Registration Dossier

Administrative data

Endpoint:
acute toxicity: dermal
Type of information:
experimental study
Adequacy of study:
key study
Study period:
jan-april 2010
Reliability:
1 (reliable without restriction)
Rationale for reliability incl. deficiencies:
guideline study

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 402 (Acute Dermal Toxicity)
Deviations:
no
Qualifier:
according to
Guideline:
EU Method B.3 (Acute Toxicity (Dermal))
Deviations:
no
Qualifier:
according to
Guideline:
EPA OPPTS 870.1200 (Acute Dermal Toxicity)
Deviations:
no
Qualifier:
according to
Guideline:
other: Japanese Ministry of Agroculture, Forestry and Fisheries (JMAFF), 12 Nousan, Notification No 8147, November 2000; including the most recent partial revisions.
Deviations:
no
GLP compliance:
yes (incl. certificate)
Test type:
standard acute method
Limit test:
no

Test material

Reference
Name:
Unnamed
Type:
Constituent
Type:
Constituent
Details on test material:
- Name of test material (as cited in study report): VC
- Molecular formula (if other than submission substance): C3H2O3
- Molecular weight (if other than submission substance): 86.05
- Physical state: clear colourless solid (at storage conditions)
- Analytical purity: >99.9%
- Batch No.: 091014
- Expiration date of the batch: 31 December 2010
- Test material will be warmed at 25°C in a warming bath to produce a liquid before use.

Test animals

Species:
rat
Strain:
other: Wistar strain, Crl:WI (Han)
Sex:
male/female
Details on test animals and environmental conditions:
TEST ANIMALS
- Source: Charles River Deutschland, Germany
- Age at study initiation: 10-12 weeks old
- Weight at study initiation: body weight variation did not exceed +/-20% of the sex mean.
- Housing: Individually housed in labeled Macrolon cages (MIII type, height 18 cm) containing sterilized sawdust as bedding material (Litalabo, S.P.P.S., Argenteuil, France) and paper as cage-enrichment (Enviro-dri, Wm. Lillico & Son (Wonham Mill Ltd), Surrey, United Kingdom).
- Diet (e.g. ad libitum): Free access to pelleted rodent diet (SM R/M-Z from SSNIFF® Spezialdiäten GmbH, Soest, Germany).
- Water (e.g. ad libitum): Free access to tap water.
- Acclimation period: The acclimatization period was at least 5 days before the start of treatment under laboratory conditions.
- Health inspection: A health inspection was performed prior to treatment, to ensure that the animals were in a
good state of health. Special attention was paid to the skin to be treated, which was intact and free from any
abnormality.

Results of analysis for diet (nutrients and contaminants), sawdust, paper and water were assessed and did not
reveal any findings that were considered to have affected the study integrity. All certificates and results of analysis
are retained in the NOTOX archives.

ENVIRONMENTAL CONDITIONS
- Temperature (°C): 20-22 (actual range)
- Humidity (%): 27-62 (actual range)
- Air changes (per hr): 15
- Photoperiod (hrs dark / hrs light): 12/12

IN-LIFE DATES: From: 13 January 2010 To: 13 April 2010

Administration / exposure

Type of coverage:
occlusive
Vehicle:
propylene glycol
Remarks:
vehicle used for treatment at 200 mg/kg bw; The test substance was doses undiluted to animals at 2000 mg/kg bw.
Details on dermal exposure:
TEST SITE
- Area of exposure: (25 cm2 for males and 18 cm2 for females)
- % coverage: 10%
- Type of wrap if used: surgical gauze (Surgy 1D) successively covered with aluminium foil and Coban elastic bandage. A piece of Micropore tape was additionally used for fixation of the bandages in females only.

REMOVAL OF TEST SUBSTANCE
- Washing (if done): tap water
- Time after start of exposure: 24 hours

TEST MATERIAL
- Amount(s) applied (volume or weight with unit): 1.471 mL/kg (2000 mg/kg bw) or 10 mL/kg (200 mg/kg bw)

One day before exposure (Day -1) an area of approximately 5x7 cm on the back of the animal was clipped.
Duration of exposure:
24 hours.
Doses:
2000 mg/kg bw
200 mg/kg bw
No. of animals per sex per dose:
5/sex/dose
Control animals:
not required
Details on study design:
VEHICLE
- Justification for choice of vehicle: The vehicle was selected based on trial formulations performed at NOTOX and
on test substance data supplied by the sponsor.

- Duration of observation period following administration: 14 days
- Frequency of observations and weighing: observations once daily; body weights on days 1 (pre-administration), 8 and 15 and at death.
- Necropsy of survivors performed: yes

-Test substance preparation: prior to weighing, the test substance was heated in a water bath with a maximum temperature of 25ºC for a maximum of 1 hour and 45 minutes. For treatment at 200 mg/kg bw, formulations (w/w) were prepared immediately prior to dosing. Homogeneity was accomplished to a visually acceptable level. Adjustment was made for specific gravity of the vehicle and of the test substance. Prior to dosing, the test substance and formulations were heated in a water bath with a maximum temperature of 25ºC for a maximum of 1 hour and 23 minutes in order to obtain homogeneity.
Statistics:
none

Results and discussion

Effect levels
Sex:
male/female
Dose descriptor:
LD50
Effect level:
> 200 - < 2 000 mg/kg bw
Based on:
test mat.
Mortality:
All animals at 2000 mg/kg bw were found dead or sacrificed in moribound condition on Day 3. No mortality occured at 200 mg/kg bw.
Clinical signs:
2000 mg/kg bw: restless behaviour, lethargy, general tremors, flat or hunched posture, uncoordinated movements, laboured respiration, shallow respiration, piloerection, water discharge from the eyes, chromodacryorrhoea (snout), ptosis and/or hypothermia.
200 mg/kg: restless behaviour, lethargy, general tremors, flat or hunched posture, shallow respiration, piloerection, chromodacryorrhoea (snout) and/or ptosis.
The animals at 200 mg/kg bw had fully recovered from the abovementioned symptoms between days 4 and 9.
General erythema, necrosis, a thickened area and/or yellow staining were seen in the treated skin-area of animals at 2000 mg/kg bw during the observation period.
Animals at 200 mg/kg bw showed a combination of the following findings in the treated skin-area during the observation period: general, maculate or focal erythema, scars, fissures, scales, scabs, a thickened area, yellow staining and dryness.
Body weight:
The changes noted in body weight gain in males and females at 200 mg/kg bw were within the range expected for rats used in this type of study.
Gross pathology:
Macroscopic post mortem examination of the animals at 2000 mg/kg bw revealed gelatinous subcutis of the skin and/or dark red foci on the glandular mucosa of the stomach.
No abnormalities were found at macroscopic post mortem examination of the animals at 200 mg/kg bw.

Applicant's summary and conclusion

Interpretation of results:
other: toxic
Conclusions:
The dermal LD50 value of VC in Wistar rats was established to be within the range of 200-2000 mg/kg bw.
Executive summary:

Acute dermal toxicity of VC was assessed in the rat according to OECD 402.

The dermal LD50 value of VC in Wistar rats was established to be within the range of 200-2000 mg/kg bw.

Based on the mortality at 2000 mg/kg bw and on the clinical signs at 200 and 2000 mg/kg bw, no animals were treated at 1000 mg/kg bw for ethical reasons. Therefore, as a worst case scenario and for labeling and classification purposes, the dermal LD50 value of VC in Wistar rats was considered to be within the range of 200-1000 mg/kg bw.

Based on these results and considerations:

-According to the Globally Harmonized System of Classification and Labeling of Chemicals (GHS) of the United Nations (2007), VC should be classified as: Toxic in contact with skin (Category 3) for acute toxicity by the dermal route.

-According to the Regulation (EC) No 1272/2008 on classification, labeling and packaging of substances and mixtures, VC should be classified as Category 3 and should be labeled as H311: Toxic in contact with skin.