2-methyl-p-phenylenediamine

Skin irritation / corrosion

Administrative data

Endpoint:

skin irritation / corrosion

Remarks:

in vitro

Type of information:

migrated information: read-across from supporting substance (structural analogue or surrogate)

Adequacy of study:

key study

Study period:

From June 06, 2012 to June 07, 2012

Reliability:

2 (reliable with restrictions)

Rationale for reliability incl. deficiencies:

other: Study well documented, followed guideline, GLP

Data source

Reference

Reference Type:

study report

Title:

Unnamed

Year:

2012

Report date:

2012

Materials and methods

Test guideline

Qualifier:

according to guideline

Guideline:

OECD Guideline 430 (In Vitro Skin Corrosion: Transcutaneous Electrical Resistance Test Method (TER))

Deviations:

no

GLP compliance:

yes (incl. QA statement)

Remarks:

(according to UK and OECD principles of GLP)

Test material

Test material information

Constituent 1

Reference substance name:

2-methyl-p-phenylenediamine sulfate

EC Number:

210-431-8

EC Name:

2-methyl-p-phenylenediamine sulfate

Cas Number:

615-50-9

Molecular formula:

C7H10N2.H2O4S

IUPAC Name:

2-methyl-p-phenylenediamine sulphate

Constituent 2

Reference substance name:

Toluene-2,5-diamine sulfate

IUPAC Name:

Toluene-2,5-diamine sulfate

Test material form:

solid: crystalline

Details on test material:

- Name of test material: Toluene-2,5-diamine sulfate (Code # A000165)- TSIN: 23005- Substance type: Pure active substance- Physical state: Slightly reddish fine crystalline powder- Stability under test conditions: The substance on ambient storage in dryness and darkness was considered stable for more than 10 years.- Stability in solution: The solution of test material in water (99.9% and 109%) and in water/acetone 4:1 v/v (99.7% and 108%) confirmed a good stability over a time period of 8 days.- Storage condition of test material: Room temperature in the dark- Solubility: Solubility in different solvents is as follows: 1.0 weight% in DMSO 0.16 weight% in acetonitrile 5.03 g/L (pH 2.47) in water 1:1 0.6 weight% (pH 7) in acetone/water 1:1 0.3 weight% in ethanol

Test animals

Species:

rat

Strain:

Wistar

Details on test animals or test system and environmental conditions:

TEST ANIMALS- Source: One (RccHanTM:WIST) strain rat was obtained from Harlan Laboratories UK Limited, Bicester, Oxon, UK.- Age at study initiation: 21- 23 daysThe animal was in the telogen phase of hair growth and little or no hair growth was visible.- Sex: Female- Weight at study initiation: Not reported- Acclimation period: 2 daysEXPERIMENT INITIATION DATE: June 06, 2012EXPERIMENT COMPLETION DATE: June 07, 2012

Test system

Preparation of test site:

shaved

Vehicle:

unchanged (no vehicle)

Controls:

no

Amount / concentration applied:

Sufficient neat test material to cover the epidermal surface was used in this study.

Duration of treatment / exposure:

Contact period of 24 hours

Observation period:

24 hours

Number of animals:

One

Details on study design:

PRE-TEST PROCEDURES: - Skin disk preparation: The animal was shaved to remove hair from the dorsal surface. The shaved area was washed using an antibiotic wash and a second antibiotic wash was performed after 3 days. 2 days later, the animal was sacrificed using carbon dioxide followed by cervical dislocation. The dorsal skin of animal was removed as a single pelt. After removal of excess fat, the pelt was mounted epidermal side uppermost, onto a polytetrafluoroethylene (PTFE) tube. The tissue was secured in place using a rubber "O" ring. Excess tissue was trimmed away and the "O" ring/PTFE interface was sealed with soft paraffin wax. The tube was supported by a clamp inside a labeled 30 mL glass receptacle containing 10 mL electrolyte solution (154 mM MgSO4).- Skin disk quality control: Two skin discs of approximately 0.79 cm2 and a resistance value of greater than 10 kΩ were taken from the pelt and the TER (Transcutaneous Electrical Resistance) was measured as a quality control procedure. TEST PROCEDURE:- Number of replicates: 3 - Controls: 3 positive (approximately 36% hydrochloric acid) and 3 negative control (sterile distilled water) discs were assayed.- Test material or negative /positive control administration: Test material/negative control/positive control was applied for 24 hours to cover the epidermal surface of skin disk. 150 µL of distilled water was applied with test material to ensure good contact with the skin. - Removal of test substance: After 24 hours of exposure the test material was removed by washing the skin disc with a jet of warm tap water until no further test material could be removed. - Determination of Transcutaneous Electrical Resistance (TER): The TER was measured using a Wheatstone Bridge with a low voltage alternating current. Prior to measurement of the resistance, the surface tension of the skin disc was reduced by adding a sufficient volume of 70% ethanol to cover the epidermis. After the removal of ethanol, the PTFE tube was placed in the labeled receptor chamber and the tissue was hydrated by the addition of 3 mL MgSO4 solution (154 mM) to the inside of the PTFE tube. The stainless steel electrodes of the databridge were placed on either side of the skin disc. The measurement was taken and a value in Ω/kΩ per skin disc was displayed on the databridge display. The mean TER for the skin discs was calculated.

Results and discussion

In vivo

Results

Irritation parameter:

other: Transcutaneous Electrical Resistance (TER)

Basis:

mean

Time point:

other: 24 hours

Remarks on result:

other: Mean TER value of test material: 17.0 kΩ (Above the cut off TER value of 5 kΩ)

Any other information on results incl. tables

SKIN DISK QUALITY CONTROL RESULT: Two skin discs were evaluated for quality control. Each disc had to give a resistance value of greater than 10kΩ in order for the remainder of the pelt to be used in the assay. If either disc fell below the 10 kΩ threshold, the pelt was discarded. The quality control discs were then discarded and new discs from the acceptable pelt were mounted on the PTFE tubes.

In this study, the TER (Transcutaneous Electrical Resistance) of quality control skin disks were as follows:

Skin disk 1: 15.5 kΩ

Skin disk 2: 21.2 kΩ

TRANSCUTANEOUS ELECTRICAL RESISTANCE ASSAY RESULTS:

The individual and mean value of three TER measurement for test material, positive control and negative control after a 24 hours contact period are provided in below table:

Table 1: Individual and Mean TER (Transcutaneous Electrical Resistance) measurements (Study # 78116)

Treatment	Tissue Number	TER	Mean TER	Standard Deviation
Toluene-2,5-diamine sulfate	1	14.8 kΩ	17.0 kΩ	± 6.6
	2	24.4 kΩ
	3	11.8 kΩ
Positive Control (approx. 36% Hydrochloric acid)	4	753 Ω	723 Ω	±27.8
	5	698 Ω
	6	718 Ω
Negative Control (Sterile distilled water)	7	13.6 kΩ	15.7 kΩ	± 3.3
	8	19.5 kΩ
	9	14.0 kΩ

Applicant's summary and conclusion

Interpretation of results:

other: non corrosive

Remarks:

Criteria used for interpretation of results: expert judgment

Conclusions:

Toluene-2,5-diamine sulfate was considered non corrosive in an in-vitro skin corrosion (Transcutaneous Electrical Resistance (TER)) assay.

Executive summary:

The in-vitro skin corrosion test of Toluene-2,5-diamine sulfate was performed following the OECD 430 guideline (In-Vitro Skin Corrosion: Transcutaneous Electrical Resistance Test (TER)).

The dorsal skin of one female Wistar rat (Harlan Laboratories UK Limited, UK) was removed as a single pelt for test application. The integrity of the pelt was confirmed using a Quality Control test. The test material (neat) was applied to the epidermal surface of skin discs for a contact period of 24 hours. 150 µL of distilled water was applied with test material to ensure good contact with the skin.
At the end of the contact period the test material was removed using a jet of warm tap water.

Undiluted test material, the positive control (36% HCl) and the negative control (sterile distilled water) were tested in triplicate. The skin discs were evaluated for TER using a low voltage alternating current electronic databridge (Wheatstone Bridge). The measurements were taken in Ω/kΩ per skin disc and the mean TERs for the skin discs were calculated. Test materials that give a mean electrical resistance of 5 kΩ or less are considered likely to be corrosive.

The mean TER value recorded for Toluene-2,5-diamine sulfate after a 24 hour contact period was 17.0 kΩ (above the cut off value of 5 kΩ). The mean TER results of the concurrent negative and positive control discs were within the acceptable range (positive control: 723 Ω; negative control: 15.7 kΩ).

Based on above, Toluene-2,5-diamine sulfate was considered non corrosive in Transcutaneous Electrical Resistance (TER) assay (in-vitro). 

This in-vitro skin corrosion test is classified as acceptable, and satisfies the guideline requirements of the OECD 430 method.