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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
Study period:
From 7 June 1978 to 5 July 1978
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:
1979
Report date:
1979

Materials and methods

Test guideline
Qualifier:
equivalent or similar to guideline
Guideline:
OECD Guideline 412 (Subacute Inhalation Toxicity: 28-Day Study)
Deviations:
yes
Remarks:
information on temperature, humidity, and airflow were not recorded. Food consumption was not monitored.
GLP compliance:
no
Limit test:
no

Test material

Constituent 1
Chemical structure
Reference substance name:
1,4,5,6,7,7-hexachloro-8,9,10-trinorborn-5-ene-2,3-dicarboxylic anhydride
EC Number:
204-077-3
EC Name:
1,4,5,6,7,7-hexachloro-8,9,10-trinorborn-5-ene-2,3-dicarboxylic anhydride
Cas Number:
115-27-5
Molecular formula:
C9H2Cl6O3
IUPAC Name:
1,4,5,6,7,7-hexachloro-8,9,10-trinorborn-5-ene-2,3-dicarboxylic anhydride
Details on test material:
- Name of test material (as cited in study report): Chlorendic anhydride
- Physical state: White somewhat chunky powder

Test animals

Species:
rat
Strain:
Crj: CD(SD)
Sex:
male/female
Details on test animals or test system and environmental conditions:
TEST ANIMALS
- Source: Charles River
- Age at study initiation: 6 - 8 weeks
- Weight at study initiation: Male 191 to 211 g. Female 182 to 206
- Housing: Wire-mesh cages
- Diet: Purina Laboratory Chow, ad libitum
- Water: ad libitum
- Acclimation period: One week

Administration / exposure

Route of administration:
inhalation: dust
Type of inhalation exposure:
whole body
Vehicle:
air
Mass median aerodynamic diameter (MMAD):
ca. 6 µm
Geometric standard deviation (GSD):
3.16
Details on inhalation exposure:
GENERATION OF TEST ATMOSPHERE / CHAMBER DESCRIPTION
- Exposure apparatus: All exposures were conducted ln a 1 cubic meter cubical stainless steel and glass chamber with pyramidal top and bottom
- Source and rate of air: A constant chamber airflow was maintained by means of a rotary centrifugal air pump located at the exhaust side of the chamber.
- System of generating particulates/aerosols: IRAD dust generators were used for generating the dust atmospheres. The DUSTGUN consisted of a revolving plate with calibrated "cups" for transporting a known quantity of powder per unit time from a reservoir to airjet. At the air jet the powders in a "cup" were dispersed into the chamber by blowing at a rate of 8 liters per minute. The dusts emerging from the dust generators were directed into the exposure chamber air inlet and were further diluted by the incoming make-up air to the desired concentration.
- Temperature, humidity, pressure in air chamber: controlled (no additional information provided).
- Air flow rate: 8 L / min
- Method of particle size determination: Andersen@ 8 stage cascade impactor
- Treatment of exhaust air: Passed through activated charcol filter and a Cambridge Absolute filter before being further diluted with air and discharged outside of the laboratory.

TEST ATMOSPHERE
- Brief description of analytical method used: The concentration of the airborne dusts in the chamber atmosphere was determined gravimetrically using the fiberglass filter sampling technique The chamber atmosphere was drawn through a 37 mm fiberglass filter at the rate of 2 liters min for a suitable
duration A critical orifice was used for regulating the air flow rate Three samples were taken from each chamber during the 6 hour exposure period The weight of ch10rendic anhydride collected on the filter was determined and the quantity of the chlorendic anhydride powder per unit volume of chamber air was calculated in milligrams liter.
- Samples taken from breathing zone: no
Analytical verification of doses or concentrations:
yes
Details on analytical verification of doses or concentrations:
The concentrations are based upon the empirical chamber airflow rate and the powder dissemination rate The quantity of powder dissiminated was determined by weighing the quantity of powder in the reservoir of the dust generator before and after the experiment The concentration of the dusts in the chamber atmosphere was calculated from the ratio of the average rates of powder dissemination to the rate of total chamber airflow (the volume of air ejected from the dust generator plus the volume of make up air passing through the chamber per unit time).
Duration of treatment / exposure:
6 hours per day, 5 days per week for 20 exposures during a 28 day period.
Frequency of treatment:
Daily during weekdays.
Doses / concentrationsopen allclose all
Dose / conc.:
0 mg/L air (nominal)
Dose / conc.:
0.11 mg/L air (nominal)
Dose / conc.:
0.99 mg/L air (nominal)
Dose / conc.:
9.97 mg/L air (nominal)
No. of animals per sex per dose:
10
Control animals:
yes, concurrent no treatment
Details on study design:
- Dose selection rationale: not specified
Positive control:
None

Examinations

Observations and examinations performed and frequency:
CAGE SIDE OBSERVATIONS: Yes
- Time schedule: Before and after immediately after the 6-hour exposure.

DETAILED CLINICAL OBSERVATIONS: Yes
- Time schedule: Before and after immediately after the 6-hour exposure.

BODY WEIGHT: Yes
- Time schedule for examinations: Individual body weights were recorded periodically before the initiation of the study and twice weekly during the four weeks of exposure.

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: Yes
- Time schedule for examinations: Daily
- Dose groups that were examined: All

HAEMATOLOGY: Yes
- Time schedule for collection of blood: All rats at termination of study.
- Anaesthetic used for blood collection: No data
- Animals fasted: Yes, overnight.
- How many animals: All

CLINICAL CHEMISTRY: Yes
- Time schedule for collection of blood: All rats at termination of study.
- Animals fasted: Yes, overnight
- How many animals: All

URINALYSIS: No

NEUROBEHAVIOURAL EXAMINATION: No
Sacrifice and pathology:
GROSS PATHOLOGY: Yes

HISTOPATHOLOGY: Yes. Adrenals, aorta, brain (2 sections), bone-rib junction, gonad, heart (with coronary vessels), duodenum, jejunum, ileum, esophagus, colon, kidneys, liver (2 sections), lungs (2 sections), mesenteric lymph node, nasal turbinates, pancreas, pituitary, salivary gland, skin, mammary gland, spleen, stomach, thymus,. thyroids, urinary bladder, uterus, trachea and gross lesions.
Statistics:
All statistical analyses compared the treatment groups with the control group by sex. The haematological and biochemical parameters and absolute and relative organ weights were compared by analysis of variances and the appropriate t-test (for equal or unequal variances) as described by Steel and Torrie using Dunnett's multiple comparison tables to judge significance of differences.

Results and discussion

Results of examinations

Clinical signs:
effects observed, treatment-related
Description (incidence and severity):
The rats exhibited varying degrees of ocular and nasal irritation, salivation and hair loss in relation to the chlorendic anhydride concentrations. In the group exposed to 9.97 mg / liter, all the rats, except one, exhibited at various times and in varying degrees, red tinged nasal and ocular discharge when observed immediately after exposure. Salivation was also noted in some of the rats. When observed the following morning, before exposure, these clinical signs had generally disappeared. Also in half of the animals, alopecia was observed, ranging from slight thinning on the abdomen to large patches on the back of the animals in the high-concentration group. These symptoms appeared in the low and intermediate groups but with less severe effects and were exhibited by fewer animals. It should be noted that 2 of the control rats also displayed slight alopecia.
Mortality:
no mortality observed
Body weight and weight changes:
effects observed, treatment-related
Description (incidence and severity):
The male rats of the high-concentration group showed depressed weight gains as compared to the controls. The low and medium concentration groups and the female rats of the high-concentration gained weight comparable to the rats of the control group.
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:
effects observed, treatment-related
Description (incidence and severity):
The rats exhibited varying degrees of ocular irritation.
Haematological findings:
effects observed, treatment-related
Description (incidence and severity):
Hematocrit values are significantly depressed from control values in all three exposure groups. In addition, the number of erythrocytes and hemoglobin levels were significantly depressed in the low-concentration group. The hematocrit value for the low-concentration group was significantly depressed from control values. The number of erythrocyteswas significantly increased for the medium and high concentration groups while the number of leucocytes was significantly increased ln the high concentration group. Even though these changes in values for both males and females are statistically significant these values probably do not represent any biological significance since they fall within the normal range of biological variation for the species
Clinical biochemistry findings:
effects observed, treatment-related
Description (incidence and severity):
For male, the fasting blood glucose level was significantly depressed from control values in the high-concentration group. Also for males, alkaline phosphatase values were significantly increased in the intermediate and high concentration groups while the SGPT level was significantly increased in the intermediate concentration group. For females, alkaline phosphatase was significantly increased in the intermediate and high concentration groups. Even though the changes in alkaline phosphatase and SGPT values were statistically significant, these values probably do not represent an exposure related effect since these values are within the normal range of biological variation for the species
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):
Statistically significant decreases in mean relative weights of livers in males in all treated groups and the mean absolute and relative weights of thyroids in female rats in the 1.0 mgl and 10 mg/l groups were considered probably compound related. The variations in the mean weights seen in adrenals and pituitary were not consistant and their bio logical significance is unknown.
Gross pathological findings:
effects observed, treatment-related
Description (incidence and severity):
Gross lesions that were considered probably treatment related were seen in the lung and stomach In the lungs these consisted of increased incidence of dark red foci and dark red areas discoloration and in the stomach dark red or brown foci on the glandular mucosa. Other gross lesions observed in the control and the treatment groups were considered spontaneous or incidental in nature and were not treat ment related.
Neuropathological findings:
not examined
Histopathological findings: non-neoplastic:
effects observed, treatment-related
Description (incidence and severity):
Compound related microscopic changes were observed in treated groups at all levels in the lungs trachea nasal turbinates and glandular part of the stomach.
In the lungs the lesions consisted of increased incidence and intensity in interstitial inflammatory cell infiltration peribronchial lymphoid hyperplasia perivascular lymphocytic infiltration interstitial fibrosis and scattered hemorrhages Interstitial pneumonia occurred only in the treated groups.
In the trachea the changes included inflammatory cell infiltration in mucosa and submucosa and focal epithelial hyperplasia In the nasal turbinates changes seen were inflammatory cell infiltrate in mucosa and dark brown pigment and or inflammatory exudate in the nasal In the glandular part of the stomach inflammatory cell infiltrate in mucosa and submucosa and focal mucosal congestion in the 10 mg/l group and focal mucosal congestion in the 1 mg/l and O.l mg/l group were seen.
Other organs in which microscopic lesions occurred included adrenals (excessive vacuolation in cortical cells), liver (portal inflammatory cell infiltration, scattered inflammatory foci) kidney (interstitial lymphocytic infiltration healed infarct), uterus (hydrometra and cervical lymph node congestion, erythrophagocytosis). These lesions were or low or equal frequency in both the control and the high dose groups and were considered spontaneous and incidental in nature and not related to treatment.
Histopathological findings: neoplastic:
not examined

Effect levels

Key result
Dose descriptor:
NOEC
Effect level:
9 970 mg/m³ air (nominal)
Based on:
test mat.
Sex:
male/female
Basis for effect level:
other: observed effects were not considered as adverse and significant

Target system / organ toxicity

open allclose all
Critical effects observed:
no
Lowest effective dose / conc.:
0.11 mg/L air (nominal)
System:
respiratory system: upper respiratory tract
Organ:
nasal cavity
trachea
Treatment related:
yes
Dose response relationship:
not specified
Relevant for humans:
not specified
Critical effects observed:
no
Lowest effective dose / conc.:
0.11 mg/L air (nominal)
System:
respiratory system: lower respiratory tract
Organ:
lungs
Treatment related:
yes
Dose response relationship:
not specified
Relevant for humans:
not specified
Critical effects observed:
no
Lowest effective dose / conc.:
0.11 mg/L air (nominal)
System:
gastrointestinal tract
Organ:
stomach
Treatment related:
yes
Dose response relationship:
not specified
Relevant for humans:
not specified

Applicant's summary and conclusion

Conclusions:
Compound related microscopic changes of a hemorrhagic inflammatory nature in the lungs and of an inflammatory nature in the trachea nasal turbinates and stomach mucosa occurred in rats from all treated groups.
Executive summary:

Male and female rats were exposed to the dusts of chlorendic anhydride for 6 hours per day 5 days per week for 20 exposures during a 28-day exposure period according to a method similar to OECD Guideline 412 (non GLP). The average nominal exposure concentrations were 0.0, 0.11, 0.99 and 9.97 mg/l air. The aerodynamic equivalent mass medium diameter was calculated to be 6.0 micrometers with a geometric standard deviation of 3.16. Immediately after the 6-hour exposure period the rats exhibited varying degrees of occular and nasal irritation and salivation in relation to the ch10rendic anhydride concentrations. By the following morning these symptoms had generally disappeared. The rats also exhibited alopecia ranging from a slight thinning on the abdomen to large patches lost on the back of some of the rats in the high concentrations group. The low and medium concentration groups and the female rats of the high concentration group demonstrated weight gains comparable to the rats of the control group. The male rats of the high concentration exhibited decreased weight gains as compared to the controls. Statistical differences in hematocrit erythrocytes and hemoglobin concentrations were observed in male rats while statistical differences in hematocrit erythrocytes and leucocytes were seen in female rats. Similarly statistical differences in glucose alkaline phosphatase and SGPT levels were observed in male rats while statistical differences in alkaline phosphatase levels were observed in female rats. Increased incidence of dark red foci and dark red area discoloration in the lungs and dark red or brown foci in glandular part of the stomach seen at necropsy in the treated groups were probably compound related. Statistically significant decreases probably compound related were noted in the mean relative weights of livers of males from all treated groups and in the mean absolute and relative weights of thyroids in female rats from the 1.0 mg/l and 10.0 mg/l groups. Compound related microscopic changes of a hemorrhagic inflammatory nature n the lungs and of an inflammatory nature in the trachea nasal turbinates and stomach mucosa occurred in rats from all treated groups.