a significant increase in the frequency of micronucleated polychromatic erythrocytes in bone marrow in either sex or any treatment time.

Executive summary:

In an ICR mouse bone marrow micronucleus assay, 5 animals per sex per dose were treated orally with dodecane-1-thiol at doses of 0, 1250, 2500, or 5000 mg/kg bw. Bone marrow cells were harvested at 24, 48, and 72 hours post-treatment. The vehicle was corn oil, recommended by the sponsor. The test material was administered by oral gavage as a single dose of 20mL of the test material per kg body weight. 

 

Signs of toxicity included lethargy and diarrhea in all dose levels and a male with crusty eyes in the highest dose level. No mortality occurred during the study. Dodecane-1-thiol was tested at an adequate dose based on a pilot study that found no mortality at the 5000 mg/kg dose level. The positive control induced the appropriate response. There was not a significant increase in the frequency of micronucleated polychromatic erythrocytes in bone marrow in either sex or any treatment time.

 

This study received a Klimisch score of 1 and is classified as reliable without restriction because it was well-documented and followed OECD Guideline 474.