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Repeated dose toxicity: inhalation

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Administrative data

Endpoint:
short-term repeated dose toxicity: inhalation
Type of information:
experimental study
Adequacy of study:
key study
Reliability:
1 (reliable without restriction)
Rationale for reliability incl. deficiencies:
other: GLP study and Test procedure was in accordance with generally accepted scientific standards and described in sufficient details.

Data source

Referenceopen allclose all

Reference Type:
publication
Title:
Inhalation toxicity of cyclododecatriene in rats.
Author:
Bamberger JR, Scott RS, Kelly DP, Kennedy GL jr. and Elliott GS
Year:
1999
Bibliographic source:
Drug Chem. Toxicol. 22, 435-454.
Reference Type:
study report
Title:
Unnamed
Year:
1996
Reference Type:
publication
Title:
Robust summary for 1,5,9-cyclododecatriene (revised).
Author:
DuPont Safety, Health & Environmental Excellence Center, Wilmington (Del., USA)
Year:
2003
Bibliographic source:
U.S. EPA, 46 pp

Materials and methods

Test guideline
Qualifier:
no guideline followed
Principles of method if other than guideline:
A total of 8 groups of male rats (2 groups per exposure concentration; approximately 7 weeks old on the day of arrival) were exposed nose-only to vapor or a vapor/aerosol mixture of 1,5,9-cyclododecatriene.
GLP compliance:
yes
Limit test:
no

Test material

Constituent 1
Reference substance name:
1,5,9-cyclododecatriene
IUPAC Name:
1,5,9-cyclododecatriene
Details on test material:
1,5,9-cyclododecatriene of DuPont Nylon, Victoria (Texas, USA);
Purity approximately 100%

Test animals

Species:
rat
Strain:
other: Sprague Dawley  Crl:CD(R)(SD)BR
Sex:
male
Details on test animals or test system and environmental conditions:
Strain: Crl:CD(R)(SD)BR
- Source: Charles River Breeding Laboratories, Raleigh (USA)
- Age: ca. 8 weeks
- Weight at study initiation: ca. 245 g (mean)
further information : see reference (Bamberger et al)

Administration / exposure

Route of administration:
inhalation
Type of inhalation exposure:
nose only
Vehicle:
air
Remarks on MMAD:
MMAD / GSD: Particle size: vapour at 5 and 50 ppm;
 aerosol at 260 ppm;   particle size: MMAD 3.5 or 3.7 um,    35 or 36% of particles < 3 um;   98 or 99% of particles < 10 um
Details on inhalation exposure:
All exposure chambers were constructed of stainless steel and glass with a nominal internal volume of 150 L. The chambers were operated in a one-pass, flow-through mode, with air flow rates adequate to provide sufficient oxygen for test rats and enable adequate distribution of 1,5,9-cyclododecatriene in the chambers. For the 5 ppm concentration, vapor atmospheres of 1,5,9-cyclododecatriene were generated by metering the liquid test material into a heated Instatherm flask. Nitrogen was used to carry the 1,5,9-cyclododecatriene vapor to a glass transfer tube where it was mixed with filtered, houseline air and fed into the top of the exposure chamber. Atmospheres of 1,5,9-cyclododecatriene in the 50 and 260 ppm chambers were generated by metering the liquid test material into a nebulizer. Filtered, houseline air introduced into the nebulizer atomized the liquid test substance and carried the resulting vapor/aerosol mixture into a glass cyclone elutriator. At 50 ppm, the remaining aerosol vaporized in the elutriator, and the vapor was carried into the top of the exposur e chamber.
Analytical verification of doses or concentrations:
yes
Details on analytical verification of doses or concentrations:
The concentration of 1,5,9-cyclododecatriene was controlled by varying the amount of test substance delivered to the flask or nebulizer.
The atmosphere concentration of 1,5,9-cyclododecatriene was determined at approximately 60- minute intervals during each exposure by gas chromatography and gravimetric analysis for the vapor and aerosol components, respectively. Chamber airflow, temperature, relative humidity, and
oxygen concentrations were recorded.
Duration of treatment / exposure:
2 weeks
Frequency of treatment:
6 hours/day, 5 days/week, total 9 exposures
Doses / concentrationsopen allclose all
Remarks:
Doses / Concentrations:
5, 50, and 260 ppm = 33.4, 334, and 1740 mg/m3
Basis:
nominal conc.
Remarks:
Doses / Concentrations:
 5 +/- 0.14; 51 +/- 1.0; 260 +/- 5.7 ppm
Basis:
analytical conc.
No. of animals per sex per dose:
10 per group
Control animals:
yes, concurrent no treatment
Details on study design:
- Number of animals: 10 animals per group
-  2 groups per concentration:     
1 for standard toxicological evaluations      
1 for neurotoxicity testing
-Post-exposure period: 2 weeks (half of the animals)
- SATELLITE GROUPS AND REASONS THEY WERE ADDED: neurotoxicity
Positive control:
no

Examinations

Observations and examinations performed and frequency:
- Clinical signs: groupwise during exposure, individually after each  exposure
- Mortality: before / during / after exposure
- Body weight: before each exposure; daily except weekends during  recovery period
- Haematology: end of exposure period
- Biochemistry: end of exposure period
- Urinalysis: end of exposure period
Sacrifice and pathology:
ORGANS EXAMINED AT NECROPSY (MACROSCOPIC AND MICROSCOPIC): 
-5 animals per group sacrificed 1 day after last exposure; other animals sacrificed after (additional) 14 days of recovery

- Macroscopic:    weights of liver, kidneys, lungs, testes, brain;   examination of liver, kidneys, lungs, duodenum, heart, spleen, brain  (cerebrum, midbrain, cerebellum, medulla / pons), spinal cord, stomach,  jejunum, ileum, pancreas, cecum, colon, rectum, mesenteric lymph node,  thymus, adrenal glands, sciatic nerve, thyroid gland, parathyroid glands,  trachea, esophagus, pharynx / larynx, eyes, prostate, seminal vesicles,  urinary bladder, testes epididymides, sternum (with bone marrow), nose;  samples of all plus of gross lesions were saved

- Microscopic:    control and high dose animals sacrificed without recovery period: all  organs listed above (macroscopic examination);   animals from other dose groups sacrificed without recovery and control  and high dose animals sacrificed after recovery: nose, pharynx / larynx,  lungs, liver, kidneys
Other examinations:
OTHER EXAMINATIONS: 
Neurotoxicity groups   Functional observational battery (FOB) of tests   + motor activity (MA) evaluations   immediately after 4th and 9th exposures;   Neuropathology evaluation in 6/10 control and high dose animals after  9th exposure
Statistics:
- Mean, standard deviation, standard error;
- Incidences: Cochran-Armitage test for trend;
- Body & organ weights: one-way analysis of variance;
- Pairwise comparison with controls: Dunnett's test
- Homogeneity of variances: Bartlett's test
- Clinical pathology: one-way analysis of variance, Bartlett's test;  Dunnett's test for comparison of means to control; Kruskal-Wallis test  and Mann-Whitney U test when Bartlett's test showed significance  (p<0.005) 
- FOB: Cochran-Armitage test for trend, Fisher's exact test for  significance of deviations from control 
- Other test in neurobehavioral evaluations: Bartlett's test for  homogeneity (p<0.005), univariate analysis of variance, Shapiro-Wilk's  test, Levene's test, Kruskal-Wallis followed by Dunn's test, block (all  p<0.05)

Results and discussion

Results of examinations

Clinical signs:
effects observed, treatment-related
Description (incidence and severity):
No deaths but clinical signs at 260 ppm
Mortality:
mortality observed, treatment-related
Description (incidence):
No deaths but clinical signs at 260 ppm
Body weight and weight changes:
effects observed, treatment-related
Description (incidence and severity):
at 50 and 260 ppm
Food consumption and compound intake (if feeding study):
not examined
Food efficiency:
not examined
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:
effects observed, treatment-related
Description (incidence and severity):
at 260 ppm
Histopathological findings: neoplastic:
not examined
Details on results:
TOXIC RESPONSE/EFFECTS BY DOSE LEVEL: 
- Mortality and time to death: No mortalities
- Clinical signs: Irregular respiration in the high dose group. Other  clinical observations occurred sporadically in all groups including  controls and were considered incidental.
- Body weight gain: significantly reduced at >= 50 ppm, reversible
- Clinical chemistry: Compound-related or biologically adverse changes  did not occur during this study.
- Haematology: Compound-related or biologically adverse changes did not  occur during this study.
- Urinalysis: Statistically significant or biologically adverse changes  did not occur during this study.
- Organ weights: No treatment-related changes were observed. (Absolute  but not relative lung weights were reduced in the high dose group.)
- Gross pathology: No treatment-related changes were observed
- Histopathology: Effects only occurred in the nasal tissue of the high  dose group. Minimal degeneration / necrosis of olfactory epithelium  occurred in the anterior portion of the nose of 4/5 rats exposed to 260  ppm. This lesion was observed neither in the 260 ppm recovery group,  which suggests reversibility, nor in the lower dosed groups.
- Other: A diminished or absent response to an alerting stimulus was  observed in the high dose group. Mean foot splay was significantly  decreased on days 4 and 9 in the high dose group. The incidence of  defecation was decreased in the 50 and 260 ppm groups during the motor  activity assessments. Beyond this, there were no statistically  significant or toxicologically remarkable neurobehavioral differences  between the control and treatment groups.

Effect levels

Dose descriptor:
NOAEL
Effect level:
33.4 mg/m³ air
Based on:
test mat.
Sex:
male
Basis for effect level:
other: Dose corresponding to 5 ppm. Decreased of bodyweight gain was observad at 50 and 260 ppm.

Target system / organ toxicity

Critical effects observed:
not specified

Any other information on results incl. tables

The analytically determined mean concentrations of

1,5,9-cyclododecatriene in the exposure chambers were

5 ± 0.14, 51 ± 1.0, and 260 ± 5.7 ppm for the 5, 50, and

260 ppm exposure levels, respectively. Particle size

distribution taken from the 260 ppm chamber during the

study showed that the aerosol in the chamber was respirable

in rats with mass median aerodynamic diameters (MMAD)

of 3.5 or 3.7 μm; 35 or 36% of the particles were less than

3 μm and 98 or 99% of the particles were less than 10 μm.

Essentially, no particulates were observed in either the 5 or

50 ppm chambers. The chamber airflow, oxygen

concentration, relative humidity, and temperature were

33-35 L/min, 20-21%, 14-30%, and 22-26°C, respectively.

Applicant's summary and conclusion

Conclusions:
Based on the decreased of bobyweight gain, the NOAEC was 33.4 mg/m3 for male rats after inhalation exposure.
Executive summary:

A total of 8 groups of male rats (2 groups per exposure concentration; approximately 7 weeks old on the day of arrival) were exposed nose-only to vapor or a vapor/aerosol mixture of 1,5,9-cyclododecatriene.

Four groups of 10 rats per concentration were used for standard toxicological evaluations, and 4 groups of 10 rats per concentration were used for neurotoxicity testing. Individual body weights and clinical signs were recorded. During the exposure, group clinical signs were recorded. In additio n, rats were checked for a startle response to an auditory stimulus during exposure. The neurotoxicity groups were given functional observational battery (FOB) assessments and motor activity (MA) evaluations immediately after the 4th and 9th exposures. Hematologic parameters and clinical chemistry parameters were measured or calculated.

Five rats in each group designated for standard toxicology evaluations were sacrificed and necropsied on test day 12 (day following the last exposure) and test day 26. The liver, kidneys, lungs, testes, and brain were weighed. Each rat was given a complete gross examination, and representative samp were saved for histopathologic examination.

There were no mortalities during the study. Body weights for rats exposed to 5 ppm 1,5,9-cyclododecatriene were similar to those of the control during the entire experiment. Rats exposed to 50 or 260 ppm 1,5,9-cyclododecatriene had significantly lower mean body weights and mean body weight gains when compared to controls. The effects on body weight were reversible. Diminished or absent startle response to an auditory stimulus was observed in rats exposed to 260 ppm 1,5,9-cyclododecatriene when compared to controls. The response was first observed generally 2 hours into the exposure. Neither onset nor intensity changed over the 9 exposure days, and the rats were responsive prior to the next day of exposure. These effects were not observed in the other exposure groups. Test substance-related clinical signs of toxicity observed immediately after exposure included irregular respiration (260 ppm) and lethargy (260 ppm). Clinical signs of toxicity noted during and after exposure were reversible and were not present in rats prior to the next exposure.

Body weights measured as part of the functional observational battery assessment were significantly lower for rats in the 260 ppm group when compared to controls. No additional toxicologically relevant findings were observed during the functional observational battery assessments or motor activity evaluation. There were no test substance-related effects in rats noted during clinical pathology evaluations, and no abnormalities were seen during gross pathology and gross neuropathology examinations. There were no compound-related organ weight effects. Histologic effects attributable to the test substance were found in the nasal tissue of rats in the 260 ppm group. There was a minimal degeneration/necrosis of nasal olfactory epithelium observed immediately after the exposure period. These effects were reversed after the 2-week recovery period. No other test-substance related effects were observed microscopically. There were no test substance-related effects in rats noted during the microscopic neuropathologic evaluation.

Based on the decreased of bobyweight gain, the NOAEC was 33.4 mg/m3 for male rats after inhalation exposure.

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