Reaction mass of (1R,4S,4aR,8R,8aS)-9-(dichloromethylidene)-8-hydroxyoctahydro-1,4-methanonaphthalen-5(1H)-one and (1S,4R,4aS,8S,8aR)-9-(dichloromethylidene)-8-hydroxyoctahydro-1,4-methanonaphthalen-5(1H)-one and (1R,4S,4aR,8S,8aS)-9-(dichloromethylidene)-8-hydroxyoctahydro-1,4-methanonaphthalen-5(1H)-one and (1S,4R,4aS,8R,8aR)-9-(dichloromethylidene)-8-hydroxyoctahydro-1,4-methanonaphthalen-5(1H)-one

Skin sensitisation

Administrative data

Endpoint:

skin sensitisation: in vivo (LLNA)

Type of information:

experimental study

Adequacy of study:

supporting study

Study period:

2011-01-04 - 2011-01-18 (experimental phase)

Reliability:

3 (not reliable)

Rationale for reliability incl. deficiencies:

other: see 'Remark'

Remarks:

This study provides some useful information about the properties of the test item. However it has been conducted only at a screening level. No specific reference to a guideline is given in the study report. However it is assumed that the study design has been based on the main principles of a corresponding relevant guideline and that the study did not follow the corresponding guideline in detail. The level of information collected/reported is limited. In addition the study has not been performed according to GLP. As a consequence the reliability ("evaluating the inherent quality of a test report or publication relating to preferably standardized methodology and the way the experimental procedure and results are described to give evidence of the clarity and plausibility of the findings"), relevance ("covering the extent to which data and tests are appropriate for a particular hazard identification or risk characterization") and adequacy ("defining the usefulness of data for hazard/risk assessment purposes") of the data ("data quality") are considered as limited and an overall reliability rating of 3 (= not reliable, according to the scoring system of Klimisch et al.) has been assigned.

Data source

Reference

Reference Type:

study report

Title:

Unnamed

Year:

2011

Report date:

2011

Materials and methods

Test guideline

Qualifier:

no guideline followed

Principles of method if other than guideline:

The test substance was assessed for its skin sensitising potential using the Local Lymph Node Assay in the CBA/Ca strain of mouse, according to a general study plan of the test facility. The test is designed to assess the skin sensitising potential (delayed type hypersensitivity) of the test item following topical application to the dorsal surface of the ear. Primary lymphocyte proliferation is assessed during the sensitising (induction) phase of the response.

The assay determines the level of lymphocyte proliferation in lymph nodes draining the application site of the test item. Determination of lymphocyte proliferation is quantified by measuring the incorporation of radiolabelled thymidine in the dividing lymph node cells. The proliferation response of lymph node cells was expressed as the number of radioactive disintegrations per minute per lymph node (dpm/node) and as the ratio of 3HTdR incorporation into lymph node cells of test nodes relative to that recorded for the control nodes (Stimulation Index).

GLP compliance:

no

Remarks:

However, the study was conducted in a facility operating to Good Laboratory Practice within the UK national GLP monitoring programme. The analytical characterisation of the test material has been performed according to GLP.

Type of study:

mouse local lymph node assay (LLNA)

Test material

Test material information

Constituent 1

Reference substance name:

Reaction mass of (1R,4S,4aR,8R,8aS)-9-(dichloromethylidene)-8-hydroxyoctahydro-1,4-methanonaphthalen-5(1H)-one and (1S,4R,4aS,8S,8aR)-9-(dichloromethylidene)-8-hydroxyoctahydro-1,4-methanonaphthalen-5(1H)-one and (1R,4S,4aR,8S,8aS)-9-(dichloromethylidene)-8-hydroxyoctahydro-1,4-methanonaphthalen-5(1H)-one and (1S,4R,4aS,8R,8aR)-9-(dichloromethylidene)-8-hydroxyoctahydro-1,4-methanonaphthalen-5(1H)-one

EC Number:

700-770-4

Molecular formula:

C12H14Cl2O2

IUPAC Name:

Reaction mass of (1R,4S,4aR,8R,8aS)-9-(dichloromethylidene)-8-hydroxyoctahydro-1,4-methanonaphthalen-5(1H)-one and (1S,4R,4aS,8S,8aR)-9-(dichloromethylidene)-8-hydroxyoctahydro-1,4-methanonaphthalen-5(1H)-one and (1R,4S,4aR,8S,8aS)-9-(dichloromethylidene)-8-hydroxyoctahydro-1,4-methanonaphthalen-5(1H)-one and (1S,4R,4aS,8R,8aR)-9-(dichloromethylidene)-8-hydroxyoctahydro-1,4-methanonaphthalen-5(1H)-one

In vivo test system

Test animals

Species:

mouse

Strain:

other: CBA/Ca

Sex:

not specified

Study design: in vivo (LLNA)

Vehicle:

dimethylformamide

Concentration:

25 % w/w

No. of animals per dose:

4

Details on study design:

Following a preliminary screening test, a group of four animals was treated with 50 μl (25 μl per ear) of the test item as a solution in dimethyl formamide at a concentration of 25% w/w. A further group of four animals was treated with dimethyl formamide alone.

Results and discussion

Positive control results:

not applicable (no positive controls used in study)

In vivo (LLNA)

Results

Results 1

Parameter:

SI

Remarks on result:

other: see Remark

Remarks:

Stimulation index: 1.72 (negative) The Stimulation Index is expressed as the mean radioactive incorporation for the treatment group divided by the mean radioactive incorporation of the vehicle control group. A positive result is obtained if the stimulation index is greater than 3.0.

Results 2

Parameter:

other: disintegrations per minute (DPM)

Remarks on result:

other: See Table 3 in "Any other information on results incl. tables"

Any other information on results incl. tables

Clinical observations, bodyweight and mortality data for the preliminary screening test are given in Table 1. The ear thickness measurements and mean ear thickness changes for the preliminary screening test are given in Table 2. The radioactive disintegrations per minute (dpm) per lymph node and the Stimulation Index for the main test are given in Table 3. Individual clinical observations and mortality data for the main test are given in Table 4 and individual bodyweights are given in Table 5.

Table 1: Clinical Observations, Bodyweight and Mortality Data – Preliminary Screening Test

Concentration (% w/w) in dimethyl formamide			Bodyweight (g)		Day
					1		2		3		4	5	6
		Animal Number	Day 1	Day 6	Pre-Dose	Post Dose	Pre-Dose	Post Dose	Pre-Dose	Post Dose
25		S-1	20	19	0	0	0	0Rt	0	0Rt	0Rt	0Rt	0Rt

0 = No signs of systemic toxicity

Rt = Off white residual test item on the ears

Table 2: Measurement of Ear Thickness and Mean Ear Thickness Changes – Preliminary Screening Test

Concentration (% w/w) in dimethyl formamide	Animal Number	Ear Thickness Measurement (mm)
		Day 1		Day 2		Day 3		Day 4		Day 5		Day 6
		pre-dose		post dose		post dose
		left	right	left	right	left	right	left	right	left	right	left	right
25	S-1	0.220	0.225	0.260	0.240	0.270	0.245	0.235	0.230	0.240	0.240	0.250	0.245
overall mean (mm)		0.223		0.250		0.258		0.233		0.240		0.248
overall mean ear thickness change (%)		na		12.360		15.730		4.494		7.87		11.236

na = Not applicable

Table 3: Dpm, Dpm/Node and Stimulation Index

Concentration (% w/w) in dimethyl formamide	Dpm	Dpm/Node §a	Stimulation Index §b	Result
Vehicle	984.24	123.03	na	na
25	1694.18	211.77	1.72	Negative

§a = Dpm/node obtained by dividing the Dpm value by 8 (total number of lymph nodes)

§b = Stimulation Index of 3.0 or greater indicates a positive result

na = Not applicable

Table 4: Individual Clinical Observations and Mortality Data

Concentration (% w/w) in dimethyl formamide	Animal Number	Day 1		Day 2		Day 3		Day 4	Day 5	Day 6
		Pre-Dose	Post Dose	Pre-Dose	Post Dose	Pre-Dose	Post Dose
Vehicle	1-1	0	0	0	0	0	0	0	0	0
Vehicle	1-2	0	0	0	0	0	0	0	0	0
Vehicle	1-3	0	0	0	0	0	0	0	0	0
Vehicle	1-4	0	0	0	0	0	0	0	0	0
25	2-1	0	0Rt	0	0Rt	0	0Rt	0	0	0
25	2-2	0	0Rt	0	0Rt	0	0Rt	0Rt	0Rt	0Rt
25	2-3	0	0Rt	0	0Rt	0	0Rt	0Rt	0Rt	0Rt
25	2-4	0	0Rt	0	0Rt	0	0Rt	0Rt	0Rt	0Rt

0 = No signs of systemic toxicity

Rt = Off white residual test item on the ears

Table 5: Individual Bodyweights and Bodyweight Changes

Concentration (% w/w) in dimethyl formamide	Animal Number	Bodyweight (g)		Bodyweight Change (g)
		Day 1	Day 6
Vehicle	1-1	18	17	-1
Vehicle	1-2	21	21	0
Vehicle	1-3	17	17	0
Vehicle	1-4	18	19	1
25	2-1	22	22	0
25	2-2	20	20	0
25	2-3	21	20	-1
25	2-4	22	21	-1

Applicant's summary and conclusion

Interpretation of results:

other: see Conclusions below

Conclusions:

This study provides some useful information about the properties of the test item. However it has been conducted only at a screening level. No specific reference to a guideline is given in the study report. However it is assumed that the study design has been based on the main principles of a corresponding relevant guideline and that the study did not follow the corresponding guideline in detail. The level of information collected/reported is limited. In addition the study has not been performed according to GLP.

As a consequence the reliability ("evaluating the inherent quality of a test report or publication relating to preferably standardized methodology and the way the experimental procedure and results are described to give evidence of the clarity and plausibility of the findings"), relevance ("covering the extent to which data and tests are appropriate for a particular hazard identification or risk characterization") and adequacy ("defining the usefulness of data for hazard/risk assessment purposes") of the data ("data quality") are considered as limited and an overall reliability rating of 3 (= not reliable, according to the scoring system of Klimisch et al.) has been assigned.

Executive summary:

A screening level in vivo study for skin sensitisation has been conducted (Local Lymph Node Assay (LLNA) in the mouse). The test item was considered to be a non-sensitiser under the conditions of the test.