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Administrative data

Description of key information

Key study: A Mouse local lymph node assay (LLNA) was performed according to OECD 429 (GLP study). Four CBA mice were exposed up 10, 25 and 50% distilled CNSL (two samples) in acetone/olive oil 4:1. Stimulation indexes of 29.29 and 30.90 were identified. The EC3 value was not directly determined, however, 10% of test substance elicited a stimulation index of 30 (EC3 = % test substance required to elicit a stimulation index of 3). Based on these results, the substance was determined to be a skin sensitizer category 1A.

Supporting study: A Guinea Pig Maximisation Test was performed according to OECD 406 (GLP study). Twenty guinea pigs were exposed to 1% of test item in paraffin oil at induction, and 5% and 2% at challenge. A positive response was elicited in 70% of animals when administered 1% induction and challenged with a 5% concentration. Since greater than 60% of animals responded at a 1% intradermal induction dose the substance was determined to be skin sensitizer category 1A.

Justification for selection of skin sensitisation endpoint:

The LLNA assay was selected as key study.

Key value for chemical safety assessment

Skin sensitisation

Link to relevant study records

Referenceopen allclose all

Endpoint:
skin sensitisation: in vivo (LLNA)
Type of information:
experimental study
Adequacy of study:
key study
Reliability:
1 (reliable without restriction)
Qualifier:
according to
Guideline:
OECD Guideline 429 (Skin Sensitisation: Local Lymph Node Assay)
GLP compliance:
yes (incl. certificate)
Type of study:
mouse local lymph node assay (LLNA)
Species:
mouse
Strain:
CBA
Sex:
female
Details on test animals and environmental conditions:
TEST ANIMALS
- Source: Harlan UK Limited
- Age at study initiation: 8 - 12 weeks old
- Weight at study initiation: 15 to 23 g
- Housing: suspended solid-floor polypropylene cages
- Diet (e.g. ad libitum): Certified rat and mous diet ad libitum
- Water (e.g. ad libitum): ad libitum
- Acclimation period: 5 days

ENVIRONMENTAL CONDITIONS
- Temperature (°C): 19 - 25°C
- Humidity (%): 30 to 70%
- Air changes (per hr): 15/hour
Vehicle:
acetone/olive oil (4:1 v/v)
Concentration:
10%, 25% and 50% for screening test.
No. of animals per dose:
4
Details on study design:
RANGE FINDING TESTS:
- Compound solubility: Soluble
- Irritation: Mild or moderate redness to the ears and neck was noted in animals tested with the undiluted test material, or at 50 or 25%.

MAIN STUDY
ANIMAL ASSIGNMENT AND TREATMENT
- Name of test method: OECD 429
- Criteria used to consider a positive response: Stimulation Index of 3.0 or greater

TREATMENT PREPARATION AND ADMINISTRATION: NX-4708 and NX-2023 were used undiluted and freshly prepared as a solution in acetone:olive oil 4:1. This vehicle was chosen as it produced the highest concentration that was suitable for dosing. Two groups of four mice were treated with either NX-4708 or NX-2023 at a concentration of 10% v/v in acetone/olive oil 4:1. The mice were administered a daily application of 25 μl of each test material to the dorsal surface of each ear for three consecutive days. The test material formulations were administered using an automatic micropipette and spread over the dorsal surface of the ear using the tip of the pipette.
Positive control substance(s):
hexyl cinnamic aldehyde (CAS No 101-86-0)
Statistics:
Use of statistics not indicated.
Positive control results:
Concentration: 15%
Stimulation index: 10.91
Result: positive
Key result
Parameter:
SI
Value:
29.29
Remarks on result:
other: Vehicle: N/A NX-4708: 29.29
Key result
Parameter:
SI
Value:
30.9
Remarks on result:
other:
Remarks:
Vehicle: NX-2023: 30.90
Key result
Parameter:
other: disintegrations per minute (DPM)
Value:
142 691.4
Remarks on result:
other: Vehicle: 4871.26 NX-4708: 142691.40
Key result
Parameter:
other: disintegrations per minute (DPM)
Value:
150 509.5
Remarks on result:
other:
Remarks:
Vehicle: NX-2023: 150509.50

Table 1 Clinical observations, bodyweight and mortality data - preliminary screening test - NX-4708

Concentration (% v/v) in acetone/olive oil 4:1

Animal number

Bodyweight change (g) Day

 

1

6

1

2

3

 

4

5

6

 

Pre

dose

Post dose

Pre

 dose

Post dose

Pre

dose

Post dose

 

100

S-1

20

-

0

0

0

-

-

-

-

-

-

50

S-2

20

-

0

0

0

0

-

-

-

-

-

25

S-3

17

-

0

0

0

0

-

-

-

-

-

10

S-4

19

20

0

0

0

0

0

0

0

0

0

Interpretation of results:
sensitising
Remarks:
Migrated information (EU)
Conclusions:
A positive result was obtained in a well-conducted LLNA, indicating that CNSL is able to cause skin sensitisation.
Executive summary:

In a dermal sensitization study with distilled CNSL in acetone/olive oil 4:1, four young adult CBA mice were tested using the LLNA method. Mild redness to the eyes and neck were observed in test animals. No other clinical signs of toxicity were noted and mortality did not occur. Body weights of treated animals were comparable to that of controls.

Stimulation indexes of 29.29 and 30.90 were identified, for NX-4708 and NX-2023, respectively. The EC3 value was not directly determined, however, 10% of test substance elicited a stimulation index of 30 (EC3 = % test substance required to elicit a stimulation index of 3). This indicates that CNSL is a skin sensitiser category 1A.

Endpoint:
skin sensitisation: in vitro
Data waiving:
study scientifically not necessary / other information available
Justification for data waiving:
an in vitro skin sensitisation study does not need to be conducted because adequate data from an in vivo skin sensitisation study are available
Justification for type of information:
An in vitro study does not need to be conducted since adequate data from an in vivo skin sensitisation study are available.
Endpoint:
skin sensitisation: in vivo (non-LLNA)
Type of information:
experimental study
Adequacy of study:
supporting study
Reliability:
1 (reliable without restriction)
Rationale for reliability incl. deficiencies:
other: Study conducted to OECD guidelines and to GLP.
Qualifier:
according to
Guideline:
OECD Guideline 406 (Skin Sensitisation)
GLP compliance:
yes (incl. certificate)
Type of study:
guinea pig maximisation test
Species:
guinea pig
Strain:
Dunkin-Hartley
Sex:
male/female
Details on test animals and environmental conditions:
TEST ANIMALS
- Source: David Hall Limited, UK
- Weight at study initiation: 328 to 455 g
- Housing: Solid floor polypropylene cages
- Diet (e.g. ad libitum): Guinea pig FD1 diet ad libitum
- Water (e.g. ad libitum): ad libitum
- Acclimation period: 5 days

ENVIRONMENTAL CONDITIONS
- Temperature (°C): 18-22°C
- Humidity (%): 44-64%
- Air changes (per hr): 15/hour
- Photoperiod (hrs dark / hrs light): 12 hours light/dark cycle
Route:
intradermal and epicutaneous
Vehicle:
paraffin oil
Concentration / amount:
1% induction, 5% and 2% challenge
Route:
epicutaneous, occlusive
Vehicle:
paraffin oil
Concentration / amount:
1% induction, 5% and 2% challenge
No. of animals per dose:
20
Details on study design:
RANGE FINDING TESTS:

MAIN STUDY
A. INDUCTION EXPOSURE
- No. of exposures: 2
- Exposure period: Day 1, intradermal injection; day 7, occlusive topical application for 48 hours
- Concentrations: 1% intradermal injection, 25% topical application

B. CHALLENGE EXPOSURE
- No. of exposures: 1
- Day(s) of challenge: day 21
- Exposure period: 24 hours
- Site: clipped right flank
- Concentrations: 5% and 2%
- Evaluation (hr after challenge): 24 and 48 hours after challenge removal
Positive control substance(s):
yes
Remarks:
Poison ivy oil
Reading:
1st reading
Hours after challenge:
24
Group:
test group
Dose level:
5%
No. with + reactions:
14
Total no. in group:
20
Remarks on result:
other: Reading: 1st reading. . Hours after challenge: 24.0. Group: test group. Dose level: 5%. No with. + reactions: 14.0. Total no. in groups: 20.0.
Reading:
1st reading
Hours after challenge:
24
Group:
test group
Dose level:
2%
No. with + reactions:
8
Total no. in group:
20
Remarks on result:
other: Reading: 1st reading. . Hours after challenge: 24.0. Group: test group. Dose level: 2%. No with. + reactions: 8.0. Total no. in groups: 20.0.
Reading:
1st reading
Hours after challenge:
24
Group:
positive control
Dose level:
1%
No. with + reactions:
20
Total no. in group:
20
Remarks on result:
other: Reading: 1st reading. . Hours after challenge: 24.0. Group: positive control. Dose level: 1%. No with. + reactions: 20.0. Total no. in groups: 20.0.
Interpretation of results:
sensitising
Remarks:
Migrated information Criteria used for interpretation of results: EU
Conclusions:
In a dermal sensitization study w ith cashew nutshell liquid, guinea pigs (20) w ere tested w ith in a guinea pig maximisation test. In this study, Cashew Nutshell Liquid is a dermal sensitizer.
Executive summary:

Positive reactions in 30% of test animals is required for a positive sensitising result in an adjuvant test. A positive response was elicited in 70% of animals when administered 1% induction and challenged with a 5% concentration.Greater than 60% of animals responded at a 1% intradermal induction dose, therefore sub-category 1A is appropriate.

Endpoint conclusion
Endpoint conclusion:
adverse effect observed (sensitising)

Justification for classification or non-classification

Based on the available results, the substance is classified as Skin Sensitizer Category 1A, H317 in accordance with CLP Regulation (EU) no. 1272/2008.