Hexachlorocyclopentadiene

Endpoint summary

Description of key information

The repeated dose toxicity of Hexachlorocyclopentadiene by the oral route was evaluated using a method similar to the OECD Test Guideline 408 (non-GLP). Rats received doses of 0, 10, 19, 38, 75 or 150 of Hexachlorocyclopentadiene by gavage once a day, 5 days per week, for 13 weeks. This protocol deviated from the current version of the OECD Test Guideline 408 but was designed to mimic the exposure of workers and is therefore considered as acceptable.

Six male rats receiving 150 mg/kg bw/day and one female rats receiving 75 mg/kg bw/day died as a result of the treatment. Other deaths occurred that were attributed to improper gavage technique. Treatment related clinical signs were observed in animals treated at 150 mg/kg bw/day. A significant reduction of the bodyweight was observed in male rats receiving 38 mg/kg bw/day or above and in female rats receiving 75 mg/kg bw/day or above. Significant changes in the relative weight of several organs were observed in relation to the reduction of the bodyweight. In male rats receiving 75 mg/kg bw/day and above and female rats receiving 38 mg/kg bw/day and above, effects in stomach including mass and black areas were observed and considered treatment related. Animals exposed to 38 mg/kg bw/day and above exhibited acute kidney tubular necrosis. In male rats receiving 38 mg/kg bw/day or above and female rats receiving 19 mg/kg bw/day, significant proliferation and inflammatory changes of the epithelia in the stomach were observed. This subchronic toxicity study by the oral route allowed to determine a NOAEL(systemic) of 10 mg/kg bw/day for both male and female rats, a NOAEL(local) of 10 mg/kg bw/day for female rats, and a NOAEL(local) of 19 mg/kg bw/day for male rats.

 

Several studies were performed investigating the effects of Hexachlorocyclopentadiene following a repeated exposure by inhalation. The subchronic toxicity study performed by the US NTP (1994) on rats was considered as the key study as it was performed on a standard species in compliance with the GLP. Chronic toxicity studies performed by inhalation were available, but they were considered as less reliable for the purpose of the assessment of the repeated-dose toxicity of Hexachlorocyclopentadiene as their purpose was to identify carcinogenic effects and several relevant examinations were omitted.

The US NTP (1994) used a method similar to the OECD Test Guideline 413 (GLP). Rats were exposed at concentrations of 0, 0.04, 0.15, 0.4, 1 or 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. All rats exposed to 1 or 2 ppm of the test substance died in the first 4 weeks of exposure and treatment related clinical signs considered as adverse effects were observed before their deaths, including respiratory distress, extensive coagulation necrosis of the respiratory epithelium, and acute and subacute inflammation. All rats exposed to lower concentrations survived. Animals exposed to 0.4 ppm showed listlessness and suppurative inflammation of the nose or lung (especially in male rats). 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. 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 inflammation of the upper and lower respiratory tract.

 

The repeated dose toxicity of Hexachlorocyclopentadiene by the dermal route was evaluated using a method similar to the OECD Test Guideline 410 (non-GLP). Rabbits received applications of 0.1 or 5 mg/kg bw of Hexachlorocyclopentadiene once a day, 5 days per week, for 4 weeks. No mortality was observed as a result of this study. No systemic effects including significant variation of the body weight of treated animals were observed. The test substance was severely irritating to the skin upon repeated dermal applications at the test concentrations. These findings were confirmed during the gross pathology and histopathology examination that identified various alterations of the skin. This short-term toxicity study by the dermal route performed on the registered substance allowed to determine a NOAEL(systemic) of 5 mg/kg bw/day and a LOAEL(local) of 1 mg/kg bw/day.

Key value for chemical safety assessment

Repeated dose toxicity: via oral route - systemic effects

Link to relevant study records

Reference

Endpoint:

sub-chronic toxicity: oral

Type of information:

experimental study

Adequacy of study:

key study

Study period:

1981

Reliability:

2 (reliable with restrictions)

Rationale for reliability incl. deficiencies:

test procedure in accordance with national standard methods with acceptable restrictions

Qualifier:

equivalent or similar to guideline

Guideline:

OECD Guideline 408 (Repeated Dose 90-Day Oral Toxicity Study in Rodents)

Deviations:

yes

Remarks:

Exposure 5 days per week. FOB and ophthalmological examination were not performed. Urinalysis, haematology, or clinical biochemistry were not examined

GLP compliance:

no

Limit test:

no

Species:

rat

Strain:

Fischer 344

Sex:

male/female

Details on test animals or test system and environmental conditions:

TEST ANIMALS
- Source: Charles River Breeding Laboratory
- Females (if applicable) nulliparous and non-pregnant: yes
- Age at study initiation: 52 days
- Weight at study initiation: Females: 99 - 135 g. Males: 130 - 170 g.
- Housing: Five animals per cage in polycarbonate cages.
- Diet (e.g. ad libitum): ad libitum
- Water (e.g. ad libitum): ad libitum
- Acclimation period: 20 days

ENVIRONMENTAL CONDITIONS
- Temperature (°C): 72°F to 78°F (22°C to 25°C)
- Humidity (%): 23% to 58%
- Air changes (per hr): 15 changes per hour
- Photoperiod (hrs dark / hrs light): 12h/12h

Route of administration:

oral: gavage

Vehicle:

corn oil

Details on oral exposure:

VEHICLE
- Lot/batch no.: 1-13-82A
- Amount of vehicle (if gavage): 0.5ml/100g
- Purity: 100%

Analytical verification of doses or concentrations:

yes

Details on analytical verification of doses or concentrations:

Dosage analysis for concentration verification was performed in duplicate on each dose level for the first set of mixings and a set approximately midway through the study.

Duration of treatment / exposure:

13 weeks

Frequency of treatment:

Once a day, 5 days per week. The frequency was selected to mimic the exposure of workers.

Dose / conc.:

0 mg/kg bw/day (nominal)

Dose / conc.:

10 mg/kg bw/day (nominal)

Dose / conc.:

19 mg/kg bw/day (nominal)

Dose / conc.:

38 mg/kg bw/day (nominal)

Dose / conc.:

75 mg/kg bw/day (nominal)

Dose / conc.:

150 mg/kg bw/day (nominal)

No. of animals per sex per dose:

10 animals/sex/dose

Control animals:

yes, concurrent vehicle

Details on study design:

not specified

Positive control:

No

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: Initially, 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: Not specified

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

OPHTHALMOSCOPIC EXAMINATION: No

HAEMATOLOGY: No

CLINICAL CHEMISTRY: No

URINALYSIS: No

NEUROBEHAVIOURAL EXAMINATION: No

IMMUNOLOGY: No

Sacrifice and pathology:

GROSS PATHOLOGY: Yes

HISTOPATHOLOGY: Yes

Statistics:

Not specified

Clinical signs:

effects observed, treatment-related

Description (incidence and severity):

Animals treated at 150 mg/kg bw/day displayed a ruffled fur and a slight inactivity that were considered treatment-related. Other clinical observations were attributed to the housing of animals collectively or were not considered as significant due to their low incidence.

Mortality:

mortality observed, treatment-related

Description (incidence):

6/10 male rats receiving 150 mg/kg bw/day died as a result of the treatment. 1/10 female rats receiving 75 mg/kg bw/day died as a result of the treatment. Other deaths occurred in male and female rats at different doses that were attributed to improper gavage technique.

Body weight and weight changes:

effects observed, treatment-related

Description (incidence and severity):

A significant reduction of the bodyweight was observed in male rats receiving 38 mg/kg bw/day or above. A reduction was also observed in male rats receiving 10 and 19 mg/kg bw/day.
A significant reduction of the bodyweight was observed in female rats receiving 75 mg/kg bw/day or above. A reduction was also observed in male rats receiving 10, 19 and 75 mg/kg bw/day.

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:

not examined

Clinical biochemistry 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):

In male rats, the increase in relative liver weight at the dose levels of 10, 19, 38, and 75 mg/kg bw was -23% (decrease), 5%, 18%, and 31% respectively. The increase in relative kidney weight was -5% (decrease), 13%, 23% and 39% respectively.
In female rats, the increase in relative liver weight at the dose levels of 10, 19, 38, and 75 mg/kg bw was 17%, 4%, 19%, and 33% respectively. The increase in relative kidney weight was 10%, 12%, 30% and 62% respectively.
Relative organ weights could not be calculated at 150 mg/kg bw/day due to the deaths occurring at this dose level.

Gross pathological findings:

effects observed, treatment-related

Description (incidence and severity):

In male rats receiving 75 mg/kg bw/day and above and female rats receiving 38 mg/kg bw/day and above, effects in stomach including mass and black areas were observed and considered treatment-related.
Other gross pathological findings were not considered significant or treatment-related.

Neuropathological findings:

not examined

Histopathological findings: non-neoplastic:

effects observed, treatment-related

Description (incidence and severity):

Animals exposed to 38 mg/kg bw/day and above exhibited acute kidney tubular necrosis characterised by vacuolisation, cytomegaly, karyomegaly, anisokaryosis, and tubular dilation.
In male rats receiving 38 mg/kg bw/day or above and female rats receiving 19 mg/kg bw/day, significant proliferation and inflammatory changes of the epithelia in the stomach were observed.

Histopathological findings: neoplastic:

no effects observed

Details on results:

A NOAEL(systemic) of 10 mg/kg bw/day can be derived for both male and female rats.
A NOAEL(local) of 10 mg/kg bw/day can be derived for female rats.
A NOAEL(local) of 19 mg/kg bw/day can be derived for male rats.

Key result

Dose descriptor:

NOAEL

Remarks:

systemic

Effect level:

10 mg/kg bw/day (nominal)

Based on:

test mat.

Sex:

male/female

Basis for effect level:

body weight and weight gain

clinical signs

gross pathology

histopathology: non-neoplastic

mortality

organ weights and organ / body weight ratios

Dose descriptor:

NOAEL

Remarks:

local

Effect level:

10 mg/kg bw/day (nominal)

Based on:

test mat.

Sex:

female

Basis for effect level:

histopathology: non-neoplastic

Dose descriptor:

NOAEL

Remarks:

local

Effect level:

19 mg/kg bw/day (nominal)

Based on:

test mat.

Sex:

male

Basis for effect level:

histopathology: non-neoplastic

Critical effects observed:

yes

Lowest effective dose / conc.:

19 mg/kg bw/day (nominal)

System:

urinary

Organ:

kidney

Treatment related:

yes

Dose response relationship:

yes

Relevant for humans:

not specified

Critical effects observed:

yes

Lowest effective dose / conc.:

19 mg/kg bw/day (nominal)

System:

gastrointestinal tract

Organ:

stomach

Treatment related:

yes

Dose response relationship:

yes

Relevant for humans:

not specified

Conclusions:

A subchronic toxicity study by the oral route was performed on rats using Hexachlorocyclopentadiene that allowed to determine a NOAEL(systemic) of 10 mg/kg bw/day for both male and female rats, a NOAEL(local) of 10 mg/kg bw/day for female rats, and a NOAEL(local) of 19 mg/kg bw/day for male rats.

Executive summary:

The repeated dose toxicity of Hexachlorocyclopentadiene by the oral route was evaluated using a method similar to the OECD Test Guideline 408 (non-GLP) with deviations.

Rats received doses of 0, 10, 19, 38, 75 and 150 of Hexachlorocyclopentadiene by gavage once a day, 5 days per week, for 13 weeks. The protocol was determined to mimic the exposure of workers. Mortality, clinical signs and body weight were recorded. At the end of the study, surviving animals were subjected to gross necropsy and histopathology.

Six male rats receiving 150 mg/kg bw/day and one female rats receiving 75 mg/kg bw/day died as a result of the treatment. Other deaths occurred that were attributed to improper gavage technique. Animals treated at 150 mg/kg bw/day displayed a ruffled fur and a slight inactivity that were considered treatment-related. Other clinical observations were attributed to the housing of animals collectively or were not considered as significant due to their low incidence.

A significant reduction of the bodyweight was observed in male rats receiving 38 mg/kg bw/day or above. A reduction was also observed in male rats receiving 10 and 19 mg/kg bw/day. A significant reduction of the bodyweight was observed in female rats receiving 75 mg/kg bw/day or above. A reduction was also observed in male rats receiving 10, 19 and 75 mg/kg bw/day.

In male rats, the increase in relative liver weight at the dose levels of 10, 19, 38, and 75 mg/kg bw was -23% (decrease), 5%, 18%, and 31% respectively. The increase in relative kidney weight was -5% (decrease), 13%, 23% and 39% respectively. In female rats, the increase in relative liver weight at the dose levels of 10, 19, 38 and 75 mg/kg bw was 17%, 4%, 19% and 33% respectively. The increase in relative kidney weight was 10%, 12%, 30% and 62% respectively.

In male rats receiving 75 mg/kg bw/day and above and female rats receiving 38 mg/kg bw/day and above, effects in stomach including mass and black areas were observed and considered treatment-related. Other gross pathological findings were not considered significant or treatment-related.

Animals exposed to 38 mg/kg bw/day and above exhibited acute kidney tubular necrosis characterised by vacuolisation, cytomegaly, karyomegaly, anisokaryosis, and tubular dilation. In male rats receiving 38 mg/kg bw/day or above and female rats receiving 19 mg/kg bw/day, significant proliferation and inflammatory changes of the epithelia in the stomach were observed.

This subchronic toxicity study by the oral route allowed to determine a NOAEL(systemic) of 10 mg/kg bw/day for both male and female rats, a NOAEL(local) of 10 mg/kg bw/day for female rats, and a NOAEL(local) of 19 mg/kg bw/day for male rats.

Endpoint conclusion

Endpoint conclusion:

adverse effect observed

Dose descriptor:

NOAEL

10 mg/kg bw/day

Study duration:

subchronic

Species:

rat

Quality of whole database:

test procedure in accordance with national standard methods with acceptable restrictions

Organ:

kidney

stomach

Repeated dose toxicity: inhalation - systemic effects

Link to relevant study records

Reference

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

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

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

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

Dose / conc.:

0 ppm (nominal)

Remarks:

Equivalent to 0 mg/m3.

Dose / conc.:

0.04 ppm (nominal)

Remarks:

Equivalent to 0.45 mg/m3.

Dose / conc.:

0.15 ppm (nominal)

Remarks:

Equivalent to 1.67 mg/m3.

Dose / conc.:

0.4 ppm (nominal)

Remarks:

Equivalent to 4.46 mg/m3.

Dose / conc.:

1 ppm (nominal)

Remarks:

Equivalent to 11.14 mg/m3.

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

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).

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

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

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

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

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

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.

Endpoint conclusion

Endpoint conclusion:

adverse effect observed

Dose descriptor:

NOAEC

1.67 mg/m³

Study duration:

subchronic

Species:

rat

Quality of whole database:

Comparable to guideline study with acceptable restrictions. Chronic studies available on the substance were lacking several information and were therefore not selected as the key studies.

Repeated dose toxicity: inhalation - local effects

Link to relevant study records

Reference

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

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

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

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

Dose / conc.:

0 ppm (nominal)

Remarks:

Equivalent to 0 mg/m3.

Dose / conc.:

0.04 ppm (nominal)

Remarks:

Equivalent to 0.45 mg/m3.

Dose / conc.:

0.15 ppm (nominal)

Remarks:

Equivalent to 1.67 mg/m3.

Dose / conc.:

0.4 ppm (nominal)

Remarks:

Equivalent to 4.46 mg/m3.

Dose / conc.:

1 ppm (nominal)

Remarks:

Equivalent to 11.14 mg/m3.

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

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).

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

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

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

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

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

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.

Endpoint conclusion

Endpoint conclusion:

adverse effect observed

Dose descriptor:

NOAEC

1.67 mg/m³

Study duration:

subchronic

Species:

rat

Quality of whole database:

Comparable to guideline study with acceptable restrictions. Chronic studies available on the substance were lacking several information and were therefore not selected as the key studies.

Repeated dose toxicity: dermal - systemic effects

Link to relevant study records

Reference

Endpoint:

short-term repeated dose toxicity: dermal

Type of information:

experimental study

Adequacy of study:

key study

Reliability:

2 (reliable with restrictions)

Rationale for reliability incl. deficiencies:

study well documented, meets generally accepted scientific principles, acceptable for assessment

Qualifier:

equivalent or similar to guideline

Guideline:

OECD Guideline 410 (Repeated Dose Dermal Toxicity: 21/28-Day Study)

Deviations:

yes

Remarks:

Two animals per sex and per dose had their skin abraded. No indication if the substance was held in contact with the skin.

GLP compliance:

no

Limit test:

no

Species:

rabbit

Strain:

New Zealand White

Sex:

male/female

Details on test animals or test system and environmental conditions:

TEST ANIMALS
- Source: not specified
- Females (if applicable) nulliparous and non-pregnant: not specified
- Age at study initiation: adult
- Weight at study initiation: From 2 to 3 kg
- Housing: individually
- Diet (e.g. ad libitum): ad libitum
- Water (e.g. ad libitum): ad libitum
- Acclimation period: not specified

ENVIRONMENTAL CONDITIONS
- Temperature (°C): not specified
- Humidity (%): not specified
- Air changes (per hr): not specified
- Photoperiod (hrs dark / hrs light): not specified

Type of coverage:

not specified

Vehicle:

ethanol

Details on exposure:

TEST SITE
- Area of exposure: back
- % coverage: 20
- Type of wrap if used: not specified
- Time intervals for shavings or clipplings: one week but more often if necessary

REMOVAL OF TEST SUBSTANCE
- Washing (if done): not specified
- Time after start of exposure: not specified

TEST MATERIAL
- Amount(s) applied (volume or weight with unit): 1 mL/kg
- Concentration (if solution): 0.1% w/v or 0.5% w/v
- Constant volume or concentration used: yes

VEHICLE
- Justification for use and choice of vehicle (if other than water): not specified
- Purity: not specified

USE OF RESTRAINERS FOR PREVENTING INGESTION: yes

Analytical verification of doses or concentrations:

not specified

Duration of treatment / exposure:

4 weeks

Frequency of treatment:

Once a day, 5 days a week.

Dose / conc.:

0 mg/kg bw/day (nominal)

Dose / conc.:

1 mg/kg bw/day (nominal)

Dose / conc.:

5 mg/kg bw/day (nominal)

No. of animals per sex per dose:

5 animals/sex/dose (including 2 animals/sex/dose with abraded skin).

Control animals:

yes, concurrent no treatment

Observations and examinations performed and frequency:

CAGE SIDE OBSERVATIONS: Yes
- Time schedule: Daily

DETAILED CLINICAL OBSERVATIONS: Yes
- Time schedule: Daily

DERMAL IRRITATION (if dermal study): Yes
- Time schedule for examinations: Daily

BODY WEIGHT: Yes
- Time schedule for examinations: Weekly

FOOD CONSUMPTION:
- Food consumption for each animal determined and mean daily diet consumption calculated as g food/kg body weight/day: No

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: No

OPHTHALMOSCOPIC EXAMINATION: No

HAEMATOLOGY: Yes
- Time schedule for collection of blood: Prior to the start of the study and at Day 23 of the study
- Anaesthetic used for blood collection: No data
- Animals fasted: Yes
- How many animals: All animals

CLINICAL CHEMISTRY: Yes
- Time schedule for collection of blood: Prior to the start of the study and at Day 23 of the study
- Animals fasted: Yes
- How many animals: All animals

URINALYSIS: Yes
- Time schedule for collection of urine: At Day 23 of the study
- Metabolism cages used for collection of urine: Yes
- Animals fasted: Yes

NEUROBEHAVIOURAL EXAMINATION: No

Sacrifice and pathology:

GROSS PATHOLOGY: Yes

HISTOPATHOLOGY: Yes

Statistics:

Not specified

Clinical signs:

no effects observed

Dermal irritation:

effects observed, treatment-related

Description (incidence and severity):

The test substance was severely irritating to the skin upon repeated dermal application at both concentration tested.

Mortality:

no mortality observed

Body weight and weight changes:

effects observed, treatment-related

Description (incidence and severity):

Non significant body weight losses were noted in some animals treated at 5 mg/kg bw/d.

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:

no effects observed

Behaviour (functional findings):

not examined

Immunological findings:

not examined

Organ weight findings including organ / body weight ratios:

effects observed, non-treatment-related

Description (incidence and severity):

Statically significant organ weight differences were observed between the groups but the results were within the historical data of the laboratory for rabbits of this strain and age. These differences were attributed to grouping and the small number of animals.

Gross pathological findings:

effects observed, treatment-related

Description (incidence and severity):

Gross skin changes were observed consisting of fibrosis, escharosis and slight to severe desquamation for both treated groups. No systemic effects were observed.

Neuropathological findings:

not examined

Histopathological findings: non-neoplastic:

effects observed, treatment-related

Description (incidence and severity):

Microscopic examination of the test skin sites revealed acanthosis and hyperkeratosis involving the epidermis in a few animals treated at 1 mg/kg bw/d and most animals treated at 5 mg/kg bw/d. An associated inflammation of the dermis escharosis formation, epidermal ulceration or microabscessation were also observed in some animals. No systemic effects were observed.

Histopathological findings: neoplastic:

no effects observed

Other effects:

no effects observed

Key result

Dose descriptor:

NOAEL

Remarks:

systemic

Effect level:

> 5 mg/kg bw/day (nominal)

Based on:

test mat.

Sex:

male/female

Basis for effect level:

body weight and weight gain

clinical biochemistry

clinical signs

gross pathology

haematology

histopathology: neoplastic

histopathology: non-neoplastic

mortality

organ weights and organ / body weight ratios

urinalysis

Key result

Dose descriptor:

LOAEL

Remarks:

local

Effect level:

1 mg/kg bw/day (nominal)

Based on:

test mat.

Sex:

male/female

Basis for effect level:

clinical signs

dermal irritation

gross pathology

histopathology: non-neoplastic

Key result

Critical effects observed:

yes

Lowest effective dose / conc.:

1 mg/kg bw/day (nominal)

System:

integumentary

Organ:

skin

Treatment related:

yes

Dose response relationship:

yes

Relevant for humans:

yes

Conclusions:

This short-term toxicity study by the dermal route performed on the registered substance allowed to determine a NOAEL(systemic) of 5 mg/kg bw/day and a LOAEL(local) of 1 mg/kg bw/day. The skin was the only identified target organ.

Executive summary:

The repeated dose toxicity of Hexachlorocyclopentadiene by the dermal was evaluated using a method similar to the OECD Test Guideline 410 (non-GLP) with deviations.

Rabbits received applications of 0, 1 or 5 mg/kg bw of Hexachlorocyclopentadiene once a day, 5 days per week, for 4 weeks. Mortality, clinical signs and body weight were recorded. Haematology, blood chemistry and urinalysis were investigated. At the end of the study, surviving animals were subjected to gross necropsy and histopathology.

No mortality was observed as a result of this study. No systemic effects including significant variation of the body weight of treated animals were observed.

The test substance was severely irritating to the skin upon repeated dermal application at both concentrations tested. These findings were confirmed during the gross pathology and histopathology examination. Gross skin changes were observed consisting of fibrosis, escharosis and slight to severe desquamation for both treated groups. Microscopic examination of the test skin sites revealed acanthosis and hyperkeratosis involving the epidermis in a few animals treated at 1 mg/kg bw/d and most animals treated at 5 mg/kg bw/d. An associated inflammation of the dermis escharosis formation, epidermal ulceration or microabscessation were also observed in some animals

There was no change in the blood and urine parameters at the end of the study.

This short-term toxicity study by the dermal route performed on the registered substance allowed to determine a NOAEL(systemic) of 5 mg/kg bw/day and a LOAEL(local) of 1 mg/kg bw/day. The skin was the only identified target organ.

Endpoint conclusion

Endpoint conclusion:

no adverse effect observed

Study duration:

subacute

Species:

rabbit

Quality of whole database:

study well documented, meets generally accepted scientific principles, acceptable for assessment

Repeated dose toxicity: dermal - local effects

Link to relevant study records

Reference

Endpoint:

short-term repeated dose toxicity: dermal

Type of information:

experimental study

Adequacy of study:

key study

Reliability:

2 (reliable with restrictions)

Rationale for reliability incl. deficiencies:

study well documented, meets generally accepted scientific principles, acceptable for assessment

Qualifier:

equivalent or similar to guideline

Guideline:

OECD Guideline 410 (Repeated Dose Dermal Toxicity: 21/28-Day Study)

Deviations:

yes

Remarks:

Two animals per sex and per dose had their skin abraded. No indication if the substance was held in contact with the skin.

GLP compliance:

no

Limit test:

no

Species:

rabbit

Strain:

New Zealand White

Sex:

male/female

Details on test animals or test system and environmental conditions:

TEST ANIMALS
- Source: not specified
- Females (if applicable) nulliparous and non-pregnant: not specified
- Age at study initiation: adult
- Weight at study initiation: From 2 to 3 kg
- Housing: individually
- Diet (e.g. ad libitum): ad libitum
- Water (e.g. ad libitum): ad libitum
- Acclimation period: not specified

ENVIRONMENTAL CONDITIONS
- Temperature (°C): not specified
- Humidity (%): not specified
- Air changes (per hr): not specified
- Photoperiod (hrs dark / hrs light): not specified

Type of coverage:

not specified

Vehicle:

ethanol

Details on exposure:

TEST SITE
- Area of exposure: back
- % coverage: 20
- Type of wrap if used: not specified
- Time intervals for shavings or clipplings: one week but more often if necessary

REMOVAL OF TEST SUBSTANCE
- Washing (if done): not specified
- Time after start of exposure: not specified

TEST MATERIAL
- Amount(s) applied (volume or weight with unit): 1 mL/kg
- Concentration (if solution): 0.1% w/v or 0.5% w/v
- Constant volume or concentration used: yes

VEHICLE
- Justification for use and choice of vehicle (if other than water): not specified
- Purity: not specified

USE OF RESTRAINERS FOR PREVENTING INGESTION: yes

Analytical verification of doses or concentrations:

not specified

Duration of treatment / exposure:

4 weeks

Frequency of treatment:

Once a day, 5 days a week.

Dose / conc.:

0 mg/kg bw/day (nominal)

Dose / conc.:

1 mg/kg bw/day (nominal)

Dose / conc.:

5 mg/kg bw/day (nominal)

No. of animals per sex per dose:

5 animals/sex/dose (including 2 animals/sex/dose with abraded skin).

Control animals:

yes, concurrent no treatment

Observations and examinations performed and frequency:

CAGE SIDE OBSERVATIONS: Yes
- Time schedule: Daily

DETAILED CLINICAL OBSERVATIONS: Yes
- Time schedule: Daily

DERMAL IRRITATION (if dermal study): Yes
- Time schedule for examinations: Daily

BODY WEIGHT: Yes
- Time schedule for examinations: Weekly

FOOD CONSUMPTION:
- Food consumption for each animal determined and mean daily diet consumption calculated as g food/kg body weight/day: No

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: No

OPHTHALMOSCOPIC EXAMINATION: No

HAEMATOLOGY: Yes
- Time schedule for collection of blood: Prior to the start of the study and at Day 23 of the study
- Anaesthetic used for blood collection: No data
- Animals fasted: Yes
- How many animals: All animals

CLINICAL CHEMISTRY: Yes
- Time schedule for collection of blood: Prior to the start of the study and at Day 23 of the study
- Animals fasted: Yes
- How many animals: All animals

URINALYSIS: Yes
- Time schedule for collection of urine: At Day 23 of the study
- Metabolism cages used for collection of urine: Yes
- Animals fasted: Yes

NEUROBEHAVIOURAL EXAMINATION: No

Sacrifice and pathology:

GROSS PATHOLOGY: Yes

HISTOPATHOLOGY: Yes

Statistics:

Not specified

Clinical signs:

no effects observed

Dermal irritation:

effects observed, treatment-related

Description (incidence and severity):

The test substance was severely irritating to the skin upon repeated dermal application at both concentration tested.

Mortality:

no mortality observed

Body weight and weight changes:

effects observed, treatment-related

Description (incidence and severity):

Non significant body weight losses were noted in some animals treated at 5 mg/kg bw/d.

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:

no effects observed

Behaviour (functional findings):

not examined

Immunological findings:

not examined

Organ weight findings including organ / body weight ratios:

effects observed, non-treatment-related

Description (incidence and severity):

Statically significant organ weight differences were observed between the groups but the results were within the historical data of the laboratory for rabbits of this strain and age. These differences were attributed to grouping and the small number of animals.

Gross pathological findings:

effects observed, treatment-related

Description (incidence and severity):

Gross skin changes were observed consisting of fibrosis, escharosis and slight to severe desquamation for both treated groups. No systemic effects were observed.

Neuropathological findings:

not examined

Histopathological findings: non-neoplastic:

effects observed, treatment-related

Description (incidence and severity):

Microscopic examination of the test skin sites revealed acanthosis and hyperkeratosis involving the epidermis in a few animals treated at 1 mg/kg bw/d and most animals treated at 5 mg/kg bw/d. An associated inflammation of the dermis escharosis formation, epidermal ulceration or microabscessation were also observed in some animals. No systemic effects were observed.

Histopathological findings: neoplastic:

no effects observed

Other effects:

no effects observed

Key result

Dose descriptor:

NOAEL

Remarks:

systemic

Effect level:

> 5 mg/kg bw/day (nominal)

Based on:

test mat.

Sex:

male/female

Basis for effect level:

body weight and weight gain

clinical biochemistry

clinical signs

gross pathology

haematology

histopathology: neoplastic

histopathology: non-neoplastic

mortality

organ weights and organ / body weight ratios

urinalysis

Key result

Dose descriptor:

LOAEL

Remarks:

local

Effect level:

1 mg/kg bw/day (nominal)

Based on:

test mat.

Sex:

male/female

Basis for effect level:

clinical signs

dermal irritation

gross pathology

histopathology: non-neoplastic

Key result

Critical effects observed:

yes

Lowest effective dose / conc.:

1 mg/kg bw/day (nominal)

System:

integumentary

Organ:

skin

Treatment related:

yes

Dose response relationship:

yes

Relevant for humans:

yes

Conclusions:

This short-term toxicity study by the dermal route performed on the registered substance allowed to determine a NOAEL(systemic) of 5 mg/kg bw/day and a LOAEL(local) of 1 mg/kg bw/day. The skin was the only identified target organ.

Executive summary:

The repeated dose toxicity of Hexachlorocyclopentadiene by the dermal was evaluated using a method similar to the OECD Test Guideline 410 (non-GLP) with deviations.

Rabbits received applications of 0, 1 or 5 mg/kg bw of Hexachlorocyclopentadiene once a day, 5 days per week, for 4 weeks. Mortality, clinical signs and body weight were recorded. Haematology, blood chemistry and urinalysis were investigated. At the end of the study, surviving animals were subjected to gross necropsy and histopathology.

No mortality was observed as a result of this study. No systemic effects including significant variation of the body weight of treated animals were observed.

The test substance was severely irritating to the skin upon repeated dermal application at both concentrations tested. These findings were confirmed during the gross pathology and histopathology examination. Gross skin changes were observed consisting of fibrosis, escharosis and slight to severe desquamation for both treated groups. Microscopic examination of the test skin sites revealed acanthosis and hyperkeratosis involving the epidermis in a few animals treated at 1 mg/kg bw/d and most animals treated at 5 mg/kg bw/d. An associated inflammation of the dermis escharosis formation, epidermal ulceration or microabscessation were also observed in some animals

There was no change in the blood and urine parameters at the end of the study.

This short-term toxicity study by the dermal route performed on the registered substance allowed to determine a NOAEL(systemic) of 5 mg/kg bw/day and a LOAEL(local) of 1 mg/kg bw/day. The skin was the only identified target organ.

Endpoint conclusion

Endpoint conclusion:

adverse effect observed

Dose descriptor:

LOAEL

4.33 µg/cm²

Study duration:

subacute

Species:

rabbit

Quality of whole database:

study well documented, meets generally accepted scientific principles, acceptable for assessment

Additional information

A LOAEL of 1 mg/kg bw/d was determined for the local dermal effect of Hexachlorocyclopentadiene following a repeated exposure. The substance was applied to approximately 30% of the total body surface area of the test animal (rabbit). According to the FDA (2005), the rabbit is considered to have an average body surface area of 0.15 m² (equivalent to 1500 cm²) and an average weight of 1.95 kg.

Corrected LOAEL = dose / 30% body surface

Corrected LOAEL = (NOAEL(local) * weight(rabbit)) / (1500 cm² * 30%)

Corrected LOAEL = (1 mg/kg bw * 1.95 kg bw) / (1500 cm² * 30%)

Corrected LOAEL = (1.95 mg) / (450 cm²)

Corrected LOAEL = 4.33*10^-3 mg/kg = 4.33 µg/cm²

FDA (2005) Guidance for Industry Estimating the Maximum Safe Starting Dose in Initial Clinical Trials for Therapeutics in Adult Healthy Volunteers https://www.fda.gov/downloads/drugs/guidances/ucm078932.pdf

Justification for classification or non-classification

An ovarian inflammation was observed in mice exposed to Hexachlorocyclopentadiene by inhalation for two years. The EU Risk Assessment Report (2007) considered this adverse effect to be treatment related due to a clear dose-response relationship observed in in the chronic toxicity study performed by the US NTP (1994). No such effects were observed in the other semi-chronic studies. On the basis of this effect it is proposed a Classification as STOT RE 1; H372 according to Regulation (EC) N° 1272/2008.

 

Signs of irritation of the respiratory tract were observed during repeated-dose toxicity studies performed by inhalation. These effects are consistent with the ones observed during the acute toxicity studies performed on Hexachlorocyclopentadiene. On the basis of these effects it is proposed a Classification as STOT SE 3; H335 (may cause respiratory irritation) according to Regulation (EC) N° 1272/2008.