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Toxicological information

Skin sensitisation

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Administrative data

Endpoint:
skin sensitisation: in vivo (non-LLNA)
Type of information:
experimental study
Adequacy of study:
key study
Study period:
From April 29 to June 15, 1996
Reliability:
2 (reliable with restrictions)
Rationale for reliability incl. deficiencies:
guideline study with acceptable restrictions

Data source

Reference
Reference Type:
study report
Title:
Unnamed
Year:
1996
Report Date:
1996

Materials and methods

Test guideline
Qualifier:
according to
Guideline:
OECD Guideline 406 (Skin Sensitisation)
Deviations:
yes
Remarks:
no certificate of analysis, no data on animal gender and purity of test substance
GLP compliance:
yes (incl. certificate)
Type of study:
guinea pig maximisation test
Justification for non-LLNA method:
The guinea-pigs study was performed before the REACH regulation.

Test material

Reference
Name:
Unnamed
Type:
Constituent
Test material form:
other: Colourless liquid
Details on test material:
- Name of test material : TMPTMA
- Physical state: Colourless liquid

In vivo test system

Test animals

Species:
guinea pig
Strain:
Hartley
Sex:
not specified
Details on test animals and environmental conditions:
TEST ANIMALS
- Source: David Hall Limited, Burton-on-Trent, Staffordshire, UK
- Age at study initiation: 8-12 weeks
- Weight at study initiation: 312-490 g
- Housing: Housed singly or in pairs in solid-floor polypropylene cages
- Diet (e.g. ad libitum): Guinea Pig FD1 Diet (Special Diets Services Limited, Witham, UK), ad libitum
- Water (e.g. ad libitum): Mains tap water, ad libitum
- Acclimation period: 5 days

ENVIRONMENTAL CONDITIONS
- Temperature (°C): 20-23 °C
- Humidity (%): 45-68%
- Air changes (per hour): 15/hour
- Photoperiod (hours dark / hours light): 12 hours dark / 12 hours light

Study design: in vivo (non-LLNA)

Inductionopen allclose all
Route:
intradermal and epicutaneous
Vehicle:
arachis oil
Concentration / amount:
Range finding test:
- Intradermal induction exposure: 1, 5, 10 or 25% w/v in arachis oil
- Topical induction exposure: 25, 50, 75 or 100% w/v in arachis oil
- Topical challenge exposure: 25, 50, 75 or 100% w/v in arachis oil

Main test:
- Intradermal induction exposure: 25% w/v in arachis oil
- Topical induction exposure: Undiluted
- Topical challenge exposure: 75 or 100% w/v in arachis oil
Challengeopen allclose all
Route:
epicutaneous, occlusive
Vehicle:
arachis oil
Concentration / amount:
Range finding test:
- Intradermal induction exposure: 1, 5, 10 or 25% w/v in arachis oil
- Topical induction exposure: 25, 50, 75 or 100% w/v in arachis oil
- Topical challenge exposure: 25, 50, 75 or 100% w/v in arachis oil

Main test:
- Intradermal induction exposure: 25% w/v in arachis oil
- Topical induction exposure: Undiluted
- Topical challenge exposure: 75 or 100% w/v in arachis oil
No. of animals per dose:
Range finding test:
- Intradermal induction exposure: 1 animal/dose
- Topical induction and challenge exposure: 2 animals/dose

Main test: 10 and 20 animals for control and test, respectively
Details on study design:
RANGE FINDING TESTS:
- Intradermal induction exposure: Guinea pigs (1/dose) received injections (0.1 mL) of test material at concentrations of 1, 5, 10 or 25% w/v in arachis oil and observed for systemic toxicity and erythema scores (Draize scoring system) at 24, 48 and 72 hours and 7 days.
- Topical induction exposure: Two guinea pigs [intradermally injected with Freund's Complete Adjuvant (FCA) 16 days earlier] applied with test material at concentrations of 25, 50, 75 or 100% v/v in arachis oil for 48 hours and observed for erythema and oedema at 1, 24 and 48 hours.
- Topical challenge exposure: Two guinea pigs applied with test material at concentrations of 25, 50, 75 or 100% v/v in arachis oil for 24 hours and observed for erythema and oedema at 1, 24 and 48 hours.


MAIN STUDY
A. INDUCTION EXPOSURE: INTRADERMAL
- No. of exposures: One
- Test groups: Intradermally injected with 0.1 mL of FCA, 25% w/v test material in arachis oil and 25% w/v test material emulsion in FCA on Day 0
- Control group: Intradermally injected with FCA, arachis oil or 50% w/v arachis oil in FCA on Day 0
- Site: Shoulder region on each side of mid-line
- Duration: Days 0-6

B. INDUCTION EXPOSURE: TOPICAL
- No. of exposures: One
- Exposure period: 48 hours
- Test groups: Filter paper patch saturated with undiluted test material topically applied on Day 7 via occlusive patch
- Control group: Only filter paper patch topically applied on Day 7 via occlusive patch
- Site: Shoulder region on each side of mid-line
- Frequency of applications: Single application
- Duration: Days 7-21

C. CHALLENGE EXPOSURE: TOPICAL
- No. of exposures: One
- Day of challenge: Day 21
- Exposure period: 24 hours
- Test groups: Filter paper patch saturated with 75 or 100% v/v test material in arachis oil topically applied on Day 21 via occlusive patch
- Site: 75 or 100% v/v in arachis oil was applied to left or right shorn flank, respectively
- Evaluation (hour after removal of challenge patch): 24 or 48 hours
Challenge controls:
No data
Positive control substance(s):
yes
Remarks:
Historical data: Ethyl 4-aminobenzoate 98%; 2,4-Dinitrochlorobenzene; Neomycin sulphate; 2-Mercaptobenzothiazole

Results and discussion

Positive control results:
Historical data (August 1994 to February 1996):
- Incidence of sensitisation for 98% ethyl 4-aminobenzoate, 2,4-dinitrochlorobenzene, neomycin sulphate, 2-mercaptobenzothiazole were 39% (7/18), 100% (9/9), 100% (10/10), 60% (12/20) or 70% (7/10), respectively

In vivo (non-LLNA)

Resultsopen allclose all
Reading:
1st reading
Hours after challenge:
24
Group:
test group
Dose level:
75 or 100% v/v in arachis oil
No. with + reactions:
0
Total no. in group:
20
Clinical observations:
None
Remarks on result:
no indication of skin sensitisation
Reading:
2nd reading
Hours after challenge:
48
Group:
test group
Dose level:
75or 100% v/v on arachis oil
No. with + reactions:
0
Total no. in group:
20
Clinical observations:
None
Remarks on result:
no indication of skin sensitisation
Reading:
1st reading
Hours after challenge:
24
Group:
negative control
No. with + reactions:
0
Total no. in group:
10
Remarks on result:
no indication of skin sensitisation
Reading:
1st reading
Group:
positive control
No. with + reactions:
7
Total no. in group:
10
Remarks on result:
positive indication of skin sensitisation
Remarks:
(2-mercaptobenzothiazole)

Any other information on results incl. tables

Table 1: Intradermal sighting test - summary of results

 

Animal identification

Time of observation

Concentration of test material (% w/v)

Grade of erythema at Injection Sites

Evidence of systemic toxicity

A

24 hours

1

1

None

48 hours

2

None

72 hours

3

None

7 days

0

None

B

24 hours

5

1

None

48 hours

2

None

72 hours

3

None

7 days

0

None

E

24 hours

10

1

None

48 hours

1

None

72 hours

1

None

7 days

1

None

F

24 hours

25

2

None

48 hours

2

None

72 hours

2

None

7 days

1

None

Conclusion: The concentration of the test material selected for the intradermal induction stage of the main study was 25% w/v in arachis oil BP

  

Table 2: Topical sighting test for induction application (48-hour exposure) - individual skin reactions

 

Animal identification

Concentration of test material (% v/v)

Skin Reactions (Hours After Removal of Patches)

1

24

48

Er

Oe

Other

Er

Oe

Other

Er

Oe

Other

G

100

2

1

-

1

0

-

0

0

-

75

2

1

-

1

0

-

0

0

-

50

2

0

-

0

0

-

0

0

-

25

1

0

-

0

0

-

0

0

-

H

100

2

0

-

2

0

-

1

0

-

75

2

0

-

1

0

-

0

0

-

50

1

0

-

0

0

-

0

0

-

25

0

0

-

0

0

-

0

0

-

Conclusion: The undiluted test material was selected for the main study topical induction

 

Table 3: Topical sighting test for challenge application (24-hour exposure) - individual skin reactions

Animal identification

Concentration of test material (% v/v)

Skin Reactions (Hours After Removal of Patches)

1

24

48

Er

Oe

Other

Er

Oe

Other

Er

Oe

Other

I

100

1

0

-

0

0

-

0

0

-

75

1

0

-

0

0

-

0

0

-

50

0

0

-

0

0

-

0

0

-

25

0

0

-

0

0

-

0

0

-

J

100

1

0

-

0

0

-

0

0

-

75

1

0

-

0

0

-

0

0

-

50

0

0

-

0

0

-

0

0

-

25

0

0

-

0

0

-

0

0

-

Er = erythema; Oe = oedema; - = no other reactions noted

 

Conclusion: The concentrations of the test material selected for the main study topical challenge were 100 and 75% v/v in arachis oil BP

Applicant's summary and conclusion

Interpretation of results:
GHS criteria not met
Remarks:
not skin sensitizing
Conclusions:
Under these test conditions, TMPTMA is not a skin sensitiser and is not classified according to the CLP Regulation (EC) N° (1272-2008).
Executive summary:

In a Magnusson & Kligman maximisation study (GPMT) performed according to OECD guideline 406 and in compliance with GLP, groups of 20 albino Dunkin Hartley guinea pigs were intradermally induced with three injections of 0.1 mL of FCA, 25% w/v TMPTMA in arachis oil and 25% w/v TMPTMA emulsion in FCA on Day 0 on shoulder region on each side of mid-line. After one week the same area were topically induced with 0.2 mL of undiluted test material via occluded filter paper patch for 48 hours. After 2 weeks of rest period, a challenge patch of 75 or 100% v/v test material in arachis oil was applied to left or right shorn flank, respectively. Control groups of 10 animals was included and treated with FCA, arachis oil or 50% w/v arachis oil in FCA. The test concentrations for the main study were determined from a sighting study using two animals.

 

Intradermal and topical induction indicated evidence of dermal irritation. No skin reactions were noted at the challenge sites of the test or control group animals at the 24 or 48-hour observations. TMPTMA produced a 0% (0/20) sensitisation rate and was considered to be a nonsensitiser to guinea pig skin. Historical data on positive controls (ethyl 4-aminobenzoate 98%; 2,4-dinitrochlorobenzene; neomycin sulphate; 2-mercaptobenzothiazole) exhibited evidence of sensitisation, thus confirming the susceptibility of this group of animals to dermal sensitisation.

 

Under these test conditions, TMPTMA is not classified according to the Annex VI to the Directive 67/548/EEC and the CLP Regulation (EC) N° (1272-2008).