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Toxicological information

Repeated dose toxicity: inhalation

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Administrative data

Endpoint:
chronic toxicity: inhalation
Type of information:
migrated information: read-across from supporting substance (structural analogue or surrogate)
Adequacy of study:
key study
Reliability:
1 (reliable without restriction)
Rationale for reliability incl. deficiencies:
other: NTP study, fully valid for assessment

Data source

Reference
Reference Type:
study report
Title:
Unnamed
Year:
2006
Report Date:
2012

Materials and methods

Test guideline
Qualifier:
according to
Guideline:
other: National Toxicology Program. Toxicology and Carcinogenesis study
GLP compliance:
yes

Test material

Reference
Name:
Unnamed
Type:
Constituent
Type:
Constituent
Type:
Constituent
Details on test material:
Purity/Composition: >=97%

Test animals

Species:
mouse
Strain:
B6C3F1
Sex:
male/female
Details on test animals and environmental conditions:
TEST ANIMALS
- Species: B6C3F1 mice
- Housing: All mice were housed individually.
- Acclimation period: 10-14 days

ENVIRONMENTAL CONDITIONS
Not available

Administration / exposure

Route of administration:
inhalation
Type of inhalation exposure:
not specified
Duration of treatment / exposure:
14 Weeks
Frequency of treatment:
Animals are exposed five times per week, weekdays only until the day prior to necropsy.
Doses / concentrations
Remarks:
Doses / Concentrations:
0, 25, 50, 100, 200, or 400 ppm of α-pinene for 6 h/day, 5 days per week. Calculated to correspond to a daily intake of 36, 72, 144, 288, and 576 mg/kg bw per day.
Basis:

No. of animals per sex per dose:
Treatment: 10 female and 10 male- 5 test groups
Controls: 10 female and 10 male- 1 test group
Animals for clinical lab studies: 10 female and 10 male- 5 test groups
Control animals for clinical lab studies: 10 female and 10 male- 1 test group
Details on study design:
Animals were acclimataded for a period of 10- to 14-days, then they were assigned at random treatment groups. The study included five treatment groups each one corresponded to a different concentration of the test material plus the control group. Each group included 10 animals per sex.
The animals were treated with alpha pinene at different concentrationn by inhalation route. The control group received untreated water or feed or vehicle alone. Animals were exposed five times per week, weekdays only until the day prior to necropsy.

Examinations

Observations and examinations performed and frequency:
Necropsy and Histopathologic Evaluation included observation and weight of Liver, thymus, right kidney, right testis, heart, and lung of all animals surviving until the end of the study.

Sacrifice and pathology:
A complete necropsy was performed on all treated and control animals that either die or were sacrificed. All tissues required for complete histopathology are trimmed, embedded, sectioned and stained with hematoxylin and eosin for histopathologic evaluation.


Treatment-related lesions (target organs) were identified and these organs plus gross lesions were examined to a no-effect level: (respiratory system)
Statistics:
Kaplan-Meier used for probability of survival. Statistical analyses used for possible dose-related effect on survival was Cox (1972) for testing two groups for equality; and Tarone's (1975) life table test for a dose-related trend.

Results and discussion

Results of examinations

Clinical signs:
no effects observed
Mortality:
no mortality observed
Body weight and weight changes:
no effects observed
Description (incidence and severity):
Body weight gain comparable for all test animals
Organ weight findings including organ / body weight ratios:
effects observed, treatment-related
Description (incidence and severity):
Absolute liver weights increased for both sexes at the 400 ppm and relative and absolute liver weights increased for both sexes at 200 ppm and 400. The 400 ppm male group showed decreased absolute and relative thymus weight.
Gross pathological findings:
no effects observed
Description (incidence and severity):
No gross or microscopic lesions
Histopathological findings: non-neoplastic:
effects observed, treatment-related
Details on results:
All mice survived until the study was terminated. Body weights gain were comparable for all test animals when compared to controls.
At 400 ppm, an increse of absolute liver weights was observed in both sexes. At 200 and 400 ppm, an increase of absolute liver weights were recorded for both sexes. The 400 ppm male group showed decreased absolute and relative thymus weight. No gross or microscopic lesions were associated with these organ weight findings.
Histopathological examination of male and female mice exposed to ≥100 ppm of α-pinene revealed evidence of hyperplasia of the transitional epithelium of the urinary bladder. Histopathological examination confirmed that there were no changes in clitoris, ovaries, uterus, epididymis, preputial gland, seminal vesicles, and testes compared to the control groups. Based on these observations, a NOAEL for both male and female mice was concluded to be 50 ppm.

Effect levels

Dose descriptor:
NOAEL
Effect level:
50 ppm
Sex:
male/female

Target system / organ toxicity

Critical effects observed:
not specified

Any other information on results incl. tables

Further histopathological findings from this study are being evaluated by NTP and will be publicly available in the future.

Applicant's summary and conclusion

Conclusions:
Based on the result of the study, the NOAEL for the inhalation 90-Day study is 50 ppm for males and females. The LOAEL is 100 ppm for males and females, based on the presence of transitional cell hyperplasia of the urinary bladder.
Executive summary:

A NTP (National Toxicology Program) 90 -Day study was conducted on alpha-pinene. Female and male B6C3F1 MICE were treated with alpha-pinene at concentration of 0, 25, 50, 100, 200, or 400 ppm for 6h/days per week via inhalation route for 14 weeks. Animals were observed twice per day and weighed once per week.

Histopathologic evaluation were performed on all early death animals regardless of dose group, all control animals, all animals, and all animals in the highest treatment group. Treatment-related lesions (target organs) were identified and these organs plus gross lesions were examined to a no-effect level.

All mice survived until the study was terminated. Body weights gain were comparable for all test animals when compared to controls.

At 400 ppm, an increse of absolute liver weights was observed in both sexes. At 200 and 400 ppm, an increase of absolute liver weights were recorded for both sexes. The 400 ppm male group showed decreased absolute and relative thymus weight. No gross or microscopic lesions were associated with these organ weight findings.

Histopathological examination of male and female mice exposed to ≥100 ppm of α-pinene revealed evidence of hyperplasia of the transitional epithelium of the urinary bladder. Histopathological examination confirmed that there were no changes in clitoris, ovaries, uterus, epididymis, preputial gland, seminal vesicles, and testes compared to the control groups.

Based on these observations, a NOAEL for both male and female mice was concluded to be 50 ppm and the LOAEL was 100 ppm for males and females, based on the presence of transitional cell hyperplasia of the urinary bladder.