Hexasodium 2,2'-[vinylenebis[(3-sulphonato-4,1-phenylene)imino[6-[(2-cyanoethyl)[2-(2-hydroxyethoxy)ethyl]amino]-1,3,5-triazine-4,2-diyl]imino]]bis[benzene-1,4-disulphonate]

Endpoint summary

Description of key information

Not skin sensitising

Key value for chemical safety assessment

Skin sensitisation

Link to relevant study records

Reference

Endpoint:

skin sensitisation: in vivo (non-LLNA)

Type of information:

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

Adequacy of study:

key study

Study period:

From March the 17th to Apri the 21st, 1989

Reliability:

2 (reliable with restrictions)

Rationale for reliability incl. deficiencies:

test procedure in accordance with generally accepted scientific standards and described in sufficient detail

Justification for type of information:

Justification for read across is detailed in section 13.

Reason / purpose for cross-reference:

read-across source

Qualifier:

according to guideline

Guideline:

OECD Guideline 406 (Skin Sensitisation)

Version / remarks:

adopted on 12 May, 1981

Qualifier:

according to guideline

Guideline:

EU Method B.6 (Skin Sensitisation)

GLP compliance:

yes

Type of study:

guinea pig maximisation test

Justification for non-LLNA method:

An appropriate guinea pig maximisation test was already avaiable, which would not justify conducting an additional LLNA due to animal welfare.

Species:

guinea pig

Strain:

Dunkin-Hartley

Sex:

female

Details on test animals and environmental conditions:

TEST ANIMALS
- Source: Charles River Wiga, Sulzfeld, West Germany.
- Age at start of treatment: ca. 11 weeks.
- Weight at study initiation: 288 - 424 g
- Diet: free access to standard guinea pig diet including ascorbic acid (1600 mg/kg); once a week hay was provided. Certificate of analysis was performed.
- Water: free access to tap-water, diluted with decalcified water. Certificate of analysis was performed.
- Accomodation: group housing of 2 animals per cage with wire-mesh floors (ITL, Bergen).
- Acclimation: at least 5 days, physical examination performed.

ENVIRONMENTAL CONDITIONS
- Temperature: 21 ± 3 °C
- Humidity: 30 - 70 %
- Air changes: 7.5 - 15 ACH
- Photoperiod: 12 hours dark.

Route:

intradermal

Vehicle:

physiological saline

Concentration / amount:

5 % w/w test article

Route:

epicutaneous, occlusive

Vehicle:

water

Concentration / amount:

25 % w/w test article

Day(s)/duration:

48 hours

Route:

epicutaneous, occlusive

Vehicle:

water

Concentration / amount:

vehicle and 25, 10, 5 % test article / 0.05 ml each

Day(s)/duration:

24 hours

No. of animals per dose:

PRELIMINARY TEST: 5 animals
MAIN TEST: 10 animals for control and 20 animals for experimental group

Details on study design:

RANGE FINDING TESTS - PRIMARY IRRITATION EXPERIMENT
- Induction application: intradermal and epidermal
Intradermal injections: 4 sites with 0.1 ml/site at 5 % w/w test article concentration.
Epidermal applications: left flank treated with 0.5 ml of 50 % concentration in milli-RO water; held with Coban elastic bandage for 24 hours.
- Challenge application: left flank treated with 0.05 ml of 50, 25, 10 and 5 % concentration in milli-RO water; held with Coban elastic bandage for 24 hours.
- Symptoms evaluated: erythema/necrosis and diameter for intradermal; checked after 24 and 48 hours.

MAIN STUDY
A1 INDUCTION EXPOSURE - Intradermal injections
- Site: scapular region; 4x6 cm clipped free of hair.
- Injections: 3 pairs at 0.1 ml/site
Test article: 5 % w/w with physiological saline.
FCA: 50:50 with distilled water.
Test article: 10 % w/w emulsified in 50:50 mixture of FCA.

A2 INDUCTION EXPOSURE - Epidermal applications (7 days after intradermal induction)
- Site: same scapular area, 4x6 cm clipped and shaved free of air.
- Concentration: 0.5 ml milli-RO water at 25 % w/w test article
- Bandage: micropore tape firmly secured with Coban elastic bandage.
- Duration: 48 hours
- Evaluation: reaction sites were checked for erythema and oedema immediately after removal of dressing.

B. CHALLENGE EXPOSURE (14 days after epidermal induction)
- Exposure period: 24 hours.
- Test group: 20 females.
- Control group: 10 females; treated as the same way by intradermal and epidermal inductions with the omission of test article.
- Site: 5x5 cm area on the left flank of each animal.
- Concentrations: milli-Ro water and 3 test article concentrations at 25, 10, 5 % in milli-RO water.
- Amount: 0.05 ml of each concentration.
- Bandage: micropore tape firmly secured with Coban elastic bandage.
- Evaluation: sites were assessed for swelling and redness 24 and 48 hours after removal. Positive skin reaction (grade 2 or more) were considered signs of sensitization. The sensitization rate by comparison with control group was then calculated.

OTHER
Used scoring system [Klingman et al., 1966].
No skin reaction............................. 0
Red spots.......................................1
Moderate but confluent redness....2
Redness and swelling....................3
Intense reddening and swelling........4

Challenge controls:

A positive control experiment is carried out once a year as a sensitivity check of the test system. The most recent test was carried out in December, 1988; performed under GLP conditions with a QA-check.

Positive control substance(s):

yes

Remarks:

formaldehyde

Positive control results:

Clearly positive results (90 % sensitisation rate) were obtained in the exporimental animals after challenge with 0.5 % w/w formaldehyde.

Reading:

2nd reading

Hours after challenge:

48

Group:

test chemical

Dose level:

25 % test article

No. with + reactions:

1

Total no. in group:

20

Remarks on result:

no indication of skin sensitisation

Remarks:

No additional animal showed a numerical score value higher than 2.

Reading:

2nd reading

Hours after challenge:

48

Group:

negative control

No. with + reactions:

0

Total no. in group:

10

Remarks on result:

no indication of skin sensitisation

Reading:

2nd reading

Hours after challenge:

48

Group:

positive control

Dose level:

0.5 %

No. with + reactions:

18

Total no. in group:

20

Remarks on result:

positive indication of skin sensitisation

Summary table of positive reactions to the challenge

	Test article concentration
	25 %	10 %	5 %	0 %
EXPERIMENTAL GROUP
No. animals with positive reaction	1	0	0	0
Sensitisation rate	5	0	0	0
CONTROL GROUP
No. animals with positive reaction	0	0	0	0

Individual induction and challenge readings

EXPERIMENTAL GROUP
Animal no.	Readings of induction (day 10)			Challenge								Sensitised
				1st reading				2nd reading
	Erythema	Oedema		25 %	10 %	5 %	0 %	25 %	10 %	5 %	0 %
306	0	1		0	0	0	0	1	0	0	0	no
307	0	1		2	0	0	0	1s	0	0	0	yes
308	1	1		0	0	0	0	1	0	0	0	no
309	0	0		0	0	0	0	0	0	0	0	no
310	1	1		0	0	0	0	1s	1	1s	0	no
311	1	2		0	0	0	0	0	0	0	0	no
312	0	0		1	0	0	0	1s	0	0	0	no
313	0	0		1	0	0	0	0	0	0	0	no
314	2	2		0	0	0	0	0	0	0	0	no
315	1	1		1	1	0	0	1s	1	0	0	no
316	0	0		1	0	0	0	1cs	0	0	0	no
317	1	1		0	0	0	0	0	0	0	0	no
318	2	2		0	0	0	0	0s	0	0	0	no
319	2	1		0	0	0	0	0s	0	0	0	no
320	1	1		0	0	0	0	0	0	0	0	no
321	1	1		1	0	0	0	1s	0	0	0	no
322	0	0		0	0	0	0	0	0	0	0	no
323	1	1		1	0	0	0	1s	0	0	0	no
324	1	1		0	0	0	0	1	0	0	0	no
325	0	0		0	0	0	0	0	0	0	0	no
CONTROL GROUP
Animal no.	Readings of induction (day 10)			Challenge								Sensitised
				1st reading				2nd reading
	Erythema	Oedema		25 %	10 %	5 %	0 %	25 %	10 %	5 %	0 %
326	0	0		0	0	0	0	1s	0	0	0	no
327	0	0		0	0	0	0	0	0	0	0	no
328	0	0		0	0	0	0	1s	0	0	0	no
329	0	0		0	0	0	0	0	0	0	0	no
330	0	0		0	0	0	0	1	0	0	0	no
331	0	0		1	0	0	0	1s	0	0	0	no
332	0	0		0	0	0	0	0	0	0	0	no
333	0	0		0	0	0	0	0	0	0	0	no
334	0	0		0	0	0	0	1s	0	0	0	no
335	0	0		0	0	0	0	1	0	0	0	no

Legend:

s = scaliness

c= crust formation

PRIMARY IRRITATION EXPERIMENT

25% (w/w) was chosen as the highest usable test article concentration, due to the fact that the 50% concentration was a much to solid suspension. No signs of systemic toxicity were observed during the primary irritation experiments, except of body weight loss in one animal.

OTHER FINDINGS

The average body weight gain of the experimental animals was approximately 30 % smaller than the average body weight gain of the control animals. However, body weights measured during the acclimatisation period were noted as being slightly low for animals assigned to the control group. These animals would therefore be expected to gain more weight than the animals assigned to the experimental group. Differences in body weight gain are therefore not considered to have arisen as a result of treatment.

No symptoms of systemic toxicity were observed.

No positive skin reaction were evident after the challenge exposure.

Positive control results (1988 test)

	Formaldehyde concentration
	0.50%	0.25%	0.10%	0%
EXPERIMENTAL GROUP
No. Animals with positive reaction	18	7	1	0
Sensitisation rate	90	35	5	0
CONTROL GROUP
No. Animals with positive reaction	0	0	0	0

Interpretation of results:

other: not classified, according to the CLP Regulation (EC) 1272/2008

Conclusions:

Not skin sensitising

Executive summary:

The Guinea Pig Maximisation Test was performed according to OECD 406 (1981). A total of 35 females were used, 20 for the main study and 10 for the control. The other 5 animals were used for the primary irritation experiment.

The induction phase was accomplished by both intradermal and epicutaneous application. In the challenge phase, the test substance resulted in a sensitisation rate of 5 %, when tested at the maximum testable epidermal application of 25 % w/w test article.

Conclusion

Less than 30 % of the tested population showed positive reactions, responding to an intradermal induction dose higher than 1 %.

Endpoint conclusion

Endpoint conclusion:

no adverse effect observed (not sensitising)

Additional information:

There is no information about the skin sensitisation potential of Optical Brightener 380, thus the available data on the structural analogous Similar Substance 01 have been taken into consideration for the assessment.

The read across approach can be considered reliable and appropriate to investigate the property (details for the approach are included into the IUCLID section 13).

The Guinea Pig Maximisation Test was performed according to OECD 406 (1981). A total of 35 females were used, 20 for the main study and 10 for the control. The other 5 animals were used for the primary irritation experiment.

The induction phase was accomplished by both intradermal and epicutaneous application. In the challenge phase, the test substance resulted in a sensitisation rate of 5 %, when tested at the maximum testable epidermal application of 25 % w/w test article.

Respiratory sensitisation

Endpoint conclusion

Endpoint conclusion:

no study available

Justification for classification or non-classification

According to the CLP Regulation (EC) No 1272/2008, 3.4 Respiratory or skin sensitisation section, skin sensitizer means a substance that will lead to an allergic response following skin contact.

Based on the Guinea Pig Maximisation Test (GPMT) results, a substance in considered a skin sensitizer when equal or more than 30 % to 60 % responding at intradermal induction dose > 0.1 % to ≤ 1 %; or equal or more than 30 % responding at intradermal induction dose higher than 1 %.

In the available experiment, less than 30 % of the tested population showed positive reactions, responding to an intradermal induction dose higher than 1 %.

In conclusion, Optical Brightener 380 is considered to not meet the criteria to be classified as skin sensitizer, according to the CLP Regulation (EC) No 1272/2008.