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Repeated dose toxicity: inhalation

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Administrative data

Endpoint:
short-term repeated dose toxicity: inhalation
Remarks:
other: Combined repeated exposure toxicity, reproduction and neurotoxicity screening in rats via whole-body inhalation exposures
Type of information:
experimental study
Adequacy of study:
key study
Reliability:
1 (reliable without restriction)
Rationale for reliability incl. deficiencies:
other: GLP compliant, guideline study, available as unpublished report, fully adequate for assessment
Cross-referenceopen allclose all
Reason / purpose:
reference to same study
Reason / purpose:
reference to other study

Data source

Reference
Reference Type:
study report
Title:
Unnamed
Year:
2003
Report Date:
2003

Materials and methods

Test guideline
Qualifier:
according to
Guideline:
OECD Guideline 422 (Combined Repeated Dose Toxicity Study with the Reproduction / Developmental Toxicity Screening Test)
Deviations:
no
Principles of method if other than guideline:
Not applicable
GLP compliance:
yes
Limit test:
no

Test material

Reference
Name:
Unnamed
Type:
Constituent
Details on test material:
- Name of test material (as cited in study report): 1-butene
- Analytical purity: ≥99%

Test animals

Species:
rat
Strain:
other: Crl:CD® (Sprague-Dawley) IGS BR
Sex:
male/female
Details on test animals and environmental conditions:
TEST ANIMALS
- Source: Charles River Laboratories, Kingston, New York, USA
- Age at study initiation: c.a. 8 weeks
- Weight at study initiation: Males 200-300g, females 150-250 g
- Housing: Individually in stainless steel, wire mesh cages within a 1.0 m3 glass and stainless steel whole body exposure chamber
- Diet: Certified Rodent Diet No. 5002 (PMI Nutrition International, St. Louis, MO, USA) ad libitum except during exposure
- Water: Mains water ad libitum except during exposure.
- Acclimation period: 2 weeks

ENVIRONMENTAL CONDITIONS
- Temperature: 20-25°C
- Humidity: 36-82%
- Air changes: Not reported
- Photoperiod: 12 hrs dark / 12 hrs light

IN-LIFE DATES: From: 1 July 2002 To: 25 August 2002

Administration / exposure

Route of administration:
inhalation: gas
Type of inhalation exposure:
whole body
Vehicle:
other: air
Remarks on MMAD:
MMAD / GSD: MMAD: 0.822 ± 0.1, 0.893 ± 0.2, 0.7986 ± 0.0343 and 3.6503 ± 7.0 for 0, 500, 2000 and 8000 ppm respectively
GSD: 2.019 ± 0.4, 1.869 ± 0.3, 1.893 ± 0.3 and 2.181 ± 0.8 for 0, 500, 2000 and 8000 ppm respectively
Details on inhalation exposure:
GENERATION OF TEST ATMOSPHERE / CHAMBER DESCRIPTION
The whole-body exposure chambers each had a volume of approximately 1000 L. The chambers were operated at a minimum flow rate of 200 L/minute. The final airflow was set to provide at least one air change in 5.0 minutes (12 air changes/hour) and a T99 equilibrium time of at most 23 minutes At the end of the 6-hour exposure, all animals remained in the chambers for a minimum of 30 minutes. During this time, the chambers were operated at approximately the same flow rate using clean air only. The chambers were exhausted through the in housed filtering system, which consisted of a coarse filter, a HEPA filter and activated charcoal.
For the treated groups, the test article was delivered from the cylinder via a regulator with a backpressure gauge via 1/4" tubing through a flowmeter via 1/4" tubing. The outlet of the flowmeter was regulated by a built-in metering valve. The test article laden airstream was directed into the turret of a 1.0 m3 glass and stainless steel exposure chamber via 1/4" tubing, where it was diluted with room air as it was drawn into the chamber. For controls, room air was drawn into the turret of the 1.0 m3 glass and stainless steel exposure chamber.

TEST ATMOSPHERE
During each exposure, measurements of airborne concentrations were performed in the animals' breathing zone at least 4 times using an appropriate sampling procedure and on-line GC analytical method. During each week of exposure, particle size determinations were performed using a TSI Aerodynamic Particle Sizer to characterize the aerodynamic particle size distribution of any aerosol present.

Analytical verification of doses or concentrations:
yes
Details on analytical verification of doses or concentrations:
The mean (± standard deviation) analytical (GC) concentrations for the control and the exposure groups were as follows: 0 ± 0, 524 ± 40, 2062 ± 126 and 8271 ± 683 ppm. The analytically measured exposure levels of the airborne test article were reasonably close to the targeted exposure levels.
Duration of treatment / exposure:
28 days
Frequency of treatment:
6 hours/day, 7 days/week
Doses / concentrationsopen allclose all
Remarks:
Doses / Concentrations:
0, 500, 2000, or 8000 ppm
Basis:
other: target concentrations
Remarks:
Doses / Concentrations:
0 ± 0, 524 ± 40, 2062 ± 126 and 8271 ± 683 ppm
Basis:
analytical conc.
No. of animals per sex per dose:
12
Control animals:
yes, sham-exposed
Details on study design:
The study design included the main study for repeated dose toxicity end points and reproductive/ developmental toxicity satellite groups (summarized separately). The reproductive and developmental toxicity satellite groups (12 females per exposure level) were exposed for two weeks prior to breeding, during breeding, and continuing through day 19 of gestation. Males from the main study were used to breed these females.
Positive control:
None

Examinations

Observations and examinations performed and frequency:
Effects on general toxicity, neurobehavioural activity, clinical chemistry, coagulation and haematology were evaluated.
CAGE SIDE OBSERVATIONS: Yes
- Time schedule: at least twice/day

DETAILED CLINICAL OBSERVATIONS: Yes
- Time schedule: prior to randomisation and then once/week

BODY WEIGHT: Yes
- Time schedule for examinations: weekly

FOOD CONSUMPTION: Yes
- Time schedule for examinations: weekly

FOOD EFFICIENCY: No

WATER CONSUMPTION: No

OPHTHALMOSCOPIC EXAMINATION: No

HAEMATOLOGY: Yes
- Time schedule for collection of blood: at termination
- Anaesthetic used for blood collection: Yes (carbon dioxide/oxygen)
- Animals fasted: Yes
- How many animals: 12/sex/group
- Parameters examined: erythrocyte count, haematocrit, haemoglobin, MCV, MCHC, leucocyte count (total and differential), platelet count, reticulocyte count, erythrocyte morphology, prothrombin time, activated partial thromboplastin

CLINICAL CHEMISTRY: Yes
- Time schedule for collection of blood: at termination
- Anaesthetic used for blood collection: Yes (carbon dioxide/oxygen)
- Animals fasted: Yes
- How many animals: 12/sex/group
- Parameters examined: alanine aminotransferase, aspartate aminotransferase, alkaline phosphatase, gamma glutamyl transpeptidase, urea nitrogen, creatinine, glucose, total cholesterol, triglycerides, total protein, albumin, globulin, albumin/globulin ratio, total bilirubin, sodium, potassium, chloride, calcium, phosphorus

URINALYSIS: No

NEUROBEHAVIOURAL EXAMINATION: Yes
- Time schedule for examinations: pre-test and during final week of exposure
- Dose groups that were examined: All
- Battery of functions tested: sensory activity / grip strength / motor activity / rectal temperature
Sacrifice and pathology:
GROSS PATHOLOGY: Yes (all animals including those dying spontaneously or killed moribund). The nasal pharynx was preserved but not examined microscopically.

ORGAN WEIGHTS: Yes (testes, epididymides, ovaries with oviducts, uterus with vagina, adrenals, brain with brain stem, heart, kidneys, liver, lungs, spleen, thymus)

HISTOPATHOLOGY: Yes (control and high dose only initially. Tissues examined: adrenals, bone marrow (femur), brain (medulla/pons, cerebrum and cerebellum), epididymides, heart, kidneys, large intestine (caecum, colon and rectum), liver, lungs (with mainstem bronchi), lymph nodes (mesenteric and mediastinal), mammary gland, ovary with oviduct, prostate, seminal vesicles, small intestine (duodenum, ileum and jejunum), spinal cord, spleen, stomach, testes, thymus, thyroid with parathyroids, tibial nerve, trachea, urinary bladder, uterus with vagina, all macroscopic lesions and tissue masses.
Other examinations:
none
Statistics:
Mean values of all exposure groups were compared to the mean value for the control group at each time interval. For all parameters except for organ weights, the standard one-way analysis of variance (ANOVA) using the F ratio to assess significance was used. If significant differences among the means were indicated, additional testing was performed using Dunnett's t-test to determine which means were significantly different from the control. Organ weight data was analyzed only by parametric methods. Bartlett's test was performed to determine if groups had equal variances. The standard
one-way analysis of variance (ANOVA) using the F ratio to assess significance was used. If significant differences among the means were indicated, additional tests were used to determine which means were significantly different from the control: Dunnett's t-test for homogeneous data, or Cochran and Cox's modified t-test for non-homogeneous data. All t-tests were conducted at the 5% and 1% significance levels. Motor Activity Data was analyzed using split-plot repeated measures ANOVA with model terms for group, animal within group, interval and group by interval interaction. If the group x interval interaction was statistically significant (p=0.05), indicating non-parallelism in the behavioural profile between groups, a separate one-way ANOVA for group effects was performed at each interval. If the response data passed on the parallel hypothesis, an ANOVA (using summed responses over intervals) was used to test for the overall treatment effect which constituted the level hypothesis. If any significant overall treatment group effect was found by any of the above ANOVAs, Dunnett's t-test was used to find groups that differed from control. Analyses were performed for sexes separately and combined. Treatment group effects were deemed significant at the p=0.05 level. Plots, tables, listings, and analyses were generated using SAS version 6.12 for WINDOWS.

Results and discussion

Results of examinations

Clinical signs:
no effects observed
Mortality:
no mortality observed
Body weight and weight changes:
no effects observed
Food consumption and compound intake (if feeding study):
no effects observed
Food efficiency:
not specified
Water consumption and compound intake (if drinking water study):
not examined
Ophthalmological findings:
not examined
Haematological findings:
no effects observed
Clinical biochemistry findings:
no effects observed
Urinalysis findings:
not examined
Behaviour (functional findings):
no effects observed
Organ weight findings including organ / body weight ratios:
no effects observed
Gross pathological findings:
no effects observed
Histopathological findings: non-neoplastic:
no effects observed
Histopathological findings: neoplastic:
no effects observed
Details on results:
See below

Effect levels

Dose descriptor:
NOAEC
Remarks:
Toxicity
Effect level:
8 000 other: ppm (18359 mg/m3, 18 mg/L) nominal
Sex:
male/female
Basis for effect level:
other: there were no adverse effects at 8000 ppm, the highest concentration tested

Target system / organ toxicity

Critical effects observed:
not specified

Any other information on results incl. tables

There were no microscopic findings considered to be related to exposure to 1-butene. In comparison with controls, there was a slightly increased incidence and severity of mixed inflammatory cells in the caecal mucosa of rats exposed to 1-butene at exposure levels of 2000 ppm and above. The caecal mucosa normally contains a small population of mixed inflammatory cells, which acts as a natural defence mechanism against ingested substances or organisms. Increased numbers of inflammatory cells are sometimes seen as a normal spontaneous finding, and this was evident in a few males and females from the control group in this study. Since the finding was present in the control group and there was no clear exposure level response relationship in the treated groups, the increased incidence is considered to be fortuitous and unlikely to be related to treatment with 1 -butene. Other microscopic findings occurred sporadically or showed a similar incidence in control and 1-butene-treated animals. None were considered to be associated with exposure to 1-butene.

Applicant's summary and conclusion

Conclusions:
Exposure of male and female rats to target concentrations of 500, 2000 and 8000 ppm of 1-butene resulted in no general systemic effects.
Executive summary:

Exposure to 1-butene at target concentrations of 500, 2000, 8000 ppm (approximately 1147, 4589, 18359 mg/m3) did not induce systemic toxicity in male and female rats exposed for 28 days or in pregnant female rats exposed for 14 days pre-mating, through mating and gestation to day 19. No treatment-related effects on body weight, clinical chemistry, organ weights or histopathology were found. Neurotoxicity screening also showed no effects on motor activity or functional observation battery. The NOAECs were at the highest concentration level tested