2-chlorobenzonitrile

Skin sensitisation

Administrative data

Endpoint:

skin sensitisation: in vivo (LLNA)

Type of information:

experimental study

Adequacy of study:

key study

Study period:

05.02.2014 - 18.02.2014

Reliability:

1 (reliable without restriction)

Rationale for reliability incl. deficiencies:

guideline study

Data source

Reference

Reference Type:

study report

Title:

Unnamed

Year:

2014

Report date:

2014

Materials and methods

Test guideline

Test guideline 1

Qualifier:

according to guideline

Guideline:

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

Test guideline 2

Qualifier:

according to guideline

Guideline:

EU Method B.42 (Skin Sensitisation: Local Lymph Node Assay)

Principles of method if other than guideline:

The relative humidity in the animal room was between approximately 23 - 45 % instead of 45 - 65 % for few hours on five days. This deviation to the study plan, however, does not affect the validity of the study.

GLP compliance:

yes (incl. QA statement)

Type of study:

mouse local lymph node assay (LLNA)

Test material

Test material information

Constituent 1

Reference substance name:

4-chlorobenzonitrile

EC Number:

210-765-4

EC Name:

4-chlorobenzonitrile

Cas Number:

623-03-0

Molecular formula:

C7H4ClN

IUPAC Name:

4-chlorobenzonitrile

Test material form:

not specified

Details on test material:

- Name of test material (as cited in study report): 4-Chlorobenzonitrile
- Analytical purity: 99.7 % (GC)
- Purity test date: 04.09.2013
- Lot/batch No.: 324601
- Expiration date of the lot/batch: 31.12.2015
- Storage condition of test material: room temperature

In vivo test system

Test animals

Species:

mouse

Strain:

CBA/Ca

Sex:

female

Details on test animals and environmental conditions:

TEST ANIMALS
- Source: Harlan Laboratories B.V., Postbus 6174, 5960 AD Horst, The Netherlands
- Age at study initiation:9-10 weeks (at beginning of treatment)
- Weight at study initiation: 19.7 - 23.7 g
- Housing: group housing, cage type: Makrolon Type II (pre-test) / III (main study), with wire mesh top.
- Diet (e.g. ad libitum): 2018C Teklad Global 18 % protein rodent diet (certified), ad libitum
- Water (e.g. ad libitum): tap water, ad libitum
- Acclimation period: 15 days prior to the start of dosing under test conditions after health examination. Only animals without any visible signs of illness were used for the study.

ENVIRONMENTAL CONDITIONS
- Temperature (°C):22 +/- 2 °C
- Humidity (%): appox. 45-65 % (except for few hours on 5 days, see deviation)

- Photoperiod (hrs dark / hrs light): artificial light 6.00 a.m. - 6.00 p.m.

Study design: in vivo (LLNA)

Vehicle:

dimethylformamide

Concentration:

Pre-Test: 25, 50 % (w/w)
Main-Test:0, 10, 25, 50 % (w/w)

No. of animals per dose:

Pre-Test: one animal per dose
Main-Test: four animals per dose

Details on study design:

see "any other information on material and methods incl. tables"

Positive control substance(s):

hexyl cinnamic aldehyde (CAS No 101-86-0)

Statistics:

The mean values and standard deviations of the body weights were calculated.

Results and discussion

Positive control results:

Last 10 positive control experiments see table 2 in "any other information on results incl. tables".

In vivo (LLNA)

Results

Results 1

Key result

Parameter:

SI

Value:

1

Test group / Remarks:

4 annimals, control group (DMF)

Results 2

Key result

Parameter:

SI

Value:

0.73

Test group / Remarks:

4 animals, test item 10 % (w/w), diluted in DMF

Results 3

Key result

Parameter:

SI

Value:

0.51

Test group / Remarks:

4 animals, test item 25 % (w/w), diluted in DMF

Results 4

Key result

Parameter:

SI

Value:

0.72

Test group / Remarks:

4 animals, test item 50 % (w/w), diluted in DMF

Results 5

Key result

Parameter:

EC3

Test group / Remarks:

0, 10, 25 and 50 % test item diluted in DMF

Remarks on result:

not determinable

Remarks:

All S.I. values are below the threshold value of 3.

Results 6

Parameter:

other: disintegrations per minute (DPM)

Test group / Remarks:

0, 10, 25 and 50 % test item diluted in DMF

Remarks on result:

other: see "any other information on results incl. tables"

Any other information on results incl. tables

1. Calculation and Results of Individual Data

Table 1: Calculation and results of individual data.

Test item concentration %	Group	Measurement DPM	Calculation			Result
			DPM-BG^a)[LM1]	number of lymph nodes	DPM per lymph node^b)	S.I.
---	BG I	273	---	---	---	---
---	BG II[LM2]	20	---	---	---	---
0	1	8083	7936.5	8	992.1	1.00
10	2	5957	5810.5	8	726.3	0.73
25	3	4219	4072.5	8	509.1	0.51
50	4	5829	5682.5	8	710.3	0.72

BG = Background; 1 = Control Group; 2-4=  Test Group;

^a)   =  The mean value was taken from the figures BG I and BG II;

^b)    =  Since the lymph nodes of the animals of a dose group were pooled, DPM/node was determined by dividing the measured value by the number of lymph nodes pooled

The EC3 value could not be calculated, since all S.I. values are below the threshold value of 3.

2. Viability / Mortality

No deaths occurred during the study period.

3. Clinical Signs

No symptoms of local skin irritation at the ears of the animals and no signs of systemic toxicity were observed during the study period.

4. Body Weights

The body weight of the animals, recorded prior to the first application and prior to treatment with³HTdR, was within the range commonly recorded for animals of this strain and age.

5. Positive Control Data

Table 2: The last 10 positive control experiments.

Positive Control Substance	Date	Concentration / Vehicle	S.I. value
α-Hexylcinnamaldehyde	October 2013	25% in acetone:olive oil (4+1, v/v)	5.8
	April 2013		5.9
	October 2012		5.7
	April 2012		3.7
	February 2012		4.7
	January 2012		10.8
	January 2012		7.1
	December 2011		5.9
	November 2011		5.3
	October 2011		6.1

Applicant's summary and conclusion

Interpretation of results:

not sensitising

Remarks:

Migrated information

Conclusions:

The test item 4-Chlorobenzonitrile was not a skin sensitiser under the test conditions of this study accroding to OECD 429 ("Skin Sensitisation: Local Lymph Node Assay").

Executive summary:

In a study according to OECD 429 ("Skin Sensitisation: Local Lymph Node Assay") 4-Chlorobenzonitrile formulated in DMF was assessed for its possible skin sensitising potential. The test was performed in mice using test item concentrations of 10, 25, and 50 % (w/w). The animals did not show any signs of systemic toxicity or local skin irritation during the course of the study and no cases of motality were observed. Stimulation Indices (S.I.) of 0.73, 0.51, and 0.72 were determined with the test item at concentrations of 10, 25, and 50 % (w/w) in DMF, respectively.

Therefore the test item 4-Chlorobenzonitrile was not a skin sensitiser under the test conditions of this study.