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Skin sensitisation

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Administrative data

Endpoint:
skin sensitisation: in vivo (non-LLNA)
Type of information:
other: Experimental result on formulation of Choline chloride with structural analogue
Adequacy of study:
key study
Reliability:
2 (reliable with restrictions)
Rationale for reliability incl. deficiencies:
other: Well-documented Guideline study according GLP, but performed on formulation of Choline Chloride with structural analogue (chlorocholine chloride).
Cross-reference
Reason / purpose for cross-reference:
reference to other study
Reference
Endpoint:
skin sensitisation: in vivo (non-LLNA)
Type of information:
migrated information: read-across from supporting substance (structural analogue or surrogate)
Adequacy of study:
supporting study
Study period:
1991-10 to 1991-12
Reliability:
2 (reliable with restrictions)
Rationale for reliability incl. deficiencies:
other: Well-documented guideline study according to GLP, but conducted on Read-Across substance / component of Choline chloride formulation.
Reason / purpose for cross-reference:
reference to other study
Qualifier:
according to guideline
Guideline:
OECD Guideline 406 (Skin Sensitisation)
Version / remarks:
version of 1987-02-24
Deviations:
no
GLP compliance:
yes
Remarks:
Dates of inspection: 1991-10-30, 1991-11-13, 1991-11-27, 1991-12-10
Type of study:
Buehler test
Species:
guinea pig
Strain:
other: Albino
Sex:
male/female
Details on test animals and environmental conditions:
TEST ANIMALS
- Source: Bred at the Toxicology Department, Rallis Agrochemical Research Station, Bangalore 560 058, India and procured from IVRI, Bangalore
- Age at study initiation: approx. 7 - 9 weeks
- Weight at study initiation: 407 ± 40 g
- Housing: Individually in standard rat cages (430 mm x 270 mm x 150 mm) equipped with hoppers for pelletted feed and drinking water in glass bottles.
- Diet (e.g. ad libitum): Standard 'Gold Mohur' brand pelletted guinea pig maintenance diet manufactured by M/S Lipton India Ltd., Bangalore-52, INDIA (a subsidiary of UNILEVER of ENGLAND) ad libitum. Vitamin C was supplemented thrice a week in drinking water (1 g/L).
- Water (e.g. ad libitum): Deep borewell water passed through activated charcoal filter and exposed to UV rays (Aquaguard on-line water filter cum-purifie manufactured by M/s Eureka Forbes Ltd., Bombay 400 001, INDIA in collaboration with ELECTROLUX of SWEDEN) supplied ad libitum in glass bottles with stainless steel sipper tubes.
- Acclimation period: One week in individual cages under laboratory conditions after first Veterinary examination

ENVIRONMENTAL CONDITIONS
- Temperature (°C): 22 ± 3°C
- Humidity (%): 40 - 70 %
- Air changes (per hr): 10 - 15
- Photoperiod (hrs dark / hrs light): 12 hours artificial fluorescent light and 12 hours dark.
Route:
epicutaneous, occlusive
Vehicle:
unchanged (no vehicle)
Concentration / amount:
100% test item, i.e. 720 g/L Chlormequat chloride
Route:
epicutaneous, semiocclusive
Vehicle:
unchanged (no vehicle)
Concentration / amount:
100% test item, i.e. 720 g/L Chlormequat chloride
No. of animals per dose:
Control (saline, 0.5 mL): 10
Positive control (0.4 mL): 5
Test compound (0.5 mL): 10
Details on study design:
RANGE FINDING TESTS: Dose selection was based on primary skin irritation study in rabbits

MAIN STUDY
A. INDUCTION EXPOSURE
- No. of exposures: 3
- Exposure period: -
- Test groups: Test substance
- Control group: saline
- Site: left dorso-lateral thorax
- Frequency of applications: one application a week for 3 weeks
- Duration: 6 h
- Concentrations: undiluted test item

B. CHALLENGE EXPOSURE
- No. of exposures: 2
- Day(s) of challenge: 2 and 3 weeks after last induction
- Exposure period: -
- Test groups: Test substance
- Control group: Saline
- Site: right dorso-lateral thorax (challenge), virgin site on right flank (rechallenge)
- Concentrations: undiluted test item
- Evaluation (hr after challenge): 24, 48 and 72 h

OTHER:
Challenge controls:
none
Positive control substance(s):
yes
Remarks:
1-Chloro-2,4-di-Nitrobenzene (DNCB), (Sigma, USA) Lot No.100 HO 738. A volume of 10 mL of 0.15% w/v solution of DNCB in methanol was prepared.
Positive control results:
Local Skin Reactions / positive control:
See also Table 2 for Individual Animal Data and Table 3 Summary Data. The testing of the test item including control and positive control DNBC for skin sensitizing properties revealed the following results:
A: Control Group: Guinea pigs in this group did not show local erythema or oedema during three induction challenge and rechallenge periods
B. Positive (DNCB) Control Group: First three induction periods did not cause any local reaction except for one guinea pig showing slight erythema during first hour after second induction application.
During challenge period all the animals showed slight to moderate erythema during 24 hours observation which got reduced in severity at 48 hours and was mild at 72 hours observation.
C. Treatment Group: No local skin reaction was observed during the three induction applications. However, during 72 hours observation after first challenge application two animals showed slight erythema. On rechallenge none of the animals showed any skin reaction attributable to test compound.
As a consequence, it can be concluded that the head of the laboratory has the capability to identify positive dermal sensitizers, the positive control DNBC induced the appropriate skin reactions and the results are valid.
Reading:
1st reading
Hours after challenge:
24
Group:
test chemical
Dose level:
0.5 mL undiluted test item
No. with + reactions:
0
Total no. in group:
8
Clinical observations:
Normal (8 animals), Lethargy, excess salivation (1 animal, died after induction phase), Excess salivation (1 animal, died after induction phase)
Remarks on result:
other: see Remark
Remarks:
Reading: 1st reading. . Hours after challenge: 24.0. Group: test group. Dose level: 0.5 mL undiluted test item. No with. + reactions: 0.0. Total no. in groups: 8.0. Clinical observations: Normal (8 animals), Lethargy, excess salivation (1 animal, died after induction phase), Excess salivation (1 animal, died after induction phase) .
Reading:
1st reading
Hours after challenge:
48
Group:
test chemical
Dose level:
0.5 mL undiluted test item
No. with + reactions:
0
Total no. in group:
8
Clinical observations:
Normal (8 animals), Lethargy, excess salivation (1 animal, died after induction phase), Excess salivation (1 animal, died after induction phase)
Remarks on result:
other: see Remark
Remarks:
Reading: 1st reading. . Hours after challenge: 48.0. Group: test group. Dose level: 0.5 mL undiluted test item. No with. + reactions: 0.0. Total no. in groups: 8.0. Clinical observations: Normal (8 animals), Lethargy, excess salivation (1 animal, died after induction phase), Excess salivation (1 animal, died after induction phase) .
Reading:
1st reading
Hours after challenge:
72
Group:
test chemical
Dose level:
0.5 mL undiluted test item
No. with + reactions:
2
Total no. in group:
8
Clinical observations:
Normal (8 animals), Lethargy, excess salivation (1 animal, died after induction phase), Excess salivation (1 animal, died after induction phase)
Remarks on result:
other: see Remark
Remarks:
Reading: 1st reading. . Hours after challenge: 72.0. Group: test group. Dose level: 0.5 mL undiluted test item. No with. + reactions: 2.0. Total no. in groups: 8.0. Clinical observations: Normal (8 animals), Lethargy, excess salivation (1 animal, died after induction phase), Excess salivation (1 animal, died after induction phase) .
Reading:
rechallenge
Hours after challenge:
24
Group:
test chemical
Dose level:
0.5 mL undiluted test item
No. with + reactions:
0
Total no. in group:
8
Clinical observations:
Normal (8 animals), Lethargy, excess salivation (1 animal, died after induction phase), Excess salivation (1 animal, died after induction phase)
Remarks on result:
other: see Remark
Remarks:
Reading: rechallenge. . Hours after challenge: 24.0. Group: test group. Dose level: 0.5 mL undiluted test item. No with. + reactions: 0.0. Total no. in groups: 8.0. Clinical observations: Normal (8 animals), Lethargy, excess salivation (1 animal, died after induction phase), Excess salivation (1 animal, died after induction phase) .
Reading:
rechallenge
Hours after challenge:
48
Group:
test chemical
Dose level:
0.5 mL undiluted test item
No. with + reactions:
0
Total no. in group:
8
Clinical observations:
Normal (8 animals), Lethargy, excess salivation (1 animal, died after induction phase), Excess salivation (1 animal, died after induction phase)
Remarks on result:
other: see Remark
Remarks:
Reading: rechallenge. . Hours after challenge: 48.0. Group: test group. Dose level: 0.5 mL undiluted test item. No with. + reactions: 0.0. Total no. in groups: 8.0. Clinical observations: Normal (8 animals), Lethargy, excess salivation (1 animal, died after induction phase), Excess salivation (1 animal, died after induction phase) .
Reading:
rechallenge
Hours after challenge:
72
Group:
test chemical
Dose level:
0.5 mL undiluted test item
No. with + reactions:
0
Total no. in group:
8
Clinical observations:
Normal (8 animals), Lethargy, excess salivation (1 animal, died after induction phase), Excess salivation (1 animal, died after induction phase)
Remarks on result:
other: see Remark
Remarks:
Reading: rechallenge. . Hours after challenge: 72.0. Group: test group. Dose level: 0.5 mL undiluted test item. No with. + reactions: 0.0. Total no. in groups: 8.0. Clinical observations: Normal (8 animals), Lethargy, excess salivation (1 animal, died after induction phase), Excess salivation (1 animal, died after induction phase) .
Reading:
1st reading
Hours after challenge:
24
Group:
negative control
Dose level:
0.5 mL saline
No. with + reactions:
0
Total no. in group:
9
Clinical observations:
Normal (9 animals), excess salivation (1 animal, died after induction phase)
Remarks on result:
other: see Remark
Remarks:
Reading: 1st reading. . Hours after challenge: 24.0. Group: negative control. Dose level: 0.5 mL saline. No with. + reactions: 0.0. Total no. in groups: 9.0. Clinical observations: Normal (9 animals), excess salivation (1 animal, died after induction phase).
Reading:
1st reading
Hours after challenge:
48
Group:
negative control
Dose level:
0.5 mL saline
No. with + reactions:
0
Total no. in group:
9
Clinical observations:
Normal (9 animals), excess salivation (1 animal, died after induction phase)
Remarks on result:
other: see Remark
Remarks:
Reading: 1st reading. . Hours after challenge: 48.0. Group: negative control. Dose level: 0.5 mL saline. No with. + reactions: 0.0. Total no. in groups: 9.0. Clinical observations: Normal (9 animals), excess salivation (1 animal, died after induction phase).
Reading:
1st reading
Hours after challenge:
72
Group:
negative control
Dose level:
0.5 mL saline
No. with + reactions:
0
Total no. in group:
9
Clinical observations:
Normal (9 animals), excess salivation (1 animal, died after induction phase)
Remarks on result:
other: see Remark
Remarks:
Reading: 1st reading. . Hours after challenge: 72.0. Group: negative control. Dose level: 0.5 mL saline. No with. + reactions: 0.0. Total no. in groups: 9.0. Clinical observations: Normal (9 animals), excess salivation (1 animal, died after induction phase).
Reading:
rechallenge
Hours after challenge:
24
Group:
negative control
Dose level:
0.5 mL saline
No. with + reactions:
0
Total no. in group:
9
Clinical observations:
Normal (9 animals), excess salivation (1 animal, died after induction phase)
Remarks on result:
other: see Remark
Remarks:
Reading: rechallenge. . Hours after challenge: 24.0. Group: negative control. Dose level: 0.5 mL saline. No with. + reactions: 0.0. Total no. in groups: 9.0. Clinical observations: Normal (9 animals), excess salivation (1 animal, died after induction phase).
Reading:
rechallenge
Hours after challenge:
48
Group:
negative control
Dose level:
0.5 mL saline
No. with + reactions:
0
Total no. in group:
9
Clinical observations:
Normal (9 animals), excess salivation (1 animal, died after induction phase)
Remarks on result:
other: see Remark
Remarks:
Reading: rechallenge. . Hours after challenge: 48.0. Group: negative control. Dose level: 0.5 mL saline. No with. + reactions: 0.0. Total no. in groups: 9.0. Clinical observations: Normal (9 animals), excess salivation (1 animal, died after induction phase).
Reading:
rechallenge
Hours after challenge:
72
Group:
negative control
Dose level:
0.5 mL saline
No. with + reactions:
0
Total no. in group:
9
Clinical observations:
Normal (9 animals), excess salivation (1 animal, died after induction phase)
Remarks on result:
other: see Remark
Remarks:
Reading: rechallenge. . Hours after challenge: 72.0. Group: negative control. Dose level: 0.5 mL saline. No with. + reactions: 0.0. Total no. in groups: 9.0. Clinical observations: Normal (9 animals), excess salivation (1 animal, died after induction phase).
Reading:
1st reading
Hours after challenge:
24
Group:
positive control
Dose level:
0.4 mL 0.15% w/v solution of DNBC
No. with + reactions:
5
Total no. in group:
5
Clinical observations:
Normal (2 animals), scale formation (3 animals)
Remarks on result:
other: Reading: 1st reading. . Hours after challenge: 24.0. Group: positive control. Dose level: 0.4 mL 0.15% w/v solution of DNBC. No with. + reactions: 5.0. Total no. in groups: 5.0. Clinical observations: Normal (2 animals), scale formation (3 animals).
Reading:
1st reading
Hours after challenge:
48
Group:
positive control
Dose level:
0.4 mL 0.15% w/v solution of DNBC
No. with + reactions:
5
Total no. in group:
5
Clinical observations:
Normal (2 animals), scale formation (3 animals)
Remarks on result:
other: Reading: 1st reading. . Hours after challenge: 48.0. Group: positive control. Dose level: 0.4 mL 0.15% w/v solution of DNBC. No with. + reactions: 5.0. Total no. in groups: 5.0. Clinical observations: Normal (2 animals), scale formation (3 animals).
Reading:
1st reading
Hours after challenge:
72
Group:
positive control
Dose level:
0.4 mL 0.15% w/v solution of DNBC
No. with + reactions:
5
Total no. in group:
5
Clinical observations:
Normal (2 animals), scale formation (3 animals)
Remarks on result:
other: Reading: 1st reading. . Hours after challenge: 72.0. Group: positive control. Dose level: 0.4 mL 0.15% w/v solution of DNBC. No with. + reactions: 5.0. Total no. in groups: 5.0. Clinical observations: Normal (2 animals), scale formation (3 animals).

Due to limitations of the numbers of lines in this free text field, detailed background information was attached, see file 'Suresh_SkinSens_Tables'

Interpretation of results:
GHS criteria not met
Conclusions:
The present study is a well-documented guideline study according to GLP which would normally be classified as Klimisch 1, but since it was conducted on the Read-Across substance Chlorocholine chloride, which is also a component of the Choline chloride formulation tested in endpoint study record "Skin sensitisation - Driscoll R, 1998, guinea pig, CC-CMC", it could be only classified as Klimisch 2.
However, the latter is only relevant when the results on chlormequat are used for read-across to Choline chloride, which are still reliable enough to conclude that Choline chloride is not a skin sensitiser taking into account the negative outcome of the present study. Since the chlorine moiety could lead to an altered metabolism and most likely to more reactive metabolites forming allergens by altering the bodies proteins, the outcome of this present study could most likely overestimate the actual sensitising effects of Choline chloride. Since only two animals of eight showed a very slight erythema only at one single timepoint (72h of challenge phase) and even no effects were noted during the rechallenge phase, it can be safely concluded that Chlormequat and hence Choline chloride do not exhibit sensitising properties and do therefore not need to be classified as sensitisers.
The results are fully reliable when they are used to discuss the possible effects of Chlorocholine chloride on the outcome of the study conducted on a Choline chloride / Chlormequat formulation, see "Skin sensitisation - Driscoll R, 1998, guinea pig, CC-CMC" and the endpoint summary "7.4.1 Skin sensitisation".
Executive summary:

In a dermal sensitisation study according to OECD guideline 406 and conducted under GLP with Chlormequat chloride 720 g/L, 7-9 weeks old albino guinea pigs (21 male / 4 female) were tested using the Bühler assay. Here, 0.5 mL of the test item were applied three times during induction phase and each once during the challenge and rechallenge phase. 1-Chloro-2,4-di-Nitrobenzene (DNCB) served as positive control and induced positive results.

Most animals exhibited no clinical signs except two animals of the test group (Lethargy, excess salivation (1x) and excess salivation, lung consolidation, liver abscess (1x)), which died after the induction phase, and one animal of the control group (excess salivation), which also died after the induction phase. Three out of five animals of the positive control group showed scale formation.

Only two out of eight animals of the test group showed a very slight erythema after 72h during the challenge phase, no reactions were observed during the rechallenge phase.

In this study, Chlorocholine chloride is not a dermal sensitizer, and consequently also Choline chloride does not need to be classified as skin sensitizer.

Data source

Reference
Reference Type:
study report
Title:
Unnamed
Year:
1998
Report date:
1998

Materials and methods

Test guidelineopen allclose all
Qualifier:
according to guideline
Guideline:
OECD Guideline 406 (Skin Sensitisation)
Version / remarks:
adopted 1992-07-17
Deviations:
no
Qualifier:
according to guideline
Guideline:
EU Method B.6 (Skin Sensitisation)
Version / remarks:
Directive 96/54/EEC
Deviations:
no
GLP compliance:
yes
Remarks:
including compliance statement
Type of study:
guinea pig maximisation test
Justification for non-LLNA method:
The Buehler test (in vivo) was performed before the LLNA was set as preferred test method.

Test material

Constituent 1
Reference substance name:
Chlormequat chloride 460 g/L, Choline chloride 320 g/L
IUPAC Name:
Chlormequat chloride 460 g/L, Choline chloride 320 g/L
Test material form:
other: liquid
Details on test material:
- Name of test material (as cited in study report): Chlormequat chloride 460 g/L, Choline chloride 320 g/L
- Substance type: formulation of active and read-across substance
- Physical state: liquid
- Composition of test material, percentage of components: Chlormequat chloride 460 g/L, Choline chloride 320 g/L
- Storage condition of test material: room temperature in the dark
- Other: Description: extremely pale yellow liquid

In vivo test system

Test animals

Species:
guinea pig
Strain:
Dunkin-Hartley
Sex:
male
Details on test animals and environmental conditions:
TEST ANIMALS
- Source: David Hall Limited, Burton-on-Trent, Staffordshire, UK
- Age at study initiation: approx. 8 - 12 weeks
- Weight at study initiation: 312 - 393 g
- Housing: singly or in pairs in solid-floor polypropylene cages furnished with woodflakes
- Diet (e.g. ad libitum): Guinea Pig FD1 Diet, Special Diets Services Limited, Witham, Essex, UK; ad libitum
- Water (e.g. ad libitum): tap water ad libitum
- Acclimation period: at least five days

ENVIRONMENTAL CONDITIONS
- Temperature (°C): 19 - 22 °C
- Humidity (%): 48 - 56 %
- Air changes (per hr): approx. 15
- Photoperiod (hrs dark / hrs light): 12 / 12

Other: each animal was selected at random and given a number unique within the study which was written on a small area of clipped rump using a black indelible marker-pen.

Study design: in vivo (non-LLNA)

Inductionopen allclose all
Route:
intradermal and epicutaneous
Vehicle:
water
Concentration / amount:
5% w/v solution in distilled water (Intradermal induction)
undiluted as supplied (Topical induction)
undiluted as supplied and 75% v/v in water (Topical induction)
Challengeopen allclose all
Route:
epicutaneous, occlusive
Vehicle:
water
Concentration / amount:
5% w/v solution in distilled water (Intradermal induction)
undiluted as supplied (Topical induction)
undiluted as supplied and 75% v/v in water (Topical induction)
No. of animals per dose:
10 (test group)
5 (control group)
Details on study design:
RANGE FINDING TESTS:
- Single intradermal exposure with 0.1 mL of either 1 %, 5%, 10% or 25% w/v of the test item in distilled water, sighting test, for intradermal induction
- Two guinea pigs, intradermally injected with Freund's Complete Adjuvant fifteen days earlier, were topically (occlusive) exposed to 100%, 75%, 50% and 25% v/v of the test item in distilled water, for topical induction
- 100%, 75%, 50% and 25% v/v of the test item in distilled water, 14 d after induction with Freund´s Complete Adjuvant analogue to control animals in the main study, for topical challenge

MAIN STUDY
A. INDUCTION EXPOSURE
- No. of exposures: 1 intradermal (row of three injections, see additional information), 1 topical 7 days later
- Exposure period: 48 h for topical induction
- Test groups:
Intradermal Induction: row of three injections, (1) Freund´s Complete Adjuvans (FCA) and H2O dest. 1:1, (2) 5% w/v solution of the test material in distilled water, (3) 5% w/v solution of the test material in distilled water and FCA plus H2O dest. 1:1
Topical induction: undiluted test material
- Control group:
Intradermal Induction: row of three injections, (1) Freund´s Complete Adjuvans (FCA) and H2O dest. 1:1, (2) distilled water, (3) 50% distilled water and FCA plus H2O dest. 1:1
Topical induction: bare filter paper
- Site: shoulder
- Frequency of applications: Day 1 and 7
- Duration: Total 9 d
- Concentrations: 5% w/v in water (intradermal), undiluted (topical)

B. CHALLENGE EXPOSURE
- No. of exposures:
- Day(s) of challenge: Day 21
- Exposure period: 48h
- Test groups: undiluted as supplied and 75% v/v of test item in distilled water
- Control group: undiluted as supplied and 75% v/v of test item in distilled water
- Site: right (100% test item) and left (75% v/v) shorn flank
- Concentrations: undiluted and 75% v/v
- Evaluation (hr after challenge): 24h and 48 h
Challenge controls:
5 animals, undiluted test item and 75% v/v of test item in distilled water
Positive control substance(s):
yes
Remarks:
historical control data from Neomycin Sulphate, 2-Mercaptobenzothiazole, 2,4-Dinitrochlorobenzene

Results and discussion

Positive control results:
See attached background material "Driscoll_PositiveControlData" which demonstrates clearly that the laboratory has the capability to identify positive dermal sensitizers.
The incidences of sensitization of the test animals were 60% (Neomycin Sulphate), 70-90% (2-Mercaptobenzothiazole) and 100% (2,4-Dinitrochlorobenzene).

In vivo (non-LLNA)

Resultsopen allclose all
Reading:
1st reading
Hours after challenge:
24
Group:
test chemical
Dose level:
100% of the test item as supplied
No. with + reactions:
0
Total no. in group:
10
Clinical observations:
No skin reactions or differences in body weight gain compared to control were noted.
Remarks on result:
other: see Remark
Remarks:
Reading: 1st reading. . Hours after challenge: 24.0. Group: test group. Dose level: 100% of the test item as supplied. No with. + reactions: 0.0. Total no. in groups: 10.0. Clinical observations: No skin reactions or differences in body weight gain compared to control were noted..
Reading:
2nd reading
Hours after challenge:
48
Group:
test chemical
Dose level:
100% of the test item as supplied
No. with + reactions:
0
Total no. in group:
10
Clinical observations:
No skin reactions or differences in body weight gain compared to control were noted.
Remarks on result:
other: see Remark
Remarks:
Reading: 2nd reading. . Hours after challenge: 48.0. Group: test group. Dose level: 100% of the test item as supplied. No with. + reactions: 0.0. Total no. in groups: 10.0. Clinical observations: No skin reactions or differences in body weight gain compared to control were noted..
Reading:
1st reading
Hours after challenge:
24
Group:
test chemical
Dose level:
75% v/v of the test item in distilled water
No. with + reactions:
0
Total no. in group:
10
Clinical observations:
No skin reactions or differences in body weight gain compared to control were noted.
Remarks on result:
other: see Remark
Remarks:
Reading: 1st reading. . Hours after challenge: 24.0. Group: test group. Dose level: 75% v/v of the test item in distilled water. No with. + reactions: 0.0. Total no. in groups: 10.0. Clinical observations: No skin reactions or differences in body weight gain compared to control were noted..
Reading:
2nd reading
Hours after challenge:
48
Group:
test chemical
Dose level:
75% v/v of the test item in distilled water
No. with + reactions:
0
Total no. in group:
10
Clinical observations:
No skin reactions or differences in body weight gain compared to control were noted.
Remarks on result:
other: see Remark
Remarks:
Reading: 2nd reading. . Hours after challenge: 48.0. Group: test group. Dose level: 75% v/v of the test item in distilled water. No with. + reactions: 0.0. Total no. in groups: 10.0. Clinical observations: No skin reactions or differences in body weight gain compared to control were noted..
Reading:
1st reading
Hours after challenge:
24
Group:
other: challenge control
Dose level:
100% of the test item as supplied
No. with + reactions:
0
Total no. in group:
5
Clinical observations:
No skin reactions were noted.
Remarks on result:
other: Reading: 1st reading. . Hours after challenge: 24.0. Group: other: challenge control. Dose level: 100% of the test item as supplied. No with. + reactions: 0.0. Total no. in groups: 5.0. Clinical observations: No skin reactions were noted..
Reading:
2nd reading
Hours after challenge:
48
Group:
other: challenge control
Dose level:
100% of the test item as supplied
No. with + reactions:
0
Total no. in group:
5
Clinical observations:
No skin reactions were noted.
Remarks on result:
other: Reading: 2nd reading. . Hours after challenge: 48.0. Group: other: challenge control. Dose level: 100% of the test item as supplied. No with. + reactions: 0.0. Total no. in groups: 5.0. Clinical observations: No skin reactions were noted..
Reading:
1st reading
Hours after challenge:
24
Group:
other: challenge control
Dose level:
75% v/v of the test item in distilled water
No. with + reactions:
0
Total no. in group:
5
Clinical observations:
No skin reactions were noted.
Remarks on result:
other: Reading: 1st reading. . Hours after challenge: 24.0. Group: other: challenge control. Dose level: 75% v/v of the test item in distilled water. No with. + reactions: 0.0. Total no. in groups: 5.0. Clinical observations: No skin reactions were noted..
Reading:
2nd reading
Hours after challenge:
48
Group:
other: challenge control
Dose level:
75% v/v of the test item in distilled water
No. with + reactions:
0
Total no. in group:
5
Clinical observations:
No skin reactions were noted.
Remarks on result:
other: Reading: 2nd reading. . Hours after challenge: 48.0. Group: other: challenge control. Dose level: 75% v/v of the test item in distilled water. No with. + reactions: 0.0. Total no. in groups: 5.0. Clinical observations: No skin reactions were noted..

Any other information on results incl. tables

 

TABLE 1

Challenge concentrations: Undiluted as supplied and 75 % v/v

Vehicle: Distilled water

Animal Number

Skin Reactions (Hours After Removal of Dressing)

24 Hours

48 Hours

100%

75%

100%

75%

Er

Oe

Other

Er

Oe

Other

Er

Oe

Other

Er

Oe

Other

Individual skin reactions in test animals after challenge

1

0

0

-

0

0

-

0

0

-

0

0

-

2

0

0

-

0

0

-

0

0

-

0

0

-

3

0

0

-

0

0

-

0

0

-

0

0

-

4

0

0

-

0

0

-

0

0

-

0

0

-

5

0

0

-

0

0

-

0

0

-

0

0

-

6

0

0

-

0

0

-

0

0

-

0

0

-

7

0

0

-

0

0

-

0

0

-

0

0

-

8

0

0

-

0

0

-

0

0

-

0

0

-

9

0

0

-

0

0

-

0

0

-

0

0

-

10

0

0

-

0

0

-

0

0

-

0

0

-

Individual skin reactions in control animals after challenge

11

0

0

-

0

0

-

0

0

-

0

0

-

12

0

0

-

0

0

-

0

0

-

0

0

-

13

0

0

-

0

0

-

0

0

-

0

0

-

14

0

0

-

0

0

-

0

0

-

0

0

-

15

0

0

-

0

0

-

0

0

-

0

0

-

Applicant's summary and conclusion

Interpretation of results:
not sensitising
Conclusions:
The present study is a well-documented GLP study according OECD guideline 406 (Guinea pig maximisation test) with the restriction, that it was conducted not only on Choline chloride (CC) but on a formulation with CC (320 g/L) and chlorocholine chloride (CCC, 460 g/L), whereas the latter was shown not to be a sensitizer in the study by Suresh. So in general, the study was classified as Klimisch 2 and hence, the results can be safely used to assess the sensitising potential of Choline chloride. The test item contains two relevant parts Choline chloride, i.e. 41 % of the solid ingredients, and the solvent (water) is not a known sensitizer. So since neither water nor CCC, which may also serve as a read-across substance, was shown to be a dermal sensitizer, it can be concluded that every possible positive result obtained from the formulation must be attributed to Choline chloride. Since no animal of the test group at no timepoint of observation showed any skin reaction during the challenge phase, and the dose of the test item was properly determined by preliminary sighting tests as the demanded highest non-irritating dose, it can safely be concluded that the test item did not exhibit any sensitizing properties and consequently, that Choline chloride is non-sensitizing, too. Hence, Choline chloride does not need to be classified as skin-sensitizer, neither according Regulation 1272/2008/EC nor Directive 67/548/EEC.
Executive summary:

In a dermal sensitization study according GLP and OECD guideline 406 with a formulation of Choline chloride (CC) and Chlorocholine chloride (CCC) (320 g/L CC, 460 g/L CCC) in water, 8-12 weeks old male albino Dunkin Hartley guinea pigs (10 animals/ test were tested in the Guinea pig maximization test (GPMT according to Magnus and Kligmann)). Induction was performed once intradermal (Day 1) and once topical (Day 7, occlusive), topical challenge was on Day 21 (occlusive). Historical data from Neomycin Sulphate, 2-Mercaptobenzothiazole and 2,4-Dinitrochlorobenzene served as positive controls and induced the anticipated effect.

No skin reactions were observed, neither in the test nor control group, and body weight gain of the test group was similar to control. Since no sensitizing effects were noted in none of the test animals in this study, the formulation of CC and CCC, and consequently also pure Choline chloride, is not a dermal sensitizer, and does therefore not need to be classified as sensitising to the skin, neither according Regulation 1272/2008/EC nor Directive 67/548/EEC.