Registration Dossier

Administrative data

Description of key information

 The following results were obtained for chlorendic anhydride: Oral NOAEL = 1242 mg/kg bw/day Dermal NOAEL = 2500 mg/kg bw day Inhalation NOAEC = 9970 mg/m³ The following results were obtained for chlorendic acid: Oral NOAEL (female) = 620 ppm in diet Oral NOAEL (male) = 1250 ppm in diet Inhalation NOAEC = 0.134 mg/L 

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:
From 25 July 1978 to 24 October 1978
Reliability:
2 (reliable with restrictions)
Rationale for reliability incl. deficiencies:
comparable to guideline study with acceptable restrictions
Qualifier:
equivalent or similar to
Guideline:
OECD Guideline 408 (Repeated Dose 90-Day Oral Toxicity in Rodents)
Deviations:
yes
Remarks:
information on temperature and humidity were not recorded.
GLP compliance:
no
Limit test:
no
Species:
rat
Strain:
Crj: CD(SD)
Sex:
male/female
Details on test animals and environmental conditions:
TEST ANIMALS
- Source: Charles River Breeding Laboratories, Inc., Portage, Michigan
- Weight at study initiation: Male 75 to 106 g. Female 71 to 97 g.
- Fasting period before study: not applicable
- Housing: Suspended wire-mesh cages.
- Diet: ad libitum
- Water: ad libitum

ENVIRONMENTAL CONDITIONS
- Photoperiod (hrs dark / hrs light): 12/12
Route of administration:
oral: feed
Vehicle:
unchanged (no vehicle)
Details on oral exposure:
PREPARATION OF DOSING SOLUTIONS:

DIET PREPARATION
- Rate of preparation of diet (frequency): weekly
- Mixing appropriate amounts with (Type of food): Purine Laboratory Chow
Analytical verification of doses or concentrations:
no
Duration of treatment / exposure:
90 days
Frequency of treatment:
Daily
Dose / conc.:
100 ppm
Remarks:
Treatment level at 100 ppm gave a compound consumption of 8 mg/kg bw/day for males and females.
Dose / conc.:
500 ppm
Remarks:
Treatment level at 500 ppm gave a compound consumption of 39 mg/kg bw/day for males and 45 mg/kg bw/day for females.
Dose / conc.:
2 500 ppm
Remarks:
Treatment level at 2500 ppm gave a compound consumption of 202 mg/kg bw/day for males and 226 mg/kg bw/day for females.
No. of animals per sex per dose:
15
Control animals:
yes, concurrent no treatment
Positive control:
None
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: weekly


FOOD CONSUMPTION AND COMPOUND INTAKE (if feeding study):
- Food consumption for each animal determined and mean daily diet consumption calculated as g food/kg body weight/day: Yes
- Compound intake calculated as time-weighted averages from the consumption and body weight gain data: Yes


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: Yes
- Time schedule for examinations: pre-test period and at 3 months
- Dose groups that were examined: All groups


HAEMATOLOGY: Yes
- Time schedule for collection of blood: 1, 2 and 3 months.
- Anaesthetic used for blood collection: No data
- Animals fasted: No data
- How many animals : Five rats per sex per group


CLINICAL CHEMISTRY: Yes / No / No data
- Time schedule for collection of blood:1, 2 and 3 months.
- Animals fasted: No data
- How many animals: Five rats per sex per group


URINALYSIS: Yes
- Time schedule for collection of urine: 1, 2 and 3 months.
- Metabolism cages used for collection of urine: No data
- Animals fasted: No data


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

HISTOPATHOLOGY: Yes
Statistics:
All statistical analyses compared the treatment groups with the control group by sex Body weights (week 13), food consumption (week 13), hematological biochemical and urinalysis parameters (1, 2 and 3 months) and absolute and relative organ weights (terminal sacrifice) were compared by analysis of variance (one way classification) Bartlett's test for homogeneity 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.
Clinical signs:
effects observed, treatment-related
Description (incidence and severity):
There were no signs of overt toxicity observed for the treated rats. Some incidental signs seen in a few control and or treated rats were malaligned upper incisors, soft stools, skin lesions, hair loss, lacrimation, corneal opacity, redness around eye.
Mortality:
mortality observed, treatment-related
Description (incidence):
Three high dose females died between the 5th and 13th week of study no other rats died during the study.
Body weight and weight changes:
effects observed, treatment-related
Description (incidence and severity):
The mid and high dose males and all three groups of treated females had a decreased rate of weight gain that was compound related. The group mean body weights of the mid and high dose males and high dose females were significantly less than controls at week 13.
Food consumption and compound intake (if feeding study):
effects observed, treatment-related
Description (incidence and severity):
Mid and high dose males and high dose females had decreased food consumption values over the 90-day study when compared with controls however only the food consumption of the high dose females was significantly less than the controls.
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):
Eye problems such as pale coloration of eye, decrease in size of eyeball, dilated pupil unresponsive to light, clear or white internal eye and increased distance between pupil and cornea, occurred most frequently in rats that had had blood drawn via the orbital sinus technique.
Haematological findings:
effects observed, treatment-related
Description (incidence and severity):
No compound-related effects were observed in the results of the hematologic tests. An incidental finding was the slightly elevated number of leucocytes seen for the high dose males at 1, 2 and 3 months of study. Other occasional statistically significant values were of no physiologic importance.
Clinical biochemistry findings:
effects observed, treatment-related
Description (incidence and severity):
Both treated males and females had elevated SAP activities at 1, 2 and 3 months of study (only the 2 and 3 month values of the high dose females and the 2 month value of the mid dose females were significantly greater than controls). At 3 months of study, all groups had greater SAP activities than controls. Mid and low dose males and females had SGOT activities significantly lower than controls at 3 months of study. However, these activities as well as other occasional statistically significant values were of no physiologic importance. No other compound-related effects were seen in the results of the biochemical tests.
Urinalysis findings:
effects observed, treatment-related
Description (incidence and severity):
No compound-related effects were seen in the results of the urinalyses. An incidental finding at 3 months of study was the elevated urinaly pH of two of five high dose females tested. The few statistically significant values (specific gravity) were of no physiologic importance.
Behaviour (functional findings):
not examined
Organ weight findings including organ / body weight ratios:
effects observed, treatment-related
Description (incidence and severity):
Statistically significant (p< 0.05) decreases in the mean absolute weight of spleens of female rats at the 2500 ppm dosage level, mean absolute weight of kidneys of male rats at the 2500 ppm dosage level, mean absolute weight of hearts of male rats at all dosage levels and the female rats at the 500 ppm dosage level and an increase in the mean weight of testes of male rats at the 500 ppm dosage level were noted. In respect of liver, statistically significant (p< 0.01) decreases in the mean absolute and relative weights at all dosage levels were observed. Of these variations decreases in the mean absolute weights of heart of male rats (p< 0.05) and in the mean absolute and relative weights of livers (p< 0.01) at all dosage levels, appeared to be treatment related. In the absence of any significant histomorphologic changes in these organs in the test groups, these decreases probably might be due to the overall reduction in the body weights resulting from a reduction of body fat and or extracellular body fluid.
Gross pathological findings:
effects observed, treatment-related
Description (incidence and severity):
The three rats from the 2500 ppm group that died during the course of the study did not show any compound related lesions. None of the rats that were sacrificed at the termination of the study had any compound related lesions.
Histopathological findings: non-neoplastic:
effects observed, treatment-related
Description (incidence and severity):
No compound-related microscopic lesions were observed in any of the tissues from rats that were examined from the 2500 ppm group. The microscopic lesions seen were considered spontaneous and incidental in nature.
Histopathological findings: neoplastic:
not examined
Key result
Dose descriptor:
NOAEL
Effect level:
> 2 500 ppm
Based on:
test mat.
Sex:
male/female
Basis for effect level:
clinical signs
mortality
body weight and weight gain
food consumption and compound intake
ophthalmological examination
haematology
clinical biochemistry
urinalysis
organ weights and organ / body weight ratios
gross pathology
histopathology: non-neoplastic
Critical effects observed:
no
Lowest effective dose / conc.:
2 500 ppm
System:
immune system
Organ:
spleen
Treatment related:
yes
Dose response relationship:
not specified
Relevant for humans:
not specified
Critical effects observed:
no
Lowest effective dose / conc.:
2 500 ppm
System:
hepatobiliary
Organ:
liver
Treatment related:
yes
Dose response relationship:
not specified
Relevant for humans:
not specified
Critical effects observed:
no
Lowest effective dose / conc.:
2 500 ppm
System:
male reproductive system
Organ:
testes
Treatment related:
yes
Dose response relationship:
not specified
Relevant for humans:
not specified
Critical effects observed:
no
Lowest effective dose / conc.:
2 500 ppm
System:
urinary
Organ:
kidney
Treatment related:
yes
Dose response relationship:
not specified
Relevant for humans:
not specified
Critical effects observed:
no
Lowest effective dose / conc.:
2 500 ppm
System:
cardiovascular
Organ:
heart
Treatment related:
yes
Dose response relationship:
not specified
Relevant for humans:
not specified
Conclusions:
No compound related gross lesions were seen in any of the rats from the treatment groups. No compound related microscopic lesions were seen in any of the tissues from rats that were examined from the 2500 ppm group.
Executive summary:

Chlorendic Anhydride was fed in the diet at levels of 100, 500, and 2S00 ppm to three groups of Charles River CD rats in a 90-day subacute toxicity study performed according to a method similar to OECD Testing Guideline 408 (non GLP) with deviations. There were 15 rats/sex/group. Control rats received the basal laboratory diet only. The rats were observed twice daily for signs of overt toxicity and mortality. Detailed observations, individual body weights and individual food consumption were recorded weekly. Ophthalmic examinations were conducted during the pretest period and at 3 months of study. Hematologic and biochemical tests and urinalyses were performed at 1, 2 and 3 months of study for five rats/sex/groups. Mid and high dose males and all three groups of treated females had decreased group mean body weights when compared with controls. Mid and high dose males and high dose females had decreased food consumption over the 90-day study when compared with controls. Both treated males and females had elevated SAP activities at 1, 2 and 3 months of study. Statistically significant decreases in the mean absolute weights of hearts of male rats and in the mean absolute and relative weights of livers of male and female rats at all dosage levels, appeared to be treatment related. No compound-related gross lesions were seen in any of the rats from the treatment groups. No compound-related microscopic lesions were seen in any of the tissues from rats that were examined from the 2500 ppm group.

Endpoint conclusion
Dose descriptor:
NOAEL
1 242 mg/kg bw/day
Study duration:
subacute
Species:
rat

Repeated dose toxicity: inhalation - systemic effects

Link to relevant study records
Reference
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
Qualifier:
equivalent or similar to
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
Species:
rat
Strain:
Crj: CD(SD)
Sex:
male/female
Details on test animals 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
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.
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
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.
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
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
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
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.

Endpoint conclusion
Dose descriptor:
NOAEC
9 970 mg/m³
Study duration:
subacute
Species:
rat

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
Study period:
From 13 January 1978 to 3 February 1978
Reliability:
2 (reliable with restrictions)
Rationale for reliability incl. deficiencies:
comparable to guideline study with acceptable restrictions
Qualifier:
equivalent or similar to
Guideline:
OECD Guideline 410 (Repeated Dose Dermal Toxicity: 21/28-Day Study)
Deviations:
yes
Remarks:
information on temperature and humidity were not recorded. Food consumption was not monitored.
GLP compliance:
no
Limit test:
yes
Species:
rabbit
Strain:
New Zealand White
Sex:
male/female
Details on test animals and environmental conditions:
TEST ANIMALS
- Source: Sweetwater Farms, Hillsboro, Ohio
- Weight at study initiation: Male 2124 to 2781 g. Female 2187 to 2602 g
- Diet: Purina Rabbit Chow ad libitum
- Water: ad libitum
- Acclimation period: Approximately 2 weeks.
Type of coverage:
occlusive
Vehicle:
physiological saline
Details on exposure:
TEST SITE
- Area of exposure: Dorsal
- % coverage: Approximately 10% of body surface
- Type of wrap if used: None

TEST MATERIAL
- Amount(s) applied (volume or weight with unit): 100, 500 and 2500 mg/kg

VEHICLE
- Justification for use and choice of vehicle (if other than water): physiological saline used as control
- Amount(s) applied (volume or weight with unit): 1.2 ml/kg/day

USE OF RESTRAINERS FOR PREVENTING INGESTION: yes
Analytical verification of doses or concentrations:
no
Duration of treatment / exposure:
5 days per week for 3 weeks for a total of 15 applications
Frequency of treatment:
Daily for 5 weekdays, break at week-ends for 3 weeks.
Dose / conc.:
100 mg/kg bw/day (nominal)
Dose / conc.:
500 mg/kg bw/day (nominal)
Dose / conc.:
2 500 mg/kg bw/day (nominal)
No. of animals per sex per dose:
4
Control animals:
yes, concurrent vehicle
Positive control:
None
Observations and examinations performed and frequency:
CAGE SIDE OBSERVATIONS: Yes

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: No data
- Anaesthetic used for blood collection: No data
- Animals fasted: No data
- How many animals: All

CLINICAL CHEMISTRY: Yes
- Time schedule for collection of blood: At termination of study
- Animals fasted: No data
- How many animals: All

URINALYSIS: Yes
- Time schedule for collection of urine:
- Metabolism cages used for collection of urine: No data

NEUROBEHAVIOURAL EXAMINATION: No
Sacrifice and pathology:
GROSS PATHOLOGY: YesHISTOPATHOLOGY: Yes
Statistics:
All statistical analyses compared the treatment groups with the control group by sex. At termination of the study, body weights, hematologic, biochemical and urinalysis parameters and absolute and relative organ weights were compared by analysis of variance (one-way classification), Bartlett's test for homogeneity 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.
Clinical signs:
no effects observed
Description (incidence and severity):
Incidental findings (primarily at the 2500 mg/kg/day dosage level) included: diarrhea, nasal or ocular discharge, hypoactivity, anorexia and dehydration.
Dermal irritation:
effects observed, treatment-related
Description (incidence and severity):
One or more signs of dermal irritation were present for all treated rabbits. The level of intensity and time of onset were primarily dose related. At the 100 mg/kg/day dosage level, a barely perceptible erythema was noted for all rabbits beginning in weeks 2 or 3 of treatment. At the 500 mg/kg/day dosage level, the onset of erythema (barely perceptible to slight) was usually evident by day 4 and persisted throughout most, or all or the treatment period. Other signs of dermal irritation (barely perceptible to slight) included: edema, atonia, desquamation, coriaceousness and fissuring. These signs were evident for most treated rabbits during either the second or third week of treatment. At the 2500 mg/kg/day dosage level, erythema (barely perceptible to moderate) was evident as early as day 2 and persisted throughout the study for most rabbits. Other signs of dermal irritation as noted previously (barely perceptible to moderate) were evident by, or on, day 7 and for most rabbits persisted throughout the treatment period.
Mortality:
no mortality observed
Description (incidence):
None of the rabbits died.
Body weight and weight changes:
effects observed, treatment-related
Description (incidence and severity):
Female rabbits in the 2500 mg / kg / day group had decreased body weight gains when compared to controls. Males had slight but statistically significant weight losses.
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
Description (incidence and severity):
No changes considered to be related to compound were seen in the hematologic studies.
Clinical biochemistry findings:
no effects observed
Urinalysis findings:
no effects observed
Description (incidence and severity):
No urine was collected from one male and one female rabbit in the 2500 mg kg day treatment group. All other urinalyses were similar to the control.
Behaviour (functional findings):
not examined
Immunological findings:
not examined
Organ weight findings including organ / body weight ratios:
no effects observed
Description (incidence and severity):
No compound related organ weight variations were observed
Gross pathological findings:
effects observed, treatment-related
Description (incidence and severity):
Six rabbits from the 2500 mg/kg/day group and 2 rabbits from the 500 mg/kg/day group had stomach lesions which may have been compound related. These stomach changes were described as ulcerations, erosions and yellow gray or white foci or areas in the mucosa. They were not seen in rabbits from the control or 100 mg/kg/day groups.
Neuropathological findings:
not examined
Histopathological findings: non-neoplastic:
effects observed, treatment-related
Description (incidence and severity):
Stomach erosions noted grossly and confirmed microscopically in several rabbits from the 2500 mg/kg/day group were shallow and did not extend the full thickness of the mucosa. The stomach mucosa away from the grossly noted erosions was completely normal. Erosions or other stomach alterations could not be confirmed microscopically in all stomachs in which a gross description of a lesion was made. Stomach lesions were not seen microscopically in rabbits from the control or 100 mg/kg/day groups; their occurrence in rabbits from the 500 and 2500 mg/kg/day groups was probably compound related.
Evidence of very slight to slight compound-related dermal irritation was seen in most rabbits from the 2500, 500 and 100 mg/kg/day groups. These skin changes included epidermal acanthosis and hyperkeratosis and inflammatory cell infiltrate in the dermis. The severity of these skin changes appeared somewhat dose related and the overall skin response to this compound could best be characterized as mild. Other microscopic lesions were considered spontaneous and unrelated to treatment and were typical of the usual lesions seen in untreated rabbits.
Brain lesions characterized by perivascular lymphocytic cuffing, glial nodules and lymphocytic meningitis were considered due to infestation by Encephalitozoan cunniculli, a common protozoan parasite of laboratory rabbits.
Histopathological findings: neoplastic:
not examined
Key result
Dose descriptor:
NOEL
Effect level:
> 2 500 mg/kg bw/day (nominal)
Based on:
test mat.
Sex:
male/female
Basis for effect level:
other: observed effects were not considered as adverse and significant
Critical effects observed:
no
Lowest effective dose / conc.:
500 mg/kg bw/day (nominal)
System:
gastrointestinal tract
Organ:
stomach
Treatment related:
yes
Dose response relationship:
not specified
Relevant for humans:
not specified
Conclusions:
Evidence of mild skin irritation, characterized by hyperkeratosis, acanthosis and dermal inflammatory cell infiltrate was seen at the application site in most rabbits from the 100, 500 and 2500 mg / kg / day groups and was considered compound related. Overall skin response based on microscopic examination of the application site was characterized as mild.
Executive summary:

Chlorendic Anhydride was administered to the backs of New Zealand White rabbits at dosage levels of 100 500 and 2500 mg/kg/day 5 days a week during this 3 week dermal study according to a method similar to OECD Testing Guideline 410 (non GLP) with deviations. Four male and four female rabbits were used at each dosage level and in the control group. The rabbits were observed daily for signs of overt toxicity, general behavior, dermal irritation, moribundity or mortality. Body weights were recorded weekly Hematologic, biochemical and urinalysis studies were conducted during the control period and following the 2l day treatment period. One or more of the following signs of dermal irritation were noted for all treated rabbits: erythema, edema, atonia, desquamation coriaceousness and fissuring. The number of signs observed, severity of the conditions barely perceptible to moderate and duration were dose related. Incidental findings primarily at the 2500 mg/kg/day dosage level included diarrhea, nasal or ocular discharge, hypoactivity, anorexia and dehydration. Male and female rabbits at the high dosage level had decreases in weight when compared with the controls. All rabbits survived the treatment period. No changes considered related to compound were seen in the hematologic and biochemical studies. Urinalyses were considered normal. Stomach mucosal lesions described as erosions ulcerations or light foci and areas at necropsy in rabbits from the 2500 and 500 mg/kg/day were the only gross findings at terminal sacrifice which were considered compound related. No compound related organ weight variations were observed. Microscopically grossly described stomach changes were confirmed in several rabbits from the 500 and 2500 mg/kg/day groups. These changes were attributed to compound effect. Evidence of mild skin irritation characterized by hyperkeratosis, acanthosis and dermal inflammatory cell infiltrate was seen at the application site in most rabbits from the 100, 500 and 2500 mg/kg/day groups and was considered compound related. Overall skin response based on microscopic examination of the application site was characterized as mild.

Endpoint conclusion
Dose descriptor:
NOAEL
2 500 mg/kg bw/day
Study duration:
subacute
Species:
rabbit

Additional information

Justification for classification or non-classification

The results of the Repeated Dose studies for Chlorendic Anhydride have determined that it will not be classified as toxic by these paths of entry.