Registration Dossier

Administrative data

Endpoint:
sub-chronic toxicity: dermal
Type of information:
experimental study
Adequacy of study:
key study
Reliability:
2 (reliable with restrictions)
Rationale for reliability incl. deficiencies:
other: see 'Remark'
Remarks:
OECD Guideline for Testing of Chemicals No. 410. Repeated Dose Dermal Toxicity:21/28-Day,1-8. May. 1981 State Environmental Protection Administration : The Guidelines for the Testing of Chemicals Ministry of Health,2005: Technical standards for testing of chemicals .

Data source

Reference
Reference Type:
study report
Title:
Unnamed
Year:
2010
Report Date:
2010

Materials and methods

Test guidelineopen allclose all
Qualifier:
according to
Guideline:
OECD Guideline 410 (Repeated Dose Dermal Toxicity: 21/28-Day Study)
Deviations:
no
Qualifier:
according to
Guideline:
other: State Environmental Protection Administration : The Guidelines for the Testing of Chemicals
Deviations:
no
Qualifier:
according to
Guideline:
other: Ministry of Health,2005: Technical standards for testing of chemicals .
Deviations:
no
GLP compliance:
yes (incl. certificate)
Limit test:
yes

Test material

Reference
Name:
Unnamed
Type:
Constituent

Test animals

Species:
rat
Strain:
Sprague-Dawley
Sex:
male/female
Details on test animals and environmental conditions:
Test species: Rats.
Strain: SD.
Supplier: Guangdong Medical Laboratory Animal Center [certified animal number SCXK (Guangdong) 2008-0002].
Number of animals: 25 males and 25 females.
Number of groups: 5(control group, Low-dose group, Intermediate-dose group, High-dose group and Additional group of high-dose).
Number of animal/group: 5 males and 5 females per group.
Body weight at the start of experiment: 200 ~ 274g .
Identification of animals: Tags marked with animal group number and treatment details were attached to cages. Each animal was given an unique number.
Acclimatization: Five days prior to the experiment in the test room.
Randomization: Animals were assigned to five groups randomly.


Environmental conditions
Test Room: In the Center
Animal house conditions: The test facility was an air-conditioned room with 12h artificial fluorescent light and 12h dark.
Temperature range: 20~25oC.
Humidity range: 40~70%.

Husbandry Practices
Caging: Stainless steel cages were used. Autoclaved clean dry corncob was used as the bedding material. Animals were housed in one group according to sex in cages.
Water bottle: Each cage was supplied with a polypropylene water bottle with a stainless steel nozzle.
Sanitation: Bedding material was changed daily.
Food and water: Standard pellet feed supplied by Guangdong Medical Laboratory Animal Center and ultra-pure filtered sterilized water were provided to the animals freely.
Frequency of providing feed and drinking water: Both drinking water and feed were provided ad libitum.

Administration / exposure

Type of coverage:
open
Vehicle:
water
Details on exposure:
Healthy young adult animals are acclimatized to the laboratory conditions for 5 days prior to the test. Before the test, animals are randomized and assigned to the treatment and control groups. An area of about 5×8 cm2 on the back of the animal was clipped free of hair. Repeat clipping or shaving is usually needed at approximately weekly intervals. Pepton 22 was administered on the hair-free site of the rats for 6 hours per day on a 5-day per week basis, for a period of 28 days. The rats were fasten in the stainless steel shelves for 6h, then the test site were cleaned with warm water every time. Animals of the Low-dose group were conducted in the 40 mg/kg dose level, the Intermediate-dose group were conducted in the 200 mg/kg dose level, the High-dose group were conducted in the 800 mg/kg dose level, the Additional group of high-dose were conducted in the 800 mg/kg dose level too, the control group were given distilled water instead of tested compounds, with the same procedure as all the test groups.
Analytical verification of doses or concentrations:
not specified
Duration of treatment / exposure:
28 days
Frequency of treatment:
Pepton 22 was administered on the hair-free site of the rats for 6 hours per day on a 5-day per week basis, for a period of 28 days.
Doses / concentrationsopen allclose all
Remarks:
Doses / Concentrations:
Low-dose group: 40 mg/kg
Basis:
nominal per unit body weight
Remarks:
Doses / Concentrations:
Medium-dose group: 200 mg/kg
Basis:
nominal per unit body weight
Remarks:
Doses / Concentrations:
High-dose group: 800 mg/kg
Basis:
nominal per unit body weight
Remarks:
Doses / Concentrations:
High-dose group (additional): 800 mg/kg
Basis:
nominal per unit body weight
No. of animals per sex per dose:
5 males and 5 females per group
25 males and 25 females in total
Control animals:
yes, concurrent vehicle
Details on study design:
Healthy young adult animals were acclimatized to the laboratory conditions for 5 days prior to the test. Before the test, animals are randomized and assigned to the treatment and control groups. An area of about 5×8 cm2 on the back of the animal was clipped free of hair. Repeat clipping or shaving were usually needed at approximately weekly intervals. Pepton 22 was administered on the hair-free site of the rats for 6 hours per day on a 5-day per week basis, for a period of 28 days. The rats were fasten in the stainless steel shelves for 6h, then the test site were cleaned with warm water every time. Animals of the Low-dose group were conducted at the 40 mg/kg dose level, the Medium-dose group were conducted at the 200 mg/kg dose level, the High-dose group were conducted at the 800 mg/kg dose level, the Additional high-dose group were conducted at the 800 mg/kg dose level too, the Control group were given distilled water instead of tested compounds, with the same procedure as all the test groups.
Positive control:
The Control group was given distilled water instead of tested compounds, using the same procedure as all the test groups.

Examinations

Observations and examinations performed and frequency:
Observation period: All animals were observed signs of toxicity, including the time of onset the degree and duration, everyday for a period of 28 days, but animals of Additional high-dose group scheduled for follow-up observations were kept for a further 14 days without treatment to detect recovery from, or persistence of, toxic effects.

Observation: A careful clinical examination was made once each day. Additional observations was made daily with appropriate actions taken to minimize loss of animals to the study. Necropsy or refrigeration of those animals found dead and isolation or sacrifice of weak or moribund animals.

Body weight: Body weight of each animal was recorded weekly and before necropsy.

Hematology test: Including erythrocyte count, hematocrit, hemoglobin concentration, platelet count, total and differential leucocyte count, thromboplastin time and partial thromboplastin time were investigated at the end of the test period.

Biochemistry test: Clinical biochemistry determination on blood was carried out at the end of the test period. Blood parameters of liver and kidney function are appropriate. The selection of specific tests was influenced by observations on the mode of action of the substance.
Include serum glutamic-pyruvic transaminase(ALT), serum glutamic-oxaloacetic transaminase(AST), gamma glutamyl transpeptidase(GGT), total bilirubin(TB), total serum protein(TP), albumen(ALB), blood creatinine(CRE) and urea nitrogen(BUN) measurements.

Gross necropsy: All animals in the study were subjected to a full gross necropsy which included examination of the external surface of the body, all orifices, and the cranial, thoracic and abdominal cavities and their contents. The liver, kidneys, adrenals and testes were weighed wet as soon as possible after dissection to avoid drying.
The following organs and tissues were preserved in a suitable medium for possible future histopathological examination: normal and treated skin, brain, heart, liver, spleen, lungs, kidney, paranephroi, testis, epididymides, ovaries and uteruses.

Histopathology: Histological examination was performed with microscopy on H.E. stained slides of the preserved organs and tissues of all the tested groups and the control group.
Sacrifice and pathology:
At the end of the experiment, all the animals were sacrificed to test some relevant indexes.

Results and discussion

Results of examinations

Clinical signs:
no effects observed
Dermal irritation:
not examined
Mortality:
no mortality observed
Body weight and weight changes:
no effects observed
Food consumption and compound intake (if feeding study):
no effects observed
Food efficiency:
not examined
Water consumption and compound intake (if drinking water study):
no effects observed
Ophthalmological findings:
not examined
Haematological findings:
effects observed, treatment-related
Description (incidence and severity):
platelet count PLT increased (P <0.05) in males, thromboplastin time (PT) prolonged (P <0.05) in males.
Clinical biochemistry findings:
effects observed, treatment-related
Description (incidence and severity):
Biochemistry test: Compared to the result of the indexes of the Control group, there were no significant differences both in females and males in Low-, Medium-, and High-dose group. But the result of the indexes of Additional high-dose group compared with
Urinalysis findings:
not examined
Behaviour (functional findings):
no effects observed
Organ weight findings including organ / body weight ratios:
effects observed, treatment-related
Description (incidence and severity):
Compared to the result of the indexes of the Control group, there were no significant differences in females in Low-, Medium-, and High-dose group. Compared with the High-dose group, the liver coefficients and Ovaries of the Additional high-dose group wer
Gross pathological findings:
no effects observed
Description (incidence and severity):
Despite some pathological changes of the brain, cerebellum, pituitary, kidney, lung, testis and uterus of each dose group, the regularity, frequency and the degree of injury showed no significant dose-effect relationship in different dose levels. Consider
Histopathological findings: non-neoplastic:
no effects observed
Histopathological findings: neoplastic:
no effects observed

Effect levels

open allclose all
Dose descriptor:
NOEL
Effect level:
200 mg/kg bw/day (nominal)
Based on:
test mat.
Sex:
female
Basis for effect level:
dermal irritation
Dose descriptor:
NOEL
Effect level:
200 mg/kg bw/day (nominal)
Based on:
test mat.
Sex:
male
Basis for effect level:
dermal irritation
Dose descriptor:
NOAEL
Effect level:
> 800 mg/kg bw/day (nominal)
Based on:
test mat.
Sex:
male/female
Basis for effect level:
dermal irritation

Target system / organ toxicity

Critical effects observed:
not specified

Any other information on results incl. tables

For tables and other data, please view the full report.

Applicant's summary and conclusion

Conclusions:
NOAEL determined by the report assessor
>200 mg/kg/d in males
>200 mg/kg/d in females
Executive summary:

Although the Chinese test report suggests a NOAEL of 40 mg/kg/day, there is no evidence for this in the test report and minor blood parameter changes are conisdered adaptive. A NOEL of 200 mg/kg/day is suggested by the reviewer, based on adaptive changes and findings of low statistical significance. It is considered that there were no adverse effects at the highest treated group of 800 mg/kg/day.

The NOAEL is therefore estimated to be > 800 mg/kg/day.