Registration Dossier

Administrative data

Description of key information

Dermal:
- Equivalent to OECD 424; GLP not specified; Sprague-Dawley rat; 20, 66.6, 200 mg/kg bw/day; NOAEL (neurotoxicity) 200 mg/kg bw/day (1982)

Key value for chemical safety assessment

Effect on neurotoxicity: via oral route

Endpoint conclusion
Endpoint conclusion:
no study available

Effect on neurotoxicity: via inhalation route

Endpoint conclusion
Endpoint conclusion:
no study available

Effect on neurotoxicity: via dermal route

Link to relevant study records
Reference
Endpoint:
neurotoxicity: sub-chronic dermal
Type of information:
experimental study
Adequacy of study:
key study
Study period:
Jun - Sep 1981
Reliability:
2 (reliable with restrictions)
Rationale for reliability incl. deficiencies:
study well documented, meets generally accepted scientific principles, acceptable for assessment
Reason / purpose:
reference to same study
Qualifier:
equivalent or similar to
Guideline:
OECD Guideline 424 (Neurotoxicity Study in Rodents)
Qualifier:
equivalent or similar to
Guideline:
other: OECD Guideline 411 (Subchronic Dermal Toxicity: 90-Day Study)
Principles of method if other than guideline:
The subchronic dermal application of the test substance to the backs of Sprague-Dawley rats was studied to assess potential neurotoxic and other local and systemic effects.
GLP compliance:
not specified
Limit test:
no
Specific details on test material used for the study:
- Name of test material (as cited in study report): C-178
- Physical state: liquid
- Analytical purity: 100% active ingredient
- Storage condition of test material: room temperature
Species:
rat
Strain:
Sprague-Dawley
Sex:
male/female
Details on test animals and environmental conditions:
TEST ANIMALS
- Source: Charles River Breeding Laboratories
- Age at study initiation: 41 days (age: 28 days at receipt)
- Weight at study initiation: week 0 males: 157-163 g; week 0 females: 133-139 g
- Housing: individually in elevated stainless steel cages
- Diet: ad libitum
- Water: ad libitum
- Acclimation period: 14 days


ENVIRONMENTAL CONDITIONS
- Photoperiod (hrs dark / hrs light): 12/12
Route of administration:
dermal
Vehicle:
corn oil
Details on exposure:
TEST SITE
- Area of exposure: the back of the rats
- % coverage: no data
- Type of wrap if used: not occluded no further data
- Time intervals for shavings or clipplings: all animals were clipped ca. 23 h prior to initial dose. The animals were reclipped when necessary.

TEST MATERIAL
- Amount(s) applied (volume or weight with unit): 2.077 ml/kg
- Concentration (if solution): 1.0, 3.33 and 10.0 %
- Constant volume or concentration used: yes

VEHICLE
- Justification for use and choice of vehicle (if other than water): immiscible with water
- Amount(s) applied (volume or weight with unit): 2.077 ml 7kg of the test substance in corn oil
- Concentration (if solution): 1.0, 3.33 and 10.0 % of the test substance in corn oil

USE OF RESTRAINERS FOR PREVENTING INGESTION: no data
Analytical verification of doses or concentrations:
not specified
Details on analytical verification of doses or concentrations:
the remaining samples of weekly dosing solutions for each dose were returned to the sponsor for analysis, no further data
Duration of treatment / exposure:
90 Days
Frequency of treatment:
daily, 5 days/week
Dose / conc.:
20 mg/kg bw/day (nominal)
Dose / conc.:
66.666 mg/kg bw/day (nominal)
Dose / conc.:
200 mg/kg bw/day (nominal)
No. of animals per sex per dose:
10
Control animals:
yes, concurrent vehicle
Observations and clinical examinations performed and frequency:
CAGE SIDE OBSERVATIONS: Yes
- Time schedule: twice daily
- Cage side observations: for mortality and gross signs of toxicologic or pharmacologic effects

DETAILED CLINICAL OBSERVATIONS: Yes
- Time schedule: weekly including signs of local or systemic toxicity, pharmacologic effects and palpation for tissue masses

BODY WEIGHT: Yes
- Time schedule for examinations: twice pretest, weekly during treatment and terminally (after fasting)


OTHER:
- blood was obtained from over night fasted rats via venipuncture of the orbital sinus under light ether anestehsia, the same animals were used that were intended for formalin-fixation (5/sex/dose).
- hematology upon termination: hemoglobin, hematocrit, erythrocytes, clotting time, total and differential, leukocytes, erythrocytes morphology
- clinical chemistry upon termination: serum glutamic oxaloacetic transaminase, serum glutamic pyruvic transaminase, blood urea nitrogen, fasting glucose, total protein, total bilirubin, sodium, potassium, calcium, inorganic phosphorus
- urinanalysis 6 days before termination: gross appearance, specific gravity, pH, protein, glucose, occult blood
Specific biochemical examinations:
no data
Neurobehavioural examinations performed and frequency:
- Neurological Function Evaluations: Yes
- Time interval: monthly
- Parameters examined according to a scoring system: posture, gait, muscular tone, reflexes (corneal), righting and toe-pinch
- no further data
Sacrifice and (histo)pathology:
One-half of the animals were sacrificed by exsanguination under light ether anesthesia, and selected organs and tissues were fixed
in formalin. Organ and organ-body weight ratios (adrenals, brain, liver, kidney, heart, spleen, testes with epididymides, ovaries) were determined on all of these animals only. Histopathological evaluations of 10 organs or tissues (liver, kidney, lung, heart, stomach, adrenal, pituitary, testes, ovaries,
spleen and skeletal muscle) were conducted on all formalin-fixed control animals and the high dose animals. The remaining animals were perfused
intravenously with glutaraldehyde under sodium pentobarbital anesthesia . Quantitative assessments of teased tibial nerve preparations were
performed on all glutaraldehyde-perfused animals in control animals and the high dose animals. In addition, brain, spinal cord and sciatic nerve were evaluated microscopically (hematoxylin and eosin and Luxol-fast blue staining) from these same animals.
Positive control:
no positive control included
Statistics:
Body weight, hematology and clinical chemistry parameters, organ weights and organ/body weight ratios were analyzed. Mean values of
all dose groups were compared to control at each time interval.
Clinical signs:
effects observed, non-treatment-related
Description (incidence and severity):
Weekly physical examinations of all animals failed to indicate any toxic effects of the test material other than irritation at the application site. Alopecia observed on the forepaws and legs was the most common finding in both sexes; however, the incidence was spread over all test groups, including control.
Dermal irritation (if dermal study):
effects observed, treatment-related
Description (incidence and severity):
The test substance produced moderate levels of irritation, in a dose-related manner, beginning the first week of the study. Males were generally more susceptible than females to the dermal effects of the test substance throughout the study. Dermal effects (erythema and eschar formation) were only scored as present or absent therefore, the number of times per week the effects were noted was used as a general indication of the severity of the dermal observations.
- 20 mg/kg/day: erythema noted occassionally during the final 2 months; more frequently in the initial 3 weeks; exfoliation observed in approximately one-half during weeks 2 and 3 with diminishing frequency
- 66.6 mg/kg/day: erythema noted with a somewhat higher frequency than 20 mg/kg/day, frequency in females comparable to control. Exfoliation and eschar recorded for most animals by week 3.
- 200 mg/kg/day: erythema, exfoliation, and eschar seen in most animals of both sexes beginning in week 1. Atonia was observed in one males and one female, fissures present in one female and four males. A persistant fissuring was observed in one male rat from week 2 through week 7.
Males appeared somewhat more sensitive than females to erythema and eschar formation.
Mortality:
no mortality observed
Description (incidence):
All animals survived to the scheduled termination of the study.
Body weight and weight changes:
effects observed, treatment-related
Description (incidence and severity):
Significantly reduced body weights in high dose male rats (versus control male rats) were noted from weeks 3 through 12, except during weeks 10 and 11. Male rat weights of the low and mid dose groups were also reduced in dose-related fashion, but the differences were not statistically significant in comparison to control values. No statistically significant effects on weight were seen in treated female rats, however, high dose female animals never exceeded 95% of the mean weights of control females after 2 weeks on test.
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
Description (incidence and severity):
Of the clinical chemical parameters which were determined at the study termination, none were affected in a manner suggestive of a treatment-effect.
Urinalysis findings:
effects observed, non-treatment-related
Description (incidence and severity):
Protein (2+, 100 mg/dl) was confirmed to be present in the urine of one mid-dose male and female, in two high-dose males and one high-dose female. A large amount of occult blood was also present in the urine of this one high-dose female. The specific gravity of this high-dose female and one mid-dose female was also high (>1.090). These findings suggested a possible effect of the test substance on the kidneys, but no alterations were observed in the histopathological examination to support this conclusion. Additionally, without relation to urinary volumes and creatinine, the relevance of in urinary protein concentration is questionable. All in all, the singular changes are not considered adverse by the registrant.
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):
There was a significant downward trend in male liver weights; however, this was not evident in the liver/body weight ratio and is therefore of doubtful significance. Other organ weights and organ/body weight ratios were comparable across all groups.
Gross pathological findings:
effects observed, non-treatment-related
Description (incidence and severity):
Formalin-fixed rats:
No changes, gross or microscopic, were evident which could be attributed to a systemic toxic effect of the test substance. The most common spontaneous gross necropsy findings, occurring across all groups, were inflammations around the ear tags, and slight hair loss on the extremities. No unusual microscopic pathological findings were evident which could be attributed to the topical administration of the test substance.
Neuropathological findings:
effects observed, non-treatment-related
Description (incidence and severity):
Month 1 neurological function tests showed two high-dose males with slightly reduced corneal response. All other evaluations were normal.
Month 2 neurological function tests were unremarkable in control, low and mid dose animals (both sexes). Four high dose males and three high dose females showed slightly abnormal gait described as "stilted". A slight decreased corneal reflex was observed in four males and one female. A moderately decreased toe pinch response (hindtoes only) was also present in one male rat.
Month 3 neurological function tests showed a slightly stilted gait and altered righting reflex in one male control rat and a slightly relaxed body tone in one mid dose male rat. One male high dose animal continued to exhibit a moderately decreased toe-pinch response (hindtoes only) . All
neurological observations were normal in both the control and treated female rats at Month 3.
Histopathological findings: non-neoplastic:
effects observed, non-treatment-related
Description (incidence and severity):
Glutaraldehyde-perfused rats:
Histopathological examinations of hematoxylin and eosin and Luxol-fast Blue stained slides of brain, spinal cord and sciatic nerve from ten rats (5/sex) treated with the high dose of the test substance failed to reveal any treatment-related lesions when these tissues were compared to similar ones from ten control rats (5/sex). In addition, microscopic examination of 50 teased nerve fibers from the tibial nerve of ten high-dose animals (5/sex) were comparable to those of the controls. When quantitative measurements were taken of myelinated nerve fibers in cross-section of the distal sciatic nerve, a slight shift to larger diameters could be detected in high-dose males when compared to the controls. However, there was also a slight decrease in fiber diameters in treated females. The relatively large standard deviation in data from both males and females suggest that these slight changes in fiber diameters are not significant. Moreover, the absence of lesions by more conventional histopathological examinations substantiate this conclusion.
Histopathological findings: neoplastic:
no effects observed
Dose descriptor:
NOAEL
Remarks:
for neurotoxicity
Effect level:
200 mg/kg bw/day (nominal)
Based on:
test mat.
Sex:
male/female
Basis for effect level:
other: no adverse effects observed
Critical effects observed:
yes
Lowest effective dose / conc.:
66.6 mg/kg bw/day (nominal)
System:
integumentary
Organ:
skin
Treatment related:
yes
Dose response relationship:
yes
Conclusions:
The test substance did not induce neurotoxicity and no sub-chronic toxicity other than dermal irritation and decreased body weight was observed in animals, under the conditions tested. Thus, the NOAEL for neurotoxic effects was considered to be greater than 200 mg/kg/day.
Executive summary:

The sub-chronic dermal application of the test substance to the backs of Sprague-Dawley rats was studied to assess potential neurotoxic and other local and systemic toxic effects. Three groups of 20 rats each (10/sex/level) were treated topically with 20, 66 2/3, and 200 mg/kg/day 5 days per week for 3 months. Twenty control animals (10/sex) were treated with corn oil. Solutions at appropriate levels were prepared in corn oil and a constant dose volume (2.077 ml/kg) was applied to all animals. Treatment sites were not occluded. Dermal observations were performed pretest and 5 times/week throughout the study. Clinical laboratory studies were performed at termination. Neurological function evaluations were performed at months 1, 2, and 3. At three months, all animals were terminated. One-half of the animals were sacrificed by exsanguination under light ether anesthesia, and selected organs and tissues were fixed in formalin. Organ and organ body weight ratios were determined on all of these animals only. Histopathological evaluations of 10 organs or tissues were conducted on all formalin-fixed Group 1 and 4 animals. The remaining animals were perfused intravenously with glutaraldehyde under sodium pentobarbital aesthesia. Quantitative assessments of teased tibial nerve preparations were performed on all glutaraldehyde-perfused animals in Group 1 and 4. In addition, brain, spinal cord and sciatic nerve were evaluated microscopically (hematoxylin and eosin and Luxol-fast Blue staining) from these same animals. Erythema, eschar and exfoliation were recorded during the initial week of the study and maximum frequency of these effects were seen during Weeks 2 and 3. A dose-response was noted, with males being slightly more susceptible. In the last 2 months of the study most treated animals developed an apparent tolerance to the irritant effects of the test substance. Significantly lower body weights were recorded throughout the study for Group IV males (200 mg/kg/day). Body weights for female rats and Group II and III male rats were not significantly affected. Routine toxicologic and pharmacologic signs were unremarkable throughout the study. Month 2 neurologic evaluations showed an effect on gait in 4 of 10 male and 3 of 10 female Group IV rats. Reduced corneal reflex was also seen in some rats in this group; however, Month 3 evaluations failed to show these effects. Hematological and clinical chemistry parameters appeared unaffected by treatment with the test substance. However, there was a dose-related increase in urinary protein values in both sexes. Organ weights, gross necropsy observations and microscopic studies did not reveal any systemic toxic effects. Using tibial nerve teasing techniques, no morphometric differences were found between Group land Group IV glutaraldehyde perfused rats. Quantitative assessment of tibial nerve fiber cross-sections showed slightly increased diameters for males and slightly decreased diameters for females. These changes were not considered to be treatment-related.

Endpoint conclusion
Endpoint conclusion:
no adverse effect observed
Dose descriptor:
NOAEL
200 mg/kg bw/day
Study duration:
subchronic
Species:
rat

Additional information

The sub-chronic dermal application of the test substance to the backs of Sprague-Dawley rats was studied to assess potential neurotoxic and other local and systemic toxic effects. Three groups of 20 rats each (10/sex/level) were treated topically with 20, 66 2/3, and 200 mg/kg/day 5 days per week for 3 months. Twenty control animals (10/sex) were treated with corn oil. Solutions at appropriate levels were prepared in corn oil and a constant dose volume (2.077 ml/kg) was applied to all animals. Treatment sites were not occluded. Dermal observations were performed pretest and 5 times/week throughout the study. Clinical laboratory studies were performed at termination. Neurological function evaluations were performed at months 1, 2, and 3. At three months, all animals were terminated. One-half of the animals were sacrificed by exsanguination under light ether anesthesia, and selected organs and tissues were fixed in formalin. Organ and organ body weight ratios were determined on all of these animals only. Histopathological evaluations of 10 organs or tissues were conducted on all formalin-fixed Group 1 and 4 animals. The remaining animals were perfused intravenously with glutaraldehyde under sodium pentobarbital aesthesia. Quantitative assessments of teased tibial nerve preparations were performed on all glutaraldehyde-perfused animals in Group 1 and 4. In addition, brain, spinal cord and sciatic nerve were evaluated microscopically (hematoxylin and eosin and Luxol-fast Blue staining) from these same animals. Erythema, eschar and exfoliation were recorded during the initial week of the study and maximum frequency of these effects were seen during Weeks 2 and 3. A dose-response was noted, with males being slightly more susceptible. In the last 2 months of the study most treated animals developed an apparent tolerance to the irritant effects of the test substance. Significantly lower body weights were recorded throughout the study for Group IV males (200 mg/kg/day). Body weights for female rats and Group II and III male rats were not significantly affected. Routine toxicologic and pharmacologic signs were unremarkable throughout the study. Month 2 neurologic evaluations showed an effect on gait in 4 of 10 male and 3 of 10 female Group IV rats. Reduced corneal reflex was also seen in some rats in this group; however, Month 3 evaluations failed to show these effects. Hematological and clinical chemistry parameters appeared unaffected by treatment with the test substance. However, there was a dose-related increase in urinary protein values in both sexes. Organ weights, gross necropsy observations and microscopic studies did not reveal any systemic toxic effects. Using tibial nerve teasing techniques, no morphometric differences were found between Group land Group IV glutaraldehyde perfused rats. Quantitative assessment of tibial nerve fiber cross-sections showed slightly increased diameters for males and slightly decreased diameters for females. These changes were not considered to be treatment-related.

The test substance did not induce neurotoxicity and no sub-chronic toxicity other than dermal irritation and decreased body weight was observed in animals, under the conditions tested. Thus, the NOAEL for neurotoxic effects was considered to be greater than 200 mg/kg/day.

Justification for classification or non-classification

No classification for neurotoxicity is warranted as no neurotoxic effects were observed in a respective neurotoxicity study.