3a,4,7,7a-tetrahydro-4,7-methanoindene

Repeated dose toxicity: oral

Administrative data

Endpoint:

short-term repeated dose toxicity: oral

Remarks:

Combined repeated dose and reproduction / developmental screening Test

Type of information:

experimental study

Adequacy of study:

key study

Study period:

Not specified in the study report

Reliability:

1 (reliable without restriction)

Rationale for reliability incl. deficiencies:

guideline study

Remarks:

GLP study, guideline study, available as unpublished report and summary information sheet (provided by Japan Chemical Industry Ecology-Toxicology and Information Center (JETOC)), minor restrictions in design and/or reporting but otherwise adequate for assessment.

Cross-reference

Reason / purpose for cross-reference:

reference to same study

Data source

Reference

Reference 1

Reference Type:

other: Information sheet

Title:

Unnamed

Year:

1998

Report date:

1998

Reference 2

Reference Type:

study report

Title:

Unnamed

Year:

1993

Report date:

1993

Materials and methods

Test guideline

Qualifier:

equivalent or similar to guideline

Guideline:

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

Principles of method if other than guideline:

n/a

GLP compliance:

yes

Limit test:

no

Test material

Test material information

Constituent 1

Reference substance name:

Reference substance 001

Cas Number:

77-73-6

Molecular formula:

C10H12

Details on test material:

- Name of test material (as cited in study report): dicyclopentadiene
- Analytical purity: 94.65%
- Physical state: colourless liquid with a camphor-like odour
- Lot/batch No.: D93028
- Stability under test conditions: confirmed to be stable by the manufacturer for the study period
- Storage condition of test material: room temperature

Specific details on test material used for the study:

SOURCE OF TEST MATERIAL
- Source and lot/batch No.of test material: Zeon; Lot No. D93028
- Expiration date of the lot/batch: Not specified
- Purity test date: Not specified
- Storage condition of test material: Test substance was stored at room temperature
- Stability under test conditions: stable during the study period

FORM AS APPLIED IN THE TEST (if different from that of starting material) : colorless liquid with camphor-like odor

OTHER SPECIFICS:
purity 94.65%
Molecular weight: 132.22
Boling Point: 170°C
Melting Point: -1°C
Density: 0.986
Solubility in water: 40 ppm
Vapour pressure: 9.75 mm Hg (37.7°C)

Test animals

Species:

rat

Strain:

Sprague-Dawley

Remarks:

Crj:CD

Details on species / strain selection:

n/a

Sex:

male/female

Details on test animals or test system and environmental conditions:

TEST ANIMALS
- Source: Charles River Japan, Inc.
- Age at study initiation: 8 weeks
- Weight at study initiation: males 304-339 g, females 186-227 g
- Housing: individually, except during mating, in polycarbonate cages
- Diet: CRF-1 (Oriental Yeast Co) assumed ad libitum
- Water: ultraviolet irradiated water (assumed ad libitum)
- Acclimation period: 6 days

ENVIRONMENTAL CONDITIONS
- Temperature: 20-25°C
- Humidity: 40-70%
- Air changes: approximately 12 per hr
- Photoperiod: 12 hrs dark / 12 hrs light

IN-LIFE DATES: Not reported

Administration / exposure

Route of administration:

oral: gavage

Details on route of administration:

n/a

Vehicle:

olive oil

Details on oral exposure:

PREPARATION OF DOSING SOLUTIONS: Test substance mixed with olive oil, dose rate 5 mL/kg bodyweight

VEHICLE
- Justification for use and choice of vehicle (if other than water): Olive Oil
- Amount of vehicle (if gavage): 5 mL/Kg

Analytical verification of doses or concentrations:

yes

Details on analytical verification of doses or concentrations:

Stability and achieved concentration of dosing preparations was confirmed prior to dosing.

Duration of treatment / exposure:

Males 44 days; Females from 14 days before mating through gestation and parturition until day 3 of lactation

Frequency of treatment:

once daily

Doses / concentrations

Doses / concentrations 1

Dose / conc.:

0 mg/kg bw/day (nominal)

Remarks:

Basis: nominal in olive oil

Doses / concentrations 2

Dose / conc.:

4 mg/kg bw/day (nominal)

Remarks:

Basis: nominal in olive oil

Doses / concentrations 3

Dose / conc.:

20 mg/kg bw/day (nominal)

Remarks:

Basis: nominal in olive oil

Doses / concentrations 4

Dose / conc.:

100 mg/kg bw/day (nominal)

Remarks:

Basis: nominal in olive oil

No. of animals per sex per dose:

10/sex/dose

Control animals:

yes, concurrent vehicle

Details on study design:

- Dose selection rationale: - Based on the results obtained in a 10 day oral dosing preliminary study, where doses of 0, 30, 100 and 300 mg/kg were administered.
- Rationale for animal assignment (if not random): stratified random sampling method by weight

Positive control:

n/a

Examinations

Observations and examinations performed and frequency:

CLINICAL OBSERVATIONS: Yes
- Time schedule: daily

BODY WEIGHT: Yes
- Time schedule for examinations: weekly

FOOD CONSUMPTION: Yes

FOOD EFFICIENCY: No

WATER CONSUMPTION: No

OPHTHALMOSCOPIC EXAMINATION: No

HAEMATOLOGY: Yes (males only)
- Time schedule for collection of blood: termination
- Anaesthetic used for blood collection: Yes (sodium thiopental)
- Animals fasted: Yes (assumed)
- How many animals: 10/group
- Parameters examined: red blood cell, white blood cell, platelets, haemoglobin, haematocrit, differential white cell count, reticulocyte, mean corpuscular volume, mean corpuscular haemoglobin, mean corpuscular haemoglobin concentration

CLINICAL CHEMISTRY: Yes (males only)
- Time schedule for collection of blood: termination
- Anaesthetic used for blood collection: Yes (sodium thiopental)
- Animals fasted: Yes (assumed)
- How many animals: 10/group
- Parameters examined: GOT, GPT, ALP, ¿-GTP, urea nitrogen, glucose, total cholesterol, triglycerides, creatinine, total bilirubin, total protein, albumin, A/G ratio, calcium, inorganic phosphorus, sodium, potassium, chloride

URINALYSIS: No

NEUROBEHAVIOURAL EXAMINATION: No

IMMUNOLOGY: No

Sacrifice and pathology:

GROSS PATHOLOGY: Yes

ORGAN WEIGHTS: Yes
- organs weighed: thymus, liver, kidneys, adrenals, testes, epididymes

HISTOPATHOLOGY: Yes (liver, kidney and adrenals all groups, other tissues controls and 100 mg/kg groups only):
- tissues examined: thymus, liver, kidneys, adrenals, testes, epididymes, brain, heart, spleen, ovaries,

Optional endpoint(s):

n/a

Other examinations:

n/a

Statistics:

Bartlett's test if uniformly distributed analysis of variance, Kruskal-Wallis if non-uniform for quantitative data. When significant differences found between groups,Dunnett-type test or Scheff test. Significance level of 5% or less.

Results and discussion

Results of examinations

Clinical signs:

effects observed, treatment-related

Description (incidence and severity):

Two females in the high dose (100 mg/kg) group died. In these decedents the following major observations were noted: lung congestion, enlargement of the adrenal gland, and bleeding of the gastric mucosa and thymus.

Transient salivation was observed immediately after dosing in the 100 mg/kg dose group after the start of dosing and was still observed after eight days in more than half of male and female rats. This was also seen in a few male rats in the 20 and 4 mg/kg dose groups.

Mortality:

mortality observed, treatment-related

Description (incidence):

Two females in the high dose (100 mg/kg) group died.

Body weight and weight changes:

effects observed, treatment-related

Description (incidence and severity):

Both male and female rats showed suppression of body weight gain and decrease in food consumption at 100 mg/kg.

Food consumption and compound intake (if feeding study):

effects observed, treatment-related

Description (incidence and severity):

Both male and female rats showed suppression of body weight gain and decrease in food consumption at 100 mg/kg.

Food efficiency:

not examined

Water consumption and compound intake (if drinking water study):

not examined

Ophthalmological findings:

not examined

Haematological findings:

effects observed, non-treatment-related

Description (incidence and severity):

Decrease in RBC count and hematocrit value in males in the 20 mg/kg bw/day dose group were observed as was a decrease in hemoglobin concentration in males of the 100 mg/kg bw/day dose group. These were well recognized and within the range of values of both physiological changes. There were no hematological changes ascribable to the test material in any group.

Clinical biochemistry findings:

effects observed, treatment-related

Description (incidence and severity):

Blood chemistry examinations in male rats in the 100 mg/kg bw/day dose group showed increases in glutamic oxaloacetic transaminase (GOT) and glutamic-pyruvate transaminase (GPT).

Endocrine findings:

not examined

Urinalysis findings:

not examined

Behaviour (functional findings):

not examined

Immunological findings:

not examined

Organ weight findings including organ / body weight ratios:

effects observed, treatment-related

Description (incidence and severity):

There were increased liver and kidney weights in male rats in the 100 mg/kg bw/dose group (neither achieved statistical significance) and statistically significantly increased actual and relative liver weight in males at 20 mg/kg/day.

Gross pathological findings:

effects observed, treatment-related

Description (incidence and severity):

Swelling of the liver; swollen adrenal glands; multifocal gray-white discoloration of the kidney were observed in the 100 mg/kg bw/day male rats. Enlargement of the liver in one 20 mg/Kg bw/day male rats was also observed but determined to be incidental. Other observations included polycystic kidney enlargement, testicular atrophy, enlargement of the spleen, and subcutaneous nodules.

In the 2 females in the 100 mg/kg bw/day dose group that died, the following major observations were noted: lung congestion, enlargement of the adrenal gland, and bleeding of the gastric mucosa and thymus.

Neuropathological findings:

not examined

Histopathological findings: non-neoplastic:

effects observed, treatment-related

Description (incidence and severity):

In male rats given 100 mg/kg, single cell necrosis in liver, and hyaline droplets and basophilic changes in tubular epithelium of kidneys was seen. Increase in fatty droplets in fascicular zone of adrenals was observed in both males and females in the 100 mg/kg bw/day dose group. Similar histopathological changes were seen in kidneys of 4, 20 mg/kg bw/day dose group male rats and in adrenals of 20 mg/kg bw/day dose group male rats.

Histopathological findings: neoplastic:

no effects observed

Other effects:

not examined

Details on results:

n/a

Effect levels

1

Key result

Dose descriptor:

NOAEL

Effect level:

4 mg/kg bw/day (nominal)

Based on:

test mat.

Sex:

male

Basis for effect level:

other: Systemic Toxicity

2

Key result

Dose descriptor:

NOAEL

Effect level:

20 mg/kg bw/day (nominal)

Based on:

test mat.

Sex:

female

Basis for effect level:

other: Systemic Toxicity

Target system / organ toxicity

Key result

Critical effects observed:

yes

Lowest effective dose / conc.:

20 mg/kg bw/day (nominal)

System:

immune system

Organ:

adrenal glands

Treatment related:

yes

Dose response relationship:

yes

Relevant for humans:

not specified

Any other information on results incl. tables

Table 1. Results of the Hematological Examination in Male Rats
Dose (mg/kg/day)	0	4	20	100
Number of animals	10	10	10	10
RBC (10^4/µL)	887 ± 31.1	885 ± 22.7	836 ± 28.4**	849 ± 42.1*
Ht (PCV) (%)	47.7 ± 1.42	47.4 ± 1.55	45.9 ± 1.75*	45.9 ± 1.64*
Hb (g/dL)	16.1 ± 0.52	16.0 ± 0.57	15.5 ± 0.64	15.4 ± 0.62*
Reticulo (%)	26 ± 3.5	24 ± 2.6	27 ± 4.0	30 ± 5.3
MCV (µm^3)	53.9 ± 1.04	53.6 ± 1.67	54.9 ± 1.55	54.1 ± 1.43
MCH (pg)	18.2 ± 0.51	18.1 ± 0.61	18.6 ± 0.64	18.2 ± 0.56
MCHC (%)	33.7 ± 0.58	33.8 ± 0.45	33.9 ± 0.40	33.6 ± 0.38
Plt (10^4/µL)	103.6 ± 14.82	101.3 ± 12.53	104.6 ± 16.55	110.7 ± 10.26
WBC (10^2/µL)	94 ± 27.8	98 ± 29.9	117 ± 27.6	114 ± 23.2
Differential Leukocyte Counts (%)
Lymphocytes	80 ± 5.4	81 ± 6.0	81 ± 8.1	78 ± 5.7
Neutrophils
Segmented	10 ± 4.4	12 ± 4.4	12 ± 7.3	16 ± 5.8
Band	0 ± 0.7	1 ± 0.8	0 ± 0.5	0 ± 0.5
Eosinophils	1 ± 1.1	1 ± 0.4	1 ± 1.1	1 ± 0.8
Basophils	0 ± 0.0	0 ± 0.0	0 ± 0.0	0 ± 0.0
Monocytes	9 ± 3.3	7 ± 3.6	6 ± 2.6	5 ± 2.9

Values are expressed as Mean ± S.D.

* Significantly different from the control group value at P<0.05

** Significantly different from the control group value at P<0.01

 

Table 2. Results of the Blood Chemistry Examination in Male Rats
Dose (mg/kg/day)	0	4	20	100
Number of animals	10	10	10	10
GOT (AsT) (IU/L)	63 ± 1.1	70 ± 10.3	63 ± 10.8	88 ± 24.8
GPT (AlT) (IU/L)	24 ± 4.2	22 ± 4.1	23 ± 5.2	45 ± 16.5
¿-GPT (IU/L)	0 ± 0.0	0 ± 0.0	1 ± 2.2	1 ± 2.5
ALP (IU/L)	285 ± 55.6	270 ± 62.5	257 ± 42.5	297 ± 47.3
Total Bilirubin (mg/dL)	0.1 ± 0.07	0.1 ± 0.06	0.1 ± 0.08	0.1 ± 0.08
Urea Nitrogen (mg/dL)	15.3 ± 2.86	16.2 ± 2.44	18.0 ± 7.98	15.6 ± 1.70
Creatinine (mg/dL)	0.5 ± 0.07	0.5 ± 0.07	0.5 ± 0.08	0.5 ± 0.06
Glucose (mg/dL)	148 ± 32.9	141 ± 8.7	139 ± 18.3	133 ± 7.6
Total Chol. (mg/dL)	80 ± 12.7	73 ± 17.0	85 ± 27.3	101 ± 21.0
Triglyceride (mg/dL)	73 ± 32.0	65 ± 33.7	68 ± 25.3	63 ± 30.8
Total Protein (g/dL)	6.64 ± 0.266	6.40 ± 0.307	6.43 ± 0.254	6.66 ± 0.34
Albumin (g/dL)	3.91 ± 0.109	3.78 ± 0.145	3.79 ± 0.076	3.81 ± 0.14
A/G Ratio	1.44 ± 0.086	1.44 ± 0.080	1.45 ± 0.123	1.34 ± 0.08
Inorganic Phos. (mg/dL)	7.6 ± 0.40	7.8 ± 0.32	8.1 ± 0.57	8.0 ± 0.44
Ca (mg/dL)	9.5 ± 0.24	9.2 ± 0.21**	9.6 ± 0.26	9.7 ± 0.10
Na (mEq/L)	144 ± 1.3	144 ± 0.8	143 ± 1.2	144 ± 1.0
K (mEq/L)	4.5 ± 0.22	4.3 ± 0.16	4.6 ± 0.63	4.6 ± 0.18
Cl (mEq/L)	103 ± 1.5	103 ± 1.8	102 ± 1.6	101 ± 0.9

Values are expressed as Mean ± S.D.

** Significantly different from the control group value at P<0.01

 

Table 3. Absolute and Relative Organ Weights
Dose (mg/kg/day)	0	4	20	100
Male Rats
Number of animals	10	10	10	10
Body weight (g)	494 ± 38.2	488 ± 26.0	487 ± 24.2	469 ± 29.2
Absolute Organ Weight
Thymus (mg)	310 ± 56.3	366 ± 79.1	348 ± 56.0	345 ± 56.0
Liver (g)	13.38 ± 1.434	13.04 ± 1.232	13.44 ± 0.851	15.27 ± 1.43
Kidneys (g)	2.85 ± 0.350	3.18 ± 0.251	3.42 ± 0.271*	3.33 ± 0.49
Adrenals (mg)	58.1 ± 6.37	59.8 ± 6.83	59.6 ± 7.32	61.5 ± 6.59
Testes (g)	3.12 ± 0.552	3.39 ± 0.287	3.05 ± 0.706	3.06 ± 0.67
Epididymides (g)	1.14 ± 0.167	1.17 ± 0.132	1.08 ± 0.222	1.10 ± 0.11
Relative Organ Weight
Thymus (mg%)	63 ± 12.1	76 ± 19.4	72 ± 11.3	74 ± 12.0
Liver (g%)	2.71 ± 0.141	2.67 ± 0.153	2.76 ± 0.149	3.25 ± 0.13
Kidneys (g%)	0.58 ± 0.057	0.65 ± 0.054	0.70 ± 0.058**	0.71 ± 0.08
Adrenals (mg%)	11.8 ± 1.17	12.3 ± 1.40	12.3 ± 1.39	13.2 ± 1.62
Testes (g%)	0.63 ± 0.103	0.70 ± 0.073	0.63 ± 0.146	0.66 ± 0.15
Epididymides (g%)	0.23 ± 0.035	0.24 ± 0.033	0.22 ± 0.048	0.23 ± 0.02
Female Rats
Number of animals	9	8	5	7
Body weight (g)	319 ± 23.6	314 ± 9.4	325 ± 24.3	298 ± 8.9
Absolute Organ Weight
Thymus (mg)	207 ± 48.8	223 ± 68.0	217 ± 62.4	205 ± 71.2
Liver (g)	13.46 ± 1.724	13.30 ± 1.640	13.56 ± 2.000	13.35 ± 1.77
Kidneys (g)	1.97 ± 0.201	1.89 ± 0.080	2.06 ± 0.201	1.96 ± 0.13
Adrenals (mg)	72.9 ± 7.17	71.0 ± 9.16	70.9 ± 3.86	71.3 ± 10.3
Relative Organ Weight
Thymus (mg%)	65 ± 16.6	71 ± 24.3	67 ± 18.3	69 ± 23.5
Liver (g%)	4.21 ± 0.326	4.22 ± 0.470	4.17 ± 0.353	4.48 ± 0.53
Kidneys (g%)	0.62 ± 0.030	0.60 ± 0.029	0.64 ± 0.017	0.66 ± 0.03
Adrenals (mg%)	22.9 ± 2.29	22.6 ± 2.95	22.0 ± 2.37	23.9 ± 3.24

Values are expressed as Mean ± S.D.

* Significantly different from the control group value at P<0.05

** Significantly different from the control group value at P<0.01

 

Table 4. Summary of Necropsy Findings
Sex	Males				Females
Dose (mg/kg/day)	0	4	20	100	0	4	20	100
Number of animals	10	10	10	10	10	10	10	8
Organ Findings
Liver
Enlargement	0	0	1	3	0	0	0	0
Kidneys
Discoloration	0	0	0	1	0	0	0	0
Whitish Dots	0	0	0	1	0	0	0	0
Enlargement	0	0	1	0	0	0	0	0
Multiple Cysts	0	0	1	0	0	0	0	0
Testes
Atrophy	1	0	2	1
Adrenals
Enlargement	0	0	0	1	0	0	0	0
Spleen
Enlargement	0	0	1	0	0	0	0	0
Subcutis
Nodule	0	0	0	0	0	0	1	0

 

Table 5. Summary of Histopathological Findings
Sex	Males				Females
Dose (mg/kg/day)	0	4	20	100	0	4	20	100
Number of animals	10	10	10	10	10	10	10	8
Organ Findings
Liver
Single cell necrosis	0	0	0	7	0	*	*	ND
Focal necrosis	0	0	0	0	1	*	*	ND
Multilocular biliary cyst	0	0	1	0	0	*	*	ND
Kidneys
Increase of hyaline droplets in the tubular epithelium	0	10	10	10	0	*	*	ND
Basophilic changes of the tubular epithelium	0	0	3	2	0	*	*	ND
Cystic Dilatation of renal tubules	1	0	0	0	0	*	*	ND
Multiple Cyst	0	0	1	0	0	*	*	ND
Testes
Atrophy of seminiferous tubules	1	*	2/2#	1
Adrenals
Increase of fatty droplets in the fascicular zone	0	0	3	8	0	0	0
Spleen
Hemosiderosis	0	*	*	0	1	*	*	ND
Subcutis
Mammary adenoma	*	*	*	*	*	*	1/1#	ND

# Number of animals showing lesion / Number of animals examined

* Not examined

ND – Value not determinable due to unclear presentation in study report table.

Applicant's summary and conclusion

Conclusions:

Dicyclopentadiene induced systemic toxicity in male and female rats including death of two females at the 100 mg/kg bw/day dose level. The NOAEL for systemic effects in female rats was determined to be 20 mg/kg bw/day. The NOAEL for systemic effects in male rats was determined to be 4 mg/kg/ bw/day.

Executive summary:

In a combined repeat dose toxicity study with reproduction/developmental toxicity screening, groups of 10 males and 10 females were dosed by oral gavage with solutions of 0, 4, 20 or 100 mg/kg bw/day dicyclopentadiene in olive oil. Animals were dosed for 2 weeks prior to mating and during mating (approximately 2 weeks). Males and females were then dosed through gestation until day 3 of lactation. Females were killed on day 4 of lactation and males were killed on day 45 of the study. Two females at 100 mg/kg bw/day died during the study and surviving males and females showed decreased food consumption and bodyweight gain at this dose level. Pathological changes in the liver and kidney were seen in males dosed at 100 mg/kg bw/day (single cell necrosis in the liver, hyaline droplet formation and basophilic changes in the tubular epithelium of the kidney) and an increase in fatty droplets in the adrenals was observed in both males and females in the 100 mg/kg bw/day group. Similar changes were seen in the kidney and adrenals of some male rats dosed at 20 mg/kg bw/day group male rats. The NOAEL for systemic toxicity was therefore considered to be 20 mg/kg bw/day for females and 4 mg/kg bw/day for male rats.