Naphtha (Fischer-Tropsch), light, C4-10-branched and linear

Developmental toxicity / teratogenicity

Administrative data

Endpoint:

developmental toxicity

Type of information:

experimental study

Adequacy of study:

key study

Reliability:

1 (reliable without restriction)

Rationale for reliability incl. deficiencies:

comparable to guideline study

Data source

Reference

Reference Type:

publication

Title:

Unnamed

Year:

2001

Materials and methods

Test guideline

Test guideline 1

Qualifier:

according to guideline

Guideline:

EPA OTS 798.4350 (Inhalation Developmental Toxicity Screen)

Deviations:

yes

Remarks:

exposure period was extended to encompass the phase of fetal development

Test guideline 2

Qualifier:

equivalent or similar to guideline

Guideline:

OECD Guideline 414 (Prenatal Developmental Toxicity Study)

Deviations:

not applicable

GLP compliance:

yes

Limit test:

no

Test material

Test material information

Constituent 1

Reference substance name:

unleaded gasoline vapor condensate

IUPAC Name:

unleaded gasoline vapor condensate

Details on test material:

- Name of test material (as cited in study report): unleaded gasoline vapor condensate
- Composition of test material, percentage of components: primarily of volatile short chain (C4-C8) aliphatic hydrocarbons (i.e. paraffins) with small amounts of cycloparaffins and aromatic hydrocarbons [see also below "Any other information on materials and methods incl. tables", Table 1].
- Other: There are likely to be fugitive emissions of the more volatile components of the substance 'Naphtha (Fischer-Tropsch), light, C4-C10 branched and linear' to air. The evaporative emissions of the test material unleaded gasoline are expected to be similar with 'Naphtha (Fischer-Tropsch), light, C4-C10 branched and linear'. Hence, the gasoline vapour condensate is representative for the fraction of 'Naphtha (Fischer-Tropsch), light, C4-C10 branched and linear' to which man would be exposed during processing, handling and use (accidental / infrequent).

Test animals

Species:

rat

Strain:

Sprague-Dawley

Details on test animals or test system and environmental conditions:

TEST ANIMALS
- Source: Charles River Laboratories, Portage, MI
- Age at study initiation: 71 days
- Weight at study initiation: 261-264 g
- Housing: Individually housed in suspended stainless-steel cages with wire mesh floors and fronts
- Diet (e.g. ad libitum): ad libitum
- Water (e.g. ad libitum): ad libitum
- Acclimation period: 14 days

ENVIRONMENTAL CONDITIONS
- Temperature (°C): 20-24
- Humidity (%): 38-74
- Photoperiod (hrs dark / hrs light): 12/12

Administration / exposure

Route of administration:

inhalation: vapour

Type of inhalation exposure (if applicable):

whole body

Vehicle:

other: nitrogen (purity 99.98%)

Details on exposure:

GENERATION OF TEST ATMOSPHERE / CHAMBER DESCRIPTION
- System of generating particulates/aerosols: Charging a glass-lined Pfaulder kettle (closed system) with 6746 lbs of unleaded gasoline. The sample was slowly heated and stirred as the liquid temperature was raised to 150 F, resulting in a vapor temperature of 130 F. The vapor was condensed by passing through a series of two receiving vessels chilled with cold water and dry ice, then through additional vapor traps chilled in dry ice/ isopropyl alcohol. The vapor condensate collected by this method represented 10.4% of the initial sample weight. The chilled condensate was uniformly mixed, transferred to 5-gallon containers, and shipped to the testing laboratory where it was stored at ambient temperature in a solvent storage building until use.
- Exposure apparatus: 1000-L glass and stainless steel exposure chamber (Wahlmann Mfg. Co., Timonium, MD)
- Source and rate of air: Test material was pumped directly from the storage container, refrigerated to minimize volatilization during transfer, into a countercurrent volatilization chamber
- Method of conditioning air: Gasoline vapor laden nitrogen flowed through the top of the volatilization chamber into the turret of the 1000 L exposure chamber, where it mixed with room air to the appropriate exposure concentration as it was drawn into the chamber.
- Temperature and humidity in air chamber: 20-25 °C, 34-70 %; monitored every 1/2 hour
- Air flow rate: 200 L/min
- Air change rate: 5 min
- Method of particle size determination: TSI Aerodynamic Particle Sizer at least once during every exposure
- Treatment of exhaust air: Chambers were exhausted through a system of a coarse filter, a HEPA filter, and a charcoal filter

Analytical verification of doses or concentrations:

yes

Details on analytical verification of doses or concentrations:

One sample per chamber per week was collected by syringe grab sampling and analyzed by gas chromatography (Hewlett Packard 5890II with flame ionization detector) to characterize 12 major components of the test atmosphere to demostrate stability of test vapor over the course of the study. Composition and stability of the test material were evaluated by characterizing the liquid unleaded gasoline vapor condensate and comparing the major components with the generated atmospheres throughout the study.

Details on mating procedure:

- Impregnation procedure: cohoused
- M/F ratio per cage: 1:1
- Length of cohabitation: nightly
- Verification of same strain and source of both sexes: yes
- Proof of pregnancy: vaginal plug and/or sperm in vaginal smear referred to as day 0 of pregnancy

Duration of treatment / exposure:

GD6-GD19

Frequency of treatment:

once daily, 6 h

Duration of test:

through day 20 of gestation

Doses / concentrations

Doses / concentrations 1

Remarks:

Doses / Concentrations:
2653 mg/m3
Basis:
analytical conc.

Doses / concentrations 2

Remarks:

Doses / Concentrations:
7960 mg/m3
Basis:
analytical conc.

Doses / concentrations 3

Remarks:

Doses / Concentrations:
23900 mg/m3
Basis:
analytical conc.

No. of animals per sex per dose:

24 females/dose

Control animals:

yes, sham-exposed

Examinations

Maternal examinations:

CAGE SIDE OBSERVATIONS: Yes
- Time schedule: Twice daily; during days 6 through 19, animals were evaluated pre- and post- exposure when animals were removed from the inhalation chambers.

DETAILED CLINICAL OBSERVATIONS: Yes
- Time schedule: Days 0 and 6 through 20.

BODY WEIGHT: Yes
- Time schedule for examinations: days 0, 3, 6, 9, 12, 15, 18, and 20 of gestation.

POST-MORTEM EXAMINATIONS: Yes
- Sacrifice on gestation day 20
- Organs examined: uterus and ovaries.

FOOD CONSUMPTION AND COMPOUND INTAKE (if feeding study): Yes
- Food consumption for each animal determined and mean daily diet consumption calculated as g food/kg body weight/day: Yes
- Compound intake calculated as time-weighted averages from the consumption and body weight gain data: Yes.

Ovaries and uterine content:

The ovaries and uterine content was examined after termination: Yes
Examinations included:
- Gravid uterus weight: Yes
- Number of corpora lutea: Yes
- Number of implantations: Yes
- Number of early resorptions: Yes
- Number of late resorptions: Yes
- Other: Where no uterine implants were grossly apparent, the uterus was stained with ammonium sulfide to visualize any uterine foci.

Fetal examinations:

- External examinations, weighed and sexed: Yes: [all per litter]
- Soft tissue examinations: Yes: [half per litter]
- Skeletal examinations: Yes: [half per litter]
- Head examinations: Yes: [half per litter]

Statistics:

conducted where appropriate

Indices:

no data

Historical control data:

no data

Results and discussion

Results: maternal animals

Maternal developmental toxicity

Details on maternal toxic effects:

Maternal toxic effects:no effects

Details on maternal toxic effects:
All mated females survived to scheduled sacrifice. Physical examinations performed pre- and post-exposure did not indicate any adverse effect from exposure to unleaded gasoline vapor condensate. Mean maternal body weight and weight gain during gestation were not adversely affected by treatment.

Effect levels (maternal animals)

Dose descriptor:

NOAEL

Effect level:

23 900 mg/m³ air (analytical)

Basis for effect level:

other: maternal toxicity

Results (fetuses)

Details on embryotoxic / teratogenic effects:

Embryotoxic / teratogenic effects:no effects

Details on embryotoxic / teratogenic effects:
Pregnancy rates in treated groups were statistically indistinguishable from the sham treated control group. No adverse effects of treatment were evident from uterine implantation data. There were no aborted pregnancies or premature deliveries in any group. No macroscopic abnormalities related to test material exposre were observed in postmortem examination of maternal animals. No external malformations or variations were recorded among fetuses from control or treated groups.

Effect levels (fetuses)

Dose descriptor:

NOAEL

Effect level:

23 900 mg/m³ air (analytical)

Basis for effect level:

other: fetotoxicity

Fetal abnormalities

Abnormalities:

not specified

Overall developmental toxicity

Developmental effects observed:

not specified

Any other information on results incl. tables

Table: Summary of reproduction and mean fetal weight data

	Exposure Levels (ppm)
		0	1000	3000	9000
	Number of Females Mated	24	24	24	24
	No. Pregnant (%)	24 (100.0)	22 (91.7)	22 (91.7)	21 (87.5)
	No. Pregnancies Aborted	0	0	0	0
	No. Premature Births	0	0	0	0
	No. Litters with Viable Fetuses	24	22	22	21
	Female Mortality No.	0	0	0	0
	Corpora Lutea
	Mean ± SD	16.7 ± 3.9	18.1 ± 2.1	17.3 ± 2.9	17.3 ± 2.0
	Implanation Sites
	Mean ± SD	14.8 ± 4.5	16.1 ± 2.7	16.1 ± 3.1	16.1 ± 2.4
	Preimplantation Loss Index
	Mean ± SD	0.121 ± 0.193	0.104 ± 0.131	0.071 ± 0.118	0.063 ± 0.107
	Number of Viable Fetuses	327	334	336	325
	Number of Dead Fetuses	0	0	0	0
	Mean Litter Size ± SD	13.6 ± 5.0	15.2 ± 2.6	15.3 ± 3.4	15.5 ± 2.3
	Mean No. Males ± SD	7.0 ± 3.2	7.5 ± 2.6	8.0 ± 2.7	7.6 ± 2.7
	Mean No. Female ± SD	6.6 ± 3.0	7.7 ± 2.3	7.3 ± 3.3	7.9 ± 2.0
	Resorptions
	Mean ± SD	1.2 ± 2.7	1.0 ± 0.8	0.8 ± 1.0	0.7 ± 0.7
	Resorptions/Implants Ratio
	Mean ± SD	0.075 ± 0.017	0.058 ± 0.054	0.055 ± 0.063	0.041 ± 0.044
	No. of Litters with Resorptions (%)	10(41.7)	14 (63.6)	11 (50.0)	11 (52.4)
	Mean Body Weight (g) of Viable	3.66 ± 0.33	3.71 ± 0.30	3.68 ± 0.17	3.66 ± 0.32
	Fetuses ± SD
	Male Fetuses	3.75 ± 0.36	3.80 ± 0.28	3.78 ± 0.20	3.82 ± 0.34
	Female Fetuses	3.61 ± 0.24	3.64 ± 0.36	3.54 ± 0.20	3.54 ± 0.32
	Sex Ratio of Viable Fetuses
	Total Males/Total Females	1.1	1.0	1.1	1.0

Note: Preimplantation Loss = (Corpora lutea - implants) / Corpora lutea

       No statistically significant differences

Table: Summary of fetal examinations by litter

	Exposure Levels (ppm)
	0	1000	3000	9000
No. of Litters Evaluated	24	22	22	21
Soft Tissue Observations
No. Fetuses Evaluated	168^a	172	174	169
Visceral Malformations
Micropthalmia	0	1 (4.5%)	0	0
Soft Tissue Variations
Ureter(s)-Tortuous		1 (4.5%)	1 (4.5%)	0
Skeletal Observations
No. Fetuses Evaluated	160^a	162	162	156
Skeletal Malformations	2 (8.3%)	0	1 (4.5%)	1 (4.8%)
Total Skeletal Variations	23 (95.8%)	22 (100%)	21 (95.5%)	21 (100%)
Rudimentary Ribs	9 (37.5%)	15	13 (59.1%)	16
		(68.2%)		(76.2%)

^a One fetus from one control group litter was evaluated for both soft tissue and skeletal anomalies

Applicant's summary and conclusion

Conclusions:

The maternal NOAEL and developmental NOAEL for unleaded gasoline vapor condensate were 23900 mg/m3 (analytical concentration). Under the conditions of this study, unleaded gasoline vapors did not produce evidence of developmental toxicity.
These findings do not warrant classification of the test material as a developmental hazard under the new Regulation (EC) 1272/2008 on classification, labeling, and packaging of substances and mixtures (CLP) or under the Directive 67/518/EEC for dangerous substances and Directive 1999/45/EC for preparations.

Executive summary:

Unleaded gasoline vapor condensate was administered once daily to pregnant rats on gestation days 6-19 via vapor inhalation at doses of 0, 2653, 7960, or 23900 mg/m3 (24 rats/dose) to assess for developmental toxicity. Maternal parameters (food consumption, body weight gain) monitored throughout gestation and at study termination (clinical chemistry, grossly visible abnormalities) were not adversely affected by treatment.  Reproductive parameters (number of implants, resorptions, or viable fetuses) were not adversely affected by administration of the test material at any of the dose levels tested. No evidence of abnormal development was observed during external, skeletal, or visceral examinations of fetuses from pregnant dams exposed. Thus, unleaded gasoline vapor condensate did not produce any maternal toxicity, fetal toxicity, or developmental effects in rats. Based on the study results, the maternal and developmental NOAELs were both 23900 mg/m3 (analytical concentration).

These findings do not warrant the classification of the test material as a developmental hazard under the new Regulation (EC) 1272/2008 on classification, labeling, and packaging of substances and mixtures (CLP) or under the Directive 67/518/EEC for dangerous substances and Directive 1999/45/EC for preparations.