Gases (petroleum), C3-5 olefinic-paraffinic alkylation feed

Toxicity to reproduction

Administrative data

Endpoint:

one-generation reproductive toxicity

Remarks:

based on generations indicated in Effect levels (migrated information)

Type of information:

migrated information: read-across based on grouping of substances (category approach)

Adequacy of study:

key study

Reliability:

1 (reliable without restriction)

Rationale for reliability incl. deficiencies:

other: GLP compliant (except for minor exception listed below), guideline study, available as unpublished report, no restrictions, fully adequate for assessment.

Data source

Reference

Reference Type:

study report

Title:

Unnamed

Year:

2010

Report date:

2010

Materials and methods

Test guideline

Test guideline 1

Qualifier:

according to guideline

Guideline:

OECD Guideline 422 (Combined Repeated Dose Toxicity Study with the Reproduction / Developmental Toxicity Screening Test)

Deviations:

yes

Remarks:

no lot number available of test material

Test guideline 2

Qualifier:

according to guideline

Guideline:

other: EPA OPPTS 870.3650

GLP compliance:

yes

Limit test:

no

Test material

Test material information

Constituent 1

Reference substance name:

Ethane

EC Number:

200-814-8

EC Name:

Ethane

Cas Number:

74-84-0

Molecular formula:

C2H6

IUPAC Name:

ethane

Details on test material:

- Name of test material (as cited in study report): ethane
- Supplier: MG Industries, 3 Great Valley Parkway, Malvern, Pennsylvania 19355, USA
- Substance type: Industrial gas
- Physical state: colourless gas
- Analytical purity: 99.0% per supplier
- Lot/batch No.: Not available
- Stability under test conditions: 99.86% before study, 99.88% after study
- Storage condition of test material: Ambient

Test animals

Species:

rat

Strain:

Sprague-Dawley

Sex:

male/female

Details on test animals or test system and environmental conditions:

- Species: Albino rats (Outbred) VAF/Plus®, Sprague-Dawley derived (CD®), Crl:CD®(SD)IGS BR
- Source: Charles River Laboratories, Raleigh, North Carolina 27610, USA
- Age at study initiation: Approximately 8 weeks
- Weight at study initiation: Males mean 256 g (range 230-284 G); females mean 197 g (range 167-217 g)
- Fasting period before study: None
- Housing: Individually in stainless steel suspended cages with wire mesh floors and fronts (except for mating period when 1 male and 1 female were housed together)
- Diet: Certified Rodent diet No 5002 (PMI Nutrition International, St Louis, Missouri, USA) ad libitum except during exposure
- Water: Municipal water ad libitum except during exposure
- Acclimation period: Approximately 2 weeks

ENVIRONMENTAL CONDITIONS
- Temperature: 20.7 - 23.2°C
- Humidity: 19.96-74.37%
- Air changes (per hr): Not reported
- Photoperiod: 12 hrs dark / 12 hrs light

IN-LIFE DATES: From: 24 November 2003 To: 17 January 2004

Administration / exposure

Route of administration:

inhalation: gas

Type of inhalation exposure (if applicable):

whole body

Vehicle:

other: air

Details on exposure:

GENERATION OF TEST ATMOSPHERE / CHAMBER DESCRIPTION
- Exposure apparatus: 1000 L glass and stainless steel whole-body exposure chamber
- Method of holding animals in test chamber: housed individually, the placement of animals in the chamber was rotated daily to ensure uniform exposure
- System of generating particulates/aerosols: the test substance was delivered from a single cylinder, through a regulator and two backpressure gauges via a flowmeter into the exposure chambers
- Time to T99: 23 minutes maximum
- Airflow rate: 204 Lpm
- Temperature and humidity in chamber: 20-24°C, 28-59%
- Oxygen level: at least 19%
- Air flow rate: minimum flow rate of 200 L/minute
- Air change rate: final airflow set to provide at least one air change in 5 mins (12 air changes/hour)
- Method of particle size determination: determined weekly using a TSI Aerodynamic Particle Sizer
- Treatment of exhaust air: filtered through a system which consisted of a coarse filter, a HEPA filter and an activated charcoal bed

TEST ATMOSPHERE
- Brief description of analytical method used: Infrared spectrophotometer (IR) 4 times per chamber per day
- Samples taken from breathing zone: yes

Details on mating procedure:

- M/F ratio per cage: 1:1
- Length of cohabitation: until evidence of mating was seen, or for two consecutive weeks
- Proof of pregnancy: vaginal plug / sperm in vaginal smear referred to as day 0 of pregnancy
- After successful mating each pregnant female was caged (how): individually in plastic "shoebox" cages with bedding
- After the mating period was over, females without evidence of copulation were removed from the mating cages, housed individually and monitored for visible signs of pregnancy with corresponding bodyweight gain.
- Any other deviations from standard protocol: no

Analytical verification of doses or concentrations:

yes

Details on analytical verification of doses or concentrations:

Exposure levels were determined using an infrared spectrophotometer 4 times/chamber/day. The test substance was evenly distributed within each chamber. The mean (± SD) analytical concentrations were 0.0 ± 0.0, 1599 ± 59, 5186 ± 285 and 16380 ± 626 ppm.

Duration of treatment / exposure:

Males for 2 weeks prior to mating and for an additional 28 days (minimum) after mating.
Females for 2 weeks prior to mating and gestation days 0-19.

Frequency of treatment:

6 hours/day, 7 days/week

Details on study schedule:

Females without evidence of mating that appeared to be pregnant were killed on an estimated gestation day 19.
Females that littered and their offspring were killed on post partum day 4.

Doses / concentrations

Doses / concentrations 1

Remarks:

Doses / Concentrations:
0, 1600, 5000 and 16000 ppm
Basis:
other: target concentration

Doses / concentrations 2

Remarks:

Doses / Concentrations:
0.0 ± 0.0, 1599 ± 59, 5186 ± 285 and 16380 ± 626 ppm
Basis:
analytical conc.

No. of animals per sex per dose:

12

Control animals:

yes, sham-exposed

Examinations

Parental animals: Observations and examinations:

CAGE SIDE OBSERVATIONS: Yes
- Time schedule: twice daily (mortality and clinical condition)

DETAILED CLINICAL OBSERVATIONS: Yes
- Time schedule: - Time schedule: Male rats were examined prior to randomisation and once weekly throughout the study. Female rats were examined prior to randomisation and once weekly throughout the premating period and on gestation days 0, 7, 14, 20 and lactation days 0 (except if parturition was not completed on the same day), 1 and 4.

BODY WEIGHT: Yes
- Time schedule for examinations: Male rats were weighed at randomisation and then weekly throughout the study. Females were weighed at randomisation, on the first day of exposure and twice weekly until evidence of copulation was observed, on gestation days 0, 7, 14 and 20, and on lactation days 1 and 4. Females were not fasted prior to recording terminal bodyweights.

FOOD CONSUMPTION: Yes
- Time schedule for examinations: Once weekly throughout the premating period. For pregnant or confirmed mated females, food consumption was recorded on gestation days 0-7, 7-14, 14-20 and on lactation days 1-4.

WATER CONSUMPTION: No

OPHTHALMOSCOPIC EXAMINATION: No

Oestrous cyclicity (parental animals):

No

Sperm parameters (parental animals):

During the microscopic examination of the testes, special emphasis was placed on the stages of spermatogenesis and the histopathology of interstitial testicular cell structure.

Litter observations:

Not applicable

Postmortem examinations (parental animals):

SACRIFICE
- Male animals: All surviving animals (after a minimum of 28 days post-mating).
- Maternal animals: All surviving animals (28 days post-mating).

GROSS NECROPSY: Yes (all animals).
- Tissues examined: adrenal glands, bone (sternum/femur), bone marrow, brain (medulla/pons, cerebrum and cerebellum), epididymides, heart, kidneys, caecum, colon, rectum, larynx, liver, lungs (with mainstem bronchi), lymph nodes (mesenteric and mediastinal), mammary glands (with adjacent skin), nasopharynx, ovaries (with oviducts), prostate, seminal vesicles, duodenum, ileum, jejunum, spinal cord (cervical, thoracic and lumbar), spleen, stomach, testes, thymus, thyroid with parathyroids, tibial nerve, trachea, urinary bladder, uterus with vagina and all macroscopic lesions and tissue masses.

ORGAN WEIGHTS: Yes. Adrenal glands, brain, epididymides, heart, kidneys, liver, lungs, ovaries, spleen, testes, thymus and uterus with vagina.

HISTOPATHOLOGY: Yes (male and female main study only).
- tissues examined: adrenal glands, bone (sternum/femur), brain (cerebellum, cerebrum and cerebellum), epididymes, heart, kidneys, large intestine (caecum, colon and rectum), liver, lungs (with mainstream bronchi), lymph node (mesenteric), lymph node (mediastinal), mammary glands (with adjacent skin), ovaries (with oviducts), prostate, seminal vesicles, small intestine (duodenum, ileum and jejunum), spinal cord (cervical, thoracic and lumbar), spleen, stomach, testes, thymus, thyroids with parathyroids, tibial nerve, trachea, urinary bladder, uterus with vagina, all macroscopic lesions and tissue masses.

Postmortem examinations (offspring):

Macroscopic postmortem examinations (external only) were performed on all surviving F1 pups on lactation day 4.

Statistics:

Group mean values of parameters for all the exposure groups were compared to the control group mean values at each time interval, using appropriate statistical methods.

Reproductive indices:

Male and female mating indices, pregnancy rates, male and female fertility indices, gestation indices and the incidence of dams with no viable pups, were analysed statistically.

Offspring viability indices:

Live birth index, litter survival and mean pup survival indices (days 0 and 4) were analysed statistically.

Results and discussion

Results: P0 (first parental generation)

General toxicity (P0)

Clinical signs:

no effects observed

Body weight and weight changes:

effects observed, treatment-related

Food consumption and compound intake (if feeding study):

effects observed, treatment-related

Organ weight findings including organ / body weight ratios:

no effects observed

Histopathological findings: non-neoplastic:

no effects observed

Reproductive function / performance (P0)

Reproductive function: sperm measures:

no effects observed

Reproductive performance:

no effects observed

Details on results (P0)

Almost all mated females were found pregnant and delivered live pups. Mating indices for the males were comparable to control. Mating, fertility and gestation indices for females were comparable to control. All but one female mated at the first opportunity. There were no treatment-related differences in the other reproductive parameters up to time of parturition, including percentange of females completing delivery and duration of gestation. There were no microscopic findings considered to be treatment-related. No effects on pre or post-implantation loss.

Effect levels (P0)

1

Dose descriptor:

NOAEC

Remarks:

reproductive toxicity

Effect level:

16 000 ppm

Sex:

male/female

Basis for effect level:

other: highest concentration tested

2

Dose descriptor:

NOAEC

Remarks:

reproductive toxicity

Effect level:

19 678 mg/m³ air

Sex:

male/female

Basis for effect level:

other: highest concentration tested

Results: F1 generation

General toxicity (F1)

Clinical signs:

no effects observed

Mortality / viability:

no mortality observed

Body weight and weight changes:

no effects observed

Gross pathological findings:

no effects observed

Details on results (F1)

No effects on total number of pups delivered, number of pups dying, viability (4 day survival) index, pup sex ratio or number of live pups/litter. No treatment-related differences in pup bodyweights or weight gains. No treatment-related differences in pup macroscopic findings.

Effect levels (F1)

1

Dose descriptor:

NOAEC

Generation:

F1

Effect level:

16 000 ppm

Sex:

male/female

Basis for effect level:

other: highest concentration tested

2

Dose descriptor:

NOAEC

Generation:

F1

Effect level:

19 678 mg/m³ air

Sex:

male/female

Basis for effect level:

other: highest concentration tested

Overall reproductive toxicity

Reproductive effects observed:

not specified

Applicant's summary and conclusion

Conclusions:

Inhalation exposure for up to 42 days had no effect on gestation duration, number of live and dead pups, pup abnormalities or pup sex and weights. The NOAEC for reproductive toxicity was 16000 ppm, equivalent to 19678 mg/m3.

Executive summary:

The study assessed the potential toxicity, including neurotoxicity and reproductive performance in male and female rats following ethane exposure at 1600, 5000 and 16000 ppm (highest exposure level was 50% of the lower explosive limit). It also was designed to investigate effects in both sexes on mating behaviour and on gonadal function, as well as effects on conception, development, parturition and pup survival to lactation day 4.

Male and female rats were exposed for 6 hours/day, 7 days/week for 2 weeks prior to mating initiation.Main study females were evaluated for subchronic effects and were exposed once daily (6 hours/day), seven days/week for 4 weeks (28 days). A satellite group of females was evaluated for reproductive effects only - exposed once daily (6 hours/day), seven days/week for at least two weeks prior to mating initiation, then once daily during mating and gestation (days 0-19). For satellite female rats without evidence of mating that appeared to be pregnant, exposure was terminated on the estimated gestation day 19. Main study male rats were exposed during the mating and post-mating periods until euthanized for a minimum exposure of 28 days.

There was no effect on survival. There were no exposure-related clinical effects or effects on body weight, food consumption, FOB or motor activity parameters for either sex (except the16000 ppm exposed animals showed marginally lower food consumption during the first week of exposures).There were no exposure-related differences in haematology, clinical chemistry and no macroscopic or microscopic changes at post-mortem.

lmost all mated female animals were found pregnant and delivered live pups. Mating indices for the ethane male rats were comparable to control. Mating, fertility and gestation indices for the female rats were comparable to control. All but one of the females in each group mated at the first opportunity. There were also no treatment-related differences inthe other reproductive parameters up to the time of parturition including the percent offemales completing delivery and the duration of gestation. There were no exposure-related differences in any parturition parameters including pre-implantation loss, post-implantation loss, the total number of pups delivered, the number of pups dying, the viability (4 day survival) index, the pup sex ratio and the number of live pups/litter, when compared to the air control group.There were no exposure-related differences in body weights or weight gains in the pups feeding from ethane exposed females during gestation compared to the pups feeding from air control animals.There were no macroscopic changes at post-mortem.

Exposure of male and female rats to target concentrations of 1600, 5000 or 16000 ppm of ethane by whole-body inhalation for 4-6 weeks resulted in no general systemic/neurotoxic effects apart from a very marginal reduction of food consumption during the first week of exposure at 16000 ppm and this transient difference was not considered adverse. There were no effects on fertility or reproductive performance, including offspring survival and weight development up to post-natal day 4.

A no-observed-adverse effect concentration (NOAEC) of 16000 ppm was determined for all endpoints. Equivalent to 19678 mg/m3 (MW 30.07g/mol).