Registration Dossier

Administrative data

Description of key information

The magnesium sulfonate read across substance (CAS 71768-47-5) was investigated for its sensitizing potential after repeated topical exposures in guinea pigs in a skin sensitization test according to the Buehler method (van Huygevoort, 2012, according to OECD 406, EU Method B6 and EPA OPPTS 870.2600, Key study, Klimisch 2).

According to Ball and colleagues, the LLNA typically used for this endpoint tends to overestimate the sensitization potential of surfactants (Ball et al., 2011). Since its introduction, there have been numerous publications reporting apparent false positive results in the LLNA for certain groups of chemicals, including surfactants (Garcia et al., 2010; Kreiling et al., 2008; Woolhiser et al., 1998; all cited in Ball et al., 2011).In addition, Basketter and Kimber were able to confirm that the utilization of the LLNA may lead to false positive results (Basketter and Kimber, 2011).

Hence, the test item triggered the conduct of a Buehler sensitization test.Test substance concentrations selected for the main study were based on the results of a preliminary study. In the main study, twenty experimental animals were epidermally treated on three occasions (once weekly, on Days 1, 8 and 15) with the undiluted test substance (100 % concentration) and ten control animals were similarly treated, but with vehicle alone (Kaydol White Mineral Oil). Two weeks after the last induction exposure (rest period), all animals were topically challenged with a 50% test substance concentration and the vehicle. A second challenge was performed approximately one week later with the same test substance concentrations and the vehicle.

Skin reactions were observed in response to a 50% concentration ofthe magnesium sulfonate read across substance (CAS71786-47-5) in three of the twenty experimental animals (one grade 1 skin reaction and two observations of scaliness) in the first challenge phase. These skin reactions were not confirmed at second challenge (one week later) with the same concentration.

So the number of animals exhibiting positive sensitization scores of 1 for the test substance at the 50% challenge and 50% rechallenge were 1/20 and 0/20, respectively (mean 24/48 hour Draize severity scores 0.00/0.05 and 0.0/0.0). No positive scores were observed in the vehicle control in the first and second challenge (mean 24/48 hour Draize severity scores 0.0/0.0). The only other observation was scaliness in two animals with scores of 0 in the first challenge with the 50% test material concentration and/or vehicle at the 48 hour reading.

The six-month reliability check with Alpha-hexylcinnamicaldehyde indicates that the Buehler Test as performed at WIL Research Europe is an appropriate model for testing for contact hypersensitivity.Based on these results the test substance is not considered to be a skin sensitizer.


Migrated from Short description of key information:
Read across: CAS 71786-47-5, Skin sensitisation according to the Buehler method, guinea pigs, not sensitising

Justification for selection of skin sensitisation endpoint:
well documented GLP-guideline study

Key value for chemical safety assessment

Skin sensitisation

Link to relevant study records

Referenceopen allclose all

Endpoint:
skin sensitisation: in vivo (non-LLNA)
Type of information:
experimental study
Adequacy of study:
key study
Study period:
2012-06-13 - 2012-11-12
Reliability:
2 (reliable with restrictions)
Rationale for reliability incl. deficiencies:
other: see 'Remark'
Remarks:
The Buehler type of sensitization test is selected at the request of the sponsor since this substance is a surfactant. Studies have shown that the Local Lymph Node Assay, which is typically used, over predicts sensitization potential of these substances. This study is a GPL-guideline study and well documented, however, as the study is used in a read across approach Klimisch 2 is assigned.
Qualifier:
according to
Guideline:
OECD Guideline 406 (Skin Sensitisation)
Deviations:
yes
Remarks:
Temporary deviations from the maximum level for daily mean relative humidity occurred. Evaluation: Laboratory historical data do not indicate an effect of the deviations.
Qualifier:
according to
Guideline:
EU Method B.6 (Skin Sensitisation)
Deviations:
no
Qualifier:
according to
Guideline:
EPA OPPTS 870.2600 (Skin Sensitisation)
Deviations:
no
Qualifier:
according to
Guideline:
other: Japanese Ministry of Agriculture, Forestry and Fisheries (JMAFF), 12 Nohsan, Notification No. 8147, April 2011; including the most recent partial revisions.
Deviations:
no
GLP compliance:
yes (incl. certificate)
Remarks:
Food and Consumer Product Safety Autrhority (VWA)
Type of study:
Buehler test
Justification for non-LLNA method:
Study was available on a surrogate substance.
Species:
guinea pig
Strain:
Dunkin-Hartley
Sex:
female
Details on test animals and environmental conditions:
TEST ANIMALS
- Source: Charles River Deutschland, Kisslegg, Germany
- Age at study initiation: young adult animals (approx. 5 weeks old), females were nulliparous and non-pregnant
- Housing: Group housing of maximally 5 animals per labelled Noryl cage (Tecniplast; 74 cm x 54 cm x 25 cm height) containing sterilized sawdust as bedding material (Litalabo, S.P.P.S., Argenteuil, France) and shelters (CS3B02A Play tunnels (90 mm x 5 mm x 125 mm), Datesand, Manchester, UK) as cage enrichment.
- Diet (e.g. ad libitum): Complete maintenance diet for guinea pigs (SSNIFF® MS-H or MS-H Ered; SSNIFF® Spezialdiäten GmbH, Soest, Germany). Hay (TecniLab-BMI BV, Someren, The Netherlands) was provided at least twice a week.
- Water (e.g. ad libitum): Free access to tap water.
- Acclimation period: at least 5 days before the start of treatment under laboratory conditions

ENVIRONMENTAL CONDITIONS
- Temperature (°C): 18 to 24°C
- Humidity (%): 40 to 70%
- Air changes (per hr): approximately 15 room air changes/hour
- Photoperiod (hrs dark / hrs light): 12-hour light/12-hour dark cycle

OTHER:
Identification of the animals was guaranteed via ear tattoos.
A health inspection was performed prior to treatment, to ensure that the animals are in a good state of health. Special attention was paid to the skin to be treated, which was intact and free from any abnormality.
Route:
epicutaneous, semiocclusive
Vehicle:
other: Kaydol White Mineral Oil
Concentration / amount:
undiluted (pure substance) / 50 % dilution in Kaydol White Mineral Oil
Route:
epicutaneous, occlusive
Vehicle:
other: Kaydol White Mineral Oil
Concentration / amount:
undiluted (pure substance) / 50 % dilution in Kaydol White Mineral Oil
No. of animals per dose:
Experimental group : 20 females.
Control group : 10 females.
Details on study design:
RANGE FINDING TESTS:
Preliminary irritation study
A preliminary irritation study was conducted in order to select test substance concentrations to be used in the Main Study. The selection of concentrations was based on the following criteria:
- The concentrations are well-tolerated by the animals.
- For the induction exposures: the highest possible concentration that produced mild irritation (grade 2).
- For challenge exposure: the maximum non-irritant concentration.

A series of test substance concentrations was tested. Practical feasibility of administration determined the highest starting-concentration (100 %). The test system, procedures and techniques were identical to those used during the main study, unless otherwise specified. The animals selected were between 4 and 9 weeks old. No body weights were determined.

Epidermal application:
A series of four test substance concentrations was used, the highest concentration being the maximum concentration that could technically be applied. Two different concentrations were applied (0.5 mL each) per animal to the clipped flank using Metalline patches# (2x3 cm) mounted on Medical tape, which will be held in place with Micropore tape# and subsequently Coban elastic bandage.
After 6 hours, the dressings were removed and the skin cleaned of residual test substance with Kaydol White Mineral Oil. The resulting dermal reactions were assessed for irritation 24 and 48 hours after removal of the last dressing.

Based on the results, two additional animals were treated each with two test substance concentrations of 50 and 75% to verify the correct challenge concentration.

MAIN STUDY
A. INDUCTION EXPOSURE
- No. of exposures: 3 (Days 1, 8 and 15)
- Exposure period: 6 hours (After 6 hours, the dressings were removed and skin cleaned of residual test substance using Kaydol White Mineral Oil. )
- Test groups: 0.5 mL of the undiluted test substance
- Control group: The control animals were treated as described for the experimental animals, except that, instead of the test substance, vehicle alone was administered.
- Site: The left side of the scapular region was clipped and subsequently epidermally treated with the test substance, all was held in place with Metalline patches (2x3 cm) mounted on Medical tape, which was held in place with Micropore tape and subsequently Coban elastic bandage.
- Frequency of applications: in total: trice
- Duration: 6 hours / 15 days
- Concentrations: 0.5 mL of the undiluted test substance
Evaluation: Immediately after removal of the last induction application on Day 15, the treated skin area was assessed for irritation.

B. CHALLENGE EXPOSURE
- No. of exposures: twice
- Day(s) of challenge: Day 29 and Day 36
- Exposure period: 6 hours (After 6 hours, the dressings were removed and skin cleaned of residual test substance and vehicle using Kaydol White Mineral Oil.)
- Site: The right flank of all animals was clipped and subsequently treated epidermally with the undiluted test substance and the vehicle (0.1 mL of each), using Patch Test Plasters (Curatest®, Lohmann, Almere, The Netherlands). The patches were held in place with Micropore tape and subsequently Coban elastic bandage.
- Evaluation (hr after challenge): The treated sites were assessed for challenge reactions 24 and 48 hours after removal of the dressings.

After termination, animals were sacrificed using isoflurane (Abbott B.V., Hoofddorp, The Netherlands) and an intra-peritoneal injection of Euthasol® 20% (AST Farma BV, Oudewater, The Netherlands).
Positive control substance(s):
yes
Remarks:
alpha-Hexylcinnamaldehyde
Key result
Reading:
other: first challenge
Group:
test group
Dose level:
50 % test substance concentration
No. with + reactions:
1
Total no. in group:
20
Clinical observations:
First challenge: Skin reactions of grade 1 were observed in one experimental animal and scaliness was observed in two experimental animals in response to the 50% test substance concentration. No skin reactions were evident in the control animals.
Remarks on result:
other: see Remark
Remarks:
Reading: other: first challenge. Group: test group. Dose level: 50 % test substance concentration. No with. + reactions: 1.0. Total no. in groups: 20.0. Clinical observations: First challenge: Skin reactions of grade 1 were observed in one experimental animal and scaliness was observed in two experimental animals in response to the 50% test substance concentration. No skin reactions were evident in the control animals..
Key result
Reading:
other: second challenge
Group:
test group
Dose level:
50 % test substance concentration
No. with + reactions:
0
Total no. in group:
20
Clinical observations:
To confirm the results of the first challenge, a second challenge was performed one week later. No skin reactions were evident after the second challenge exposure in the experimental and control animals.
Remarks on result:
other: see Remark
Remarks:
Reading: other: second challenge. Group: test group. Dose level: 50 % test substance concentration. No with. + reactions: 0.0. Total no. in groups: 20.0. Clinical observations: To confirm the results of the first challenge, a second challenge was performed one week later. No skin reactions were evident after the second challenge exposure in the experimental and control animals. .

PRELIMINARY IRRITATION STUDY

 

The results of the epidermal exposures for the selection of suitable test substance concentrations for the main study are described in Table 1. Based on the results, the undiluted test substance was selected for the three epidermal induction exposures. A 50 % test substance concentration was selected for the challenge phase.

 

MAINSTUDY

 

The skin effects observed immediately after the last (3rd) induction exposure are given in Table 2.

 

Challenge phase:

First challenge: Skin reactions of grade 1 were observed in one experimental animal and scaliness was observed in two experimental animals in response to the 50 % test substance concentration (see Table 2). No skin reactions were evident in the control animals.

Second challenge:

To confirm the results of the first challenge, a second challenge was performed one week later. No skin reactions were evident after the second challenge exposure in the experimental and control animals (see Table 3).

 

No mortality occurred and no symptoms of systemic toxicity were observed in the animals of the main study.

 

Body weights and body weight gain of experimental animals remained in the same range as controls over the study period (see Table 4).

 

Skin reactions were observed in response to a 50 % concentration of 71786-47-5 in three of the twenty experimental animals (one grade 1 skin reaction and two observations of scaliness) in the first challenge phase. These skin reactions were not confirmed at second challenge with the same concentration. No skin reactions were observed in the control animals in response to the 50 % first and second challenge exposure.

 

The six-month reliability check with Alpha-hexylcinnamicaldehyde indicates that the Buehler Test as performed at WIL Research Europe is an appropriate model for testing for contact hypersensitivity.


Table 1: preliminary irritation study   
SKIN REACTIONS AFTER EPIDERMAL EXPOSURE 
Animal Conc. % 24 hours after exposure 48 hours after exposure
number Erythema Oedema Erythema Oedema
(grade) (grade) (grade) (grade)
1 100 0 1 1 0
50 0 0 0 0
2 100 0 1 0 0
50 0 0 0 0
3 20 0 0 0 0
10 0 0 0 0
4 20 0 0 0 0
10 0 0 0 0
1 75 0 0 1 0
50 0 0 0 0
2 75 0 0 0 0
50 0 0 0 0
Note: It was noted that the identification of the animals used in the preliminary study was not unique. The raw data of the study specifies that the animals were of separate deliveries from the supplier and treated on different occasions.

Table 2: INDUCTION AND FIRST CHALLENGE READINGS
  Induction First Challenge
Animal Day 15 Day 30 Day 31
24 Hour Reading 48 Hour Reading
No Readings Readings Readings
100% # 50% vehicle 50% vehicle
  Er Oe        
      Control         
91 0 0 0 0 0 0
92 0 0 0 0 0 0
93 0 0 0 0 0 0
94 0 0 0 0 0 0
95 0 0 0 0 0 0
96 0 0 0 0 0 0
97 0 0 0 0 0 0
98 0 0 0 0 0 0
99 0p 0 0 0 0 0
100 0 0 0 0 0 0
      Experimental       
101 1 0 0 0 0 0
102 1p 0 0 0 0 0
103 1 0 0 0 0 0
104 1 0 0 0 0p 0
105 2 0 0 0 0 0
106 1 0 0 0 0 0
107 2p 0 0 0 1 0
108 0 0 0 0 0 0
109 1 0 0 0 0 0
110 1 0 0 0 0 0
111 1p 0 0 0 0 0
112 1 0 0 0 0 0
113 2 0 0 0 0 0
114 1p 0 0 0 0 0
115 1 0 0 0 0 0
116 1 0 0 0 0 0
117 1 0 0 0 0 0
118 1 0 0 0 0 0
119 2p 0 0 0 0 0
120 1 0 0 0 0p 0p
#. Test substance concentration (experimental animals) or vehicle (control animals).
p. Scaliness
Er: Erythema
Oe: Oedema
Vehicle: Kaydol White Mineral Oil.

Table 3: SECOND CHALLENGE READINGS
  Challenge
Animal Day 37 Day 38
No Readings Readings
50% vehicle 50% vehicle
Control       
91 0 0 0 0
92 0 0 0 0
93 0 0 0 0
94 0 0 0 0
95 0 0 0 0
96 0 0 0 0
97 0 0 0 0
98 0 0 0 0
99 0 0 0 0
100 0 0 0 0
Experimental
101 0 0 0 0
102 0 0 0 0
103 0 0 0 0
104 0 0 0 0
105 0 0 0 0
106 0 0 0 0
107 0 0 0 0
108 0 0 0 0
109 0 0 0 0
110 0 0 0 0
111 0 0 0 0
112 0 0 0 0
113 0 0 0 0
114 0 0 0 0
115 0 0 0 0
116 0 0 0 0
117 0 0 0 0
118 0 0 0 0
119 0 0 0 0
120 0 0 0 0
#. Test substance concentration (experimental animals) or vehicle (control animals).
Er: Erythema.
Oe: Oedema.
Vehicle: Kaydol White Mineral Oil.

 Table 4: BODY WEIGHTS (GRAM)
SEX/DOSE LEVEL ANIMAL DAY 1 DAY 38
FEMALES CONTROL 
    91 309 449
92 302 448
93 310 525
94 342 557
95 323 475
96 345 526
97 336 509
98 350 524
99 319 467
100 346 509
    MEAN   328 499
ST.DEV.   18 37
N   10 10
FEMALES EXPERIMENTAL
    101 337 493
102 352 561
103 334 510
104 359 534
105 306 493
106 340 552
107 321 478
108 333 550
109 323 456
110 348 558
111 280 519
112 353 507
113 333 510
114 306 459
115 310 497
116 339 572
117 334 525
118 332 485
119 326 543
120 332 544
   
    MEAN   330 517
ST.DEV.   19 34
N   20 20
Interpretation of results:
not sensitising
Remarks:
Migrated information Criteria used for interpretation of results: other: EU-GHS
Conclusions:
The study was performed according to the OECD Guideline 406 without deviations and according to the good laboratory practice principles, it is considered to be of high quality (reliability Klimisch 2). The criteria of validity of the test system are fulfilled. The test material did not induce a reproducible sensitisation on the intact skin of guinea pigs. The test material was considered not to be sensitising under the conditions of the test.
Executive summary:

Benzenesulfonic Acid mono and dialkylderivs magnesium salt, neutral, in diluents oil TBN = 20 (CAS 71768 -47 -5) was investigated for its sensitizing potential after repeated topical exposures in guinea pigs in a skin sensitization test according to the Buehler method (van Huygevoort, 2012, according to OECD 406, EU Method B6 and EPA OPPTS 870.2600, Klimisch 2). The Buehler type of sensitization test is selected at the request of the sponsor, since this substance is a surfactant. Studies have shown that the LLNA, which is typically used, over predicts sensitization potential of these surfactants. Test substance concentrations selected for the main study were based on the results of a preliminary study. In the main study, twenty experimental animals were epidermally treated on three occasions (once weekly, on Days 1, 8 and 15) with the undiluted test substance (100 % concentration) and ten control animals were similarly treated, but with vehicle alone (Kaydol White Mineral Oil). Two weeks after the last induction exposure (rest period), all animals were topically challenged with a 50% test substance concentration and the vehicle. A second challenge was performed approximately one week later with the same test substance concentrations and the vehicle.

Skin reactions were observed in response to a 50% concentration of the magnesium sulfonate read across substance (CAS 71786-47-5) in three of the twenty experimental animals (one grade 1 skin reaction and two observations of scaliness) in the first challenge phase.  These skin reactions were not confirmed at second challenge (one week later) with the same concentration.

So the number of animals exhibiting positive sensitization scores of 1 for the test substance at the 50% challenge and 50% rechallenge were 1/20 and 0/20, respectively (mean 24/48 hour Draize severity scores 0.00/0.05 and 0.0/0.0). No positive scores were observed in the vehicle control in the first and second challenge (mean 24/48 hour Draize severity scores 0.0/0.0). The only other observation was scaliness in two animals with scores of 0 in the first challenge with the 50% test material concentration and/or vehicle at the 48 hour reading. 

The six-month reliability check with Alpha-hexylcinnamicaldehyde indicates that the Buehler Test as performed at WIL Research Europe is an appropriate model for testing for contact hypersensitivity. Based on these results the test substance is not considered to be a skin sensitizer.

Endpoint:
skin sensitisation: in vitro
Data waiving:
study scientifically not necessary / other information available
Justification for data waiving:
other:
Endpoint conclusion
Endpoint conclusion:
no adverse effect observed (not sensitising)

Respiratory sensitisation

Endpoint conclusion
Endpoint conclusion:
no study available

Justification for classification or non-classification

The magnesium sulfonate read across substance, (CAS 71786-47-5), did not show skin sensitising potential in a GLP-guideline test according to the Buehler method. Therefore, the magnesium sulfonate target substance does not meet the criteria for classification and labelling for skin sensitisation in accordance with European regulation (EC) No. 1272/2008.