Hexachlorocyclopentadiene

Repeated dose toxicity: inhalation

Administrative data

Endpoint:

sub-chronic toxicity: inhalation

Type of information:

experimental study

Adequacy of study:

key study

Study period:

From 25 October 1983 to 27 January 1984

Reliability:

2 (reliable with restrictions)

Rationale for reliability incl. deficiencies:

comparable to guideline study with acceptable restrictions

Data source

Reference

Reference Type:

study report

Title:

Unnamed

Year:

1994

Report date:

1994

Materials and methods

Test guideline

Qualifier:

equivalent or similar to guideline

Guideline:

OECD Guideline 413 (Subchronic Inhalation Toxicity: 90-Day Study)

Deviations:

yes

Remarks:

No FOB and ophthalmological examination were performed.

GLP compliance:

yes

Limit test:

no

Test material

Test material information

Constituent 1

Reference substance name:

Hexachlorocyclopentadiene

EC Number:

201-029-3

EC Name:

Hexachlorocyclopentadiene

Cas Number:

77-47-4

Molecular formula:

C5Cl6

IUPAC Name:

hexachlorocyclopentadiene

impurity 1

Reference substance name:

Hexachlorobuta-1,3-diene

EC Number:

201-765-5

EC Name:

Hexachlorobuta-1,3-diene

Cas Number:

87-68-3

Molecular formula:

C4Cl6

IUPAC Name:

1,1,2,3,4,4-hexachlorobuta-1,3-diene

Test material form:

liquid

Test animals

Species:

rat

Strain:

Fischer 344

Sex:

male/female

Details on test animals or test system and environmental conditions:

TEST ANIMALS
- Source: Frederick Cancer Research Facility
- Females (if applicable) nulliparous and non-pregnant: yes
- Age at study initiation: 6 weeks
- Weight at study initiation: Females: 101 to 108g / Males: 118 to 127g
- Housing: individually in stainless steel cages (Hazleton Systems)
- Diet (e.g. ad libitum): NIH-07 pelleted rodent diet ad libitum
- Water (e.g. ad libitum): Tap water ad libitum
- Acclimation period: 2 weeks

DETAILS OF FOOD AND WATER QUALITY:

ENVIRONMENTAL CONDITIONS
- Temperature (°C): 20°C-21°C
- Humidity (%): 35%-65%
- Air changes (per hr): 20 changes/h
- Photoperiod (hrs dark / hrs light): 12h/12h

Administration / exposure

Route of administration:

inhalation: vapour

Type of inhalation exposure:

whole body

Vehicle:

air

Details on inhalation exposure:

GENERATION OF TEST ATMOSPHERE / CHAMBER DESCRIPTION
- Exposure apparatus: Stainless steel whole-body inhalation chamber (Hazleton Systems)
- Source and rate of air: fresh air
- System of generating particulates/aerosols: Vaporizer. Gardner Type CN condensation nuclei detector used to ensure the generation of vapour and not of aerosol.

TEST ATMOSPHERE
- Brief description of analytical method used: On-line gas chromatograph with electron capture detector.
- Samples taken from breathing zone: yes

Analytical verification of doses or concentrations:

yes

Details on analytical verification of doses or concentrations:

A gas chromatograph with an electron capture detector was used. The system was a 3% OV-225 coating on a 100/120 mesh Gas Chrom Q column and an argon/methane (9O:lO) carrier gas at a flow rate of 30 mL/minute. Column was maintained isothermally at 125°C.

Duration of treatment / exposure:

13 weeks

Frequency of treatment:

6 hours per day, 5 days per week

Doses / concentrations

Doses / concentrations 1

Dose / conc.:

0 ppm (nominal)

Remarks:

Equivalent to 0 mg/m3.

Doses / concentrations 2

Dose / conc.:

0.04 ppm (nominal)

Remarks:

Equivalent to 0.45 mg/m3.

Doses / concentrations 3

Dose / conc.:

0.15 ppm (nominal)

Remarks:

Equivalent to 1.67 mg/m3.

Doses / concentrations 4

Dose / conc.:

0.4 ppm (nominal)

Remarks:

Equivalent to 4.46 mg/m3.

Doses / concentrations 5

Dose / conc.:

1 ppm (nominal)

Remarks:

Equivalent to 11.14 mg/m3.

Doses / concentrations 6

Dose / conc.:

2 ppm (nominal)

Remarks:

Equivalent to 22.28 mg/m3.

No. of animals per sex per dose:

10 animals/sex/dose.

Control animals:

yes, concurrent no treatment

Details on study design:

- Dose selection rationale: Not specified

Positive control:

No

Examinations

Observations and examinations performed and frequency:

CAGE SIDE OBSERVATIONS: Yes
- Time schedule: Twice daily

DETAILED CLINICAL OBSERVATIONS: Yes
- Time schedule: Weekly

BODY WEIGHT: Yes
- Time schedule for examinations: At beginning, weekly, and at termination

FOOD EFFICIENCY:
- Body weight gain in kg/food consumption in kg per unit time X 100 calculated as time-weighted averages from the consumption and body weight gain data: No

WATER CONSUMPTION AND COMPOUND INTAKE (if drinking water study): No

OPHTHALMOSCOPIC EXAMINATION: No

HAEMATOLOGY: Yes
- Time schedule for collection of blood: Days 4, 16, 46 and 93
- Anaesthetic used for blood collection: Not specified
- Animals fasted: Not specified
- How many animals: All animals

CLINICAL CHEMISTRY: Yes
- Time schedule for collection of blood: Days 4, 16, 46 and 93
- Anaesthetic used for blood collection: Not specified
- Animals fasted: Not specified
- How many animals: All animals

URINALYSIS: Yes
- Time schedule for collection of urine: Days 3, 15, 45, 92
- Metabolism cages used for collection of urine: Yes
- Animals fasted: Not specified

NEUROBEHAVIOURAL EXAMINATION: No

IMMUNOLOGY: No

Sacrifice and pathology:

GROSS PATHOLOGY: Yes

HISTOPATHOLOGY: Yes

Statistics:

Statistical analyses for possible dose-related effects on survival used Cox’s (1972) method for testing two groups for equality and Tarone’s (1975) life table test to identify dose-related trends.
The probability of survival was estimated by the product-limit procedure of Kaplan and Meier (1958).

Results and discussion

Results of examinations

Clinical signs:

effects observed, treatment-related

Description (incidence and severity):

Listlessness was observed in rats exposed to 0.4, 1 and 2 ppm. Respiratory distress was observed in rats exposed to 1 and 2 ppm.

Mortality:

mortality observed, treatment-related

Description (incidence):

All rats exposed to 1 or 2 ppm of test substance died in the first 4 weeks of exposure. All rats exposed to lower concentrations survived.

Body weight and weight changes:

effects observed, treatment-related

Description (incidence and severity):

Final mean body weight and mean body weight gain of male rats exposed to 0.4 ppm were significantly lower than those of the control animals.

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:

effects observed, non-treatment-related

Description (incidence and severity):

Statistically significant differences in haematology parameters were observed but were not attributed to the test substance as they were not persistent nor dose-related.

Clinical biochemistry findings:

effects observed, non-treatment-related

Description (incidence and severity):

Statistically significant differences in clinical biochemistry parameters were observed but were not attributed to the test substance as they were not persistent nor dose-related.

Urinalysis findings:

effects observed, non-treatment-related

Description (incidence and severity):

Statistically significant differences in urinalysis parameters were observed but were not attributed to the test substance as they were not persistent nor dose-related.

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):

Lung weights of male rats exposed to 0.4 ppm of test substance were significantly greater than those of the controls. Other differences in organ weights were likely related to the lower body weights observed in exposed rats.

Gross pathological findings:

effects observed, treatment-related

Description (incidence and severity):

In rats exposed to 1 and 2 ppm of test substance, an extensive coagulation necrosis of the respiratory epithellium of the nose, larynx, trachea, bronchi, and bronchioles was observed.

Neuropathological findings:

not examined

Histopathological findings: non-neoplastic:

effects observed, treatment-related

Description (incidence and severity):

In rats exposed to 1 and 2 ppm of test substance, acute and subacute inflammation was observed in the respiratory tract consisting of vascular congestion, edema, accumulation of fibrin, and infiltrates of neutrophils and mononuclear cells. In some animals fibrinosuppurative exudate and suppurative alveolar inflammation were observed.
In rats exposed to 0.4 ppm, focal or multifocal suppurative inflammation of the nose or lung was observed (especially in male rats). Focal squamous metaplasia was observed in some male rats exposed to 0.4 ppm and some male and female rats exposed to 1 and 2 ppm.

Histopathological findings: neoplastic:

no effects observed

Effect levels

1

Key result

Dose descriptor:

NOAEC

Remarks:

systemic

Effect level:

1.67 mg/m³ air (nominal)

Based on:

test mat.

Sex:

male/female

Basis for effect level:

body weight and weight gain

clinical signs

mortality

2

Dose descriptor:

NOAEC

Remarks:

local

Effect level:

1.67 mg/m³ air (nominal)

Based on:

test mat.

Sex:

male/female

Basis for effect level:

gross pathology

histopathology: non-neoplastic

organ weights and organ / body weight ratios

Target system / organ toxicity

1

Key result

Critical effects observed:

yes

Lowest effective dose / conc.:

4.46 mg/m³ air (nominal)

System:

respiratory system: upper respiratory tract

Organ:

larynx

nasal cavity

trachea

Treatment related:

yes

Dose response relationship:

yes

Relevant for humans:

not specified

2

Critical effects observed:

yes

Lowest effective dose / conc.:

4.46 mg/m³ air (nominal)

System:

respiratory system: lower respiratory tract

Organ:

bronchi

bronchioles

lungs

Treatment related:

yes

Dose response relationship:

yes

Relevant for humans:

not specified

Applicant's summary and conclusion

Conclusions:

A subchronic toxicity study by inhalation was performed on rats using Hexachlorocyclopentadiene that allowed to determine NOAEC of 0.15 ppm (equivalent to 1.67 mg/m3) for both local and systemic effects based on mortality, body weight, and effects on upper and lower respiratory tract.

Executive summary:

The repeated dose toxicity of Hexachlorocyclopentadiene by inhalation was evaluated using a method similar to the OECD Test Guideline 413 (GLP) with deviations.

Rats were exposed at concentrations of 0, 0.04, 0.15, 0.4, 1 and 2 ppm of Hexachlorocyclopentadiene in air(equivalent to 0, 0.45, 1.67, 4.46, 11.14 and 22.28 mg/m3) for 6 hours per day, 5 days per week, for 13 weeks. Mortality, clinical signs, body weight, and haematology, clinical biochemistry, and urinalysis parameters were recorded. At the end of the study, surviving animals were subjected to gross necropsy and histopathology. All rats exposed to 1 or 2 ppm of test substance died in the first 4 weeks of exposure. All rats exposed to lower concentrations survived. Listlessness was observed in rats exposed to 0.4, 1 and 2 ppm. Respiratory distress was observed in rats exposed to 1 and 2 ppm. Final mean body weight and mean body weight gain of male rats exposed to 0.4 ppm were significantly lower than those of the control animals. Lung weights of male rats exposed to 0.4 ppm of test substance were significantly greater than those of the controls. Other differences in organ weights were likely related to the lower body weights observed in exposed rats. In rats exposed to 1 and 2 ppm of test substance, an extensive coagulation necrosis of the respiratory epithellium of the nose, larynx, trachea, bronchi, and bronchioles was observed. In rats exposed to 1 and 2 ppm of test substance, acute and subacute inflammation was observed consisting of vascular congestion, edema, accumulation of fibrin, and infiltrates of neutrophils and mononuclear cells. In some animals fibrinosuppurative exudate and suppurative alveolar inflammation were observed. In rats exposed to 0.4 ppm, focal or multifocal suppurative inflammation of the nose or lung was observed (especially in male rats). Focal squamous metaplasia was observed in some male rats exposed to 0.4 ppm and some male and female rats exposed to 1 and 2 ppm. Statistically significant differences in haematology, clinical biochemestry, and urinalysis parameters were observed but were not attributed to the test substance as they were not persistent nor dose-related.

This subchronic toxicity study by inhalation allowed to determine a NOAEC of 0.15 ppm (equivalent to 1.67 mg/m3) for both local and systemic effects based on mortality, body weight, and upper and lower respiratory tract.