(3R)-1,1,3-trimethyl-2,3-dihydro-1H-inden-4-amine

Skin sensitisation

Administrative data

Endpoint:

skin sensitisation: in vivo (LLNA)

Type of information:

experimental study

Adequacy of study:

key study

Study period:

02 Oct - 14 Nov 2013

Reliability:

2 (reliable with restrictions)

Rationale for reliability incl. deficiencies:

comparable to guideline study with acceptable restrictions

Remarks:

GLP compliance unknown; deviations from guideline (3 instead of 4 animals / group, no information on positive control)

Data source

Reference

Reference Type:

study report

Title:

Unnamed

Year:

2013

Report date:

2013

Materials and methods

Test guideline

Qualifier:

equivalent or similar to guideline

Guideline:

OECD Guideline 429 (Skin Sensitisation: Local Lymph Node Assay)

Version / remarks:

adopted 22 Jul 2010

Deviations:

yes

Remarks:

- 3 instead of 4 animals/group - No information on positive control - No statistical analysis performed

GLP compliance:

not specified

Remarks:

in-house safety study

Type of study:

mouse local lymph node assay (LLNA)

Test material

Test material information

Constituent 1

Reference substance name:

(3R-)1,1,3-trimethylindan-4-amine

EC Number:

836-285-0

Cas Number:

125349-37-3

Molecular formula:

C12H17N

IUPAC Name:

(3R-)1,1,3-trimethylindan-4-amine

In vivo test system

Test animals

Species:

mouse

Strain:

other: CBA/JCrlj

Sex:

female

Details on test animals and environmental conditions:

TEST ANIMALS
- Age at study initiation: 9 weeks (preliminary study); 11 weeks (main study 1); 8 weeks (main study 2)
- Weight at study initiation: preliminary study: 20.66 - 21.60 g; main study 1: 20.66 - 25.08 g; main study 2: 18.22 - 21.32 g
- Housing: The animals were housed in suspended stainless-steel cages comprising stainless-steel wire mesh front and floor, and aluminium walls (W 176 × D 302 × H 130 mm, Yamato Scientific Co., Ltd.), 3 animals per cage. Once a week, each cage was exchanged for a washed, sterilised cage.
- Diet: solid feed for mice (CRF-1, Oriental Yeast Co., Ltd.); ad libitum
- Water: tap water via an automatic water supply unit; ad libitum
- Acclimation period: not specified

ENVIRONMENTAL CONDITIONS
- Temperature (°C): 21 - 25
- Humidity (%): 40 - 70
- Air changes (per hr): ≥ 10
- Photoperiod: 12 / 12

IN-LIFE DATES:
Preliminary study: 02 - 07 Oct 2013
Main study 1: 16 - 23 Oct 2013
Main study 2: 06 - 14 Nov 2013

Study design: in vivo (LLNA)

Vehicle:

acetone/olive oil (4:1 v/v)

Concentration:

preliminary study and main study 1: 5%, 25% (acetone/olive oil (4:1 v/v)), 100% (stock solution)
main study 2: 50% (acetone/olive oil (4:1 v/v))

No. of animals per dose:

preliminary study: 1 animal per concentration
main study 1 & 2: 3 animals per concentration and negative control

Details on study design:

PRE-SCREEN TESTS
- Compound solubility: The test substance dissolved at 50% (v/v) in acetone/olive oil (4:1 v/v). Therefore, acetone/olive oil (4:1 v/v) was selected as the medium.
- Irritation: No irritation was observed after applying the test substance at any concentration.
- Lymph node weights: In the 5%, 25% and stock solution (100%) application groups the weights were 0.0046 g, 0.0063 g and 0.0061 g, respectively.

MAIN STUDY
ANIMAL ASSIGNMENT AND TREATMENT
- Name of test method: 3H-methyl thymidine incorporation
- Criteria used to consider a positive response: The ratios [radioactivity in the groups with the test substance applied (dpm) / radioactivity in the control group (dpm)] were calculated, and a ratio (stimulation index) of ≥ 3 was assessed as positive.

TREATMENT PREPARATION AND ADMINISTRATION
For the preliminary study, acetone/olive oil (4:1 v/v) solutions with the test substance at 5% and 25%, and the stock solution (100%) were applied, respectively, to both ears of the mice at 0.025 mL/site for 3 consecutive days (1 mouse / concentration). The presence/absence of irritation (erythema) was verified on days 2, 3 and 6, counting the first day of application as day 1. On day 6, the auricular lymph nodes were collected and weighed.
For main study 1, concentrations of the test substance were set to on the basis of the results of the preliminary study (5% and 25% acetone/olive oil (4:1 v/v), and stock solution (100%)) and were applied open to both ears of the mice at 0.025 mL/site for 3 consecutive days (3 mice / concentration). Also, a control group was established wherein only acetone/olive oil (4:1 v/v) was applied. The presence/absence of irritation (erythema) was verified on days 2, 3 and 6, counting the first day of application as day 1. On day 6, [methyl-3H]thymidine that had been prepared to a concentration of 2.96 MBq (80 µCi)/mL using PBS was administered intravenously at 0.25 mL/animal. 5 hours later, cells were harvested from the auricular lymph nodes, and the radioactivity (dpm) was measured using a liquid scintillation counter.
For main study 2, the treatment concentration was selected based on the results of main study 1 (50% acetone/olive oil (4:1 v/v)) and applied to both ears of each of the 3 mice at 0.025 mL/site on 3 consecutive days. Also, a control group was established wherein only acetone/olive oil (4:1 v/v) was applied. The presence/absence of irritation (erythema) was verified on days 2, 3 and 6, counting the first day of application as day 1. On day 6, [methyl-3H]thymidine that had been prepared to a concentration of 2.96 MBq (80 µCi)/mL using PBS was administered intravenously at 0.25 mL/animal; 5 hours later, cells were harvested from the auricular lymph nodes, and the radioactivity (dpm) was measured using a liquid scintillation counter.

Positive control substance(s):

not specified

Statistics:

not performed

Results and discussion

Positive control results:

no data

In vivo (LLNA)

Results

Results 1

Key result

Parameter:

EC3

Value:

> 25 - < 50

Test group / Remarks:

50%
main study 2

Results 2

Key result

Parameter:

SI

Value:

6.85

Test group / Remarks:

50%
main study 2

Results 3

Key result

Parameter:

SI

Value:

2.5

Test group / Remarks:

25%
main study 1

Results 4

Key result

Parameter:

SI

Value:

0.637

Test group / Remarks:

5%
main study 1

Cellular proliferation data / Observations:

DETAILS ON STIMULATION INDEX CALCULATION:
Main study 1: The SI of the 5% and 25% treatment group was 0.637 and 2.5, respectively. The stock solution (100%) group was excluded from the study based on effects > 5% decrease in body weight of the animals, which was deemed unsuitable for evaluating sensitisation. The SI ratios were below 3, but due to the gap between the excluded high dose (100%) and the medium dose (25%), the study was performed again in a 50% application group (main study 2).
Main study 2: The SI of the 50% treatment group was 6.85.

EC3 CALCULATION:
The ratio was ≥ 3 in the 50% application group, and so the test substance was judged to be positive (25%
CLINICAL OBSERVATIONS
Main study 1: In the stock solution (100%) group, after completion of administration on day 2, decreased spontaneous movement and emaciation were observed in 1/3 animals. No irritation was observed at any concentration.
Main study 2: On day 3, very mild erythema was observed in 1/3 animals in the 50% application group. No clinical signs were noted.

BODY WEIGHTS:
Main study 1: Measuring the body weights in the stock solution (100%) group revealed a mean body weight decrease of 1.74 g. The mean body weight increases after test substance application in the control group, the 5% and 25% application groups were 1.24 g, 0.15 g and 1.83 g, respectively.
Main study 2: The mean body weight increases in the control group and the 50% application group were 1.31 g and 0.90 g, respectively.

LYMPH NODE WEIGHTS:
Main study 1: The mean lymph node weights in the control group and the 5% and 25% application groups were 0.0052 g, 0.0053 g and 0.0085 g, respectively.
Main study 2: The mean lymph node weights in the control group and the 50% application group were 0.0044 g and 0.0084 g, respectively.

Any other information on results incl. tables

Table 1: Results of main study 1

Dose (%)	Body weight (g)				Local reaction (ears)*			Lymph node weights (g)			Radioactivity
	Day 1	Day 6	Weight increase	Mean weight increase	Day 2	Day 3	Day 6	Individual	Group total	Group mean	Dpm	ratio
0^#	23.21	24.85	1.64	1.24	0	0	0	0.0041	0.0157	0.0052	1492
	23.85	24.26	0.41		0	0	0	0.0066
	22.76	24.43	1.67		0	0	0	0.0050
5a	25.08	24.73	-0.35	-0.15	0	0	0	0.0051	0.0160	0.0053	950	0.637
	25.00	25.43	0.43		0	0	0	0.0054
	24.91	24.37	-0.54		0	0	0	0.0055
25a	24.26	26.76	2.50	1.83	0	0	0	0.0094	0.0255	0.0085	3730	2.50
	22.78	23.67	0.89		0	0	0	0.0074
	21.61	23.70	2.09		0	0	0	0.0087
100b	20.66	-	-	-	0	-	-	-	-	-	-	-
	24.32	-	-		0	-	-	-
	24.10	-	-		0	-	-	-

#: Acetone/olive oil (4:1 v/v)

a Solution (acetone/olive oil (4:1 v/v) as medium) b Stock solution

*0: No erythema, 1: Very mild erythema, 2: Obvious erythema, 3: Moderate to severe erythema, 4: Severe erythema (beet redness) to slight crusting

 

Table 2: Results of main study 2

Dose (%)	Body weight (g)				Local reaction (ears)*				Lymph node weights (g)			Radioactivity
	Day 1	Day 6	Weight increase	Mean weight increase	Day 2	Day 3		Day 6	Individual	Group total	Group mean	Dpm	ratio
0^#	18.22	19.87	1.65	1.31	0		0	0	0.0053	0.0133	0.0044	754
	21.05	22.50	1.45		0		0	0	0.0037
	19.62	20.46	0.84		0		0	0	0.0043
50a	18.25	19.81	1.56	0.90	0		0	0	0.0100	0.0253	0.0084	5163	6.85
	20.47	21.01	0.54		0		0	0	0.0074
	21.32	21.92	0.60		0		0	0	0.0079

#: Acetone/olive oil (4:1 v/v)

a Solution (acetone/olive oil (4:1 v/v) as medium)

*0: No erythema, 1: Very mild erythema, 2: Obvious erythema, 3: Moderate to severe erythema, 4: Severe erythema (beet redness) to slight crusting

 

Applicant's summary and conclusion

Interpretation of results:

other: Skin sens 1B (H317) according to Regulation (EC) No 1272/2008.

Conclusions:

Under the present test conditions, the test substance (3R-)2,3-dihydro-1,1,3-trimethyl-1H-inden-4amine was assessed to be positive of exhibiting skin sensitising potential and meets the classification criteria Skin sens 1B (H317) according to Regulation (EC) No 1272/2008.