[No public or meaningful name is available]

Skin sensitisation

Administrative data

Endpoint:

skin sensitisation: in vivo (LLNA)

Type of information:

experimental study

Adequacy of study:

key study

Study period:

2012-03-01 to 2012-03-14

Reliability:

1 (reliable without restriction)

Data source

Reference

Reference Type:

study report

Title:

Unnamed

Year:

2012

Materials and methods

Test guideline

Test guideline 1

Qualifier:

according to guideline

Guideline:

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

Deviations:

no

Test guideline 2

Qualifier:

according to guideline

Guideline:

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

Deviations:

no

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:

Reaction mass of tri- µ -(2-ethylhexanoato-O)-bis(N,N',N''-trimethyl-1,4,7-triazacyclononane-N,N',N'')dimanganese and µ -(acetato-O)-di- µ -(2-ethylhexanoato-O)-bis(N,N',N''-trimethyl-1,4,7-triazacyclononane-N,N',N'')dimanganese

IUPAC Name:

Reaction mass of tri- µ -(2-ethylhexanoato-O)-bis(N,N',N''-trimethyl-1,4,7-triazacyclononane-N,N',N'')dimanganese and µ -(acetato-O)-di- µ -(2-ethylhexanoato-O)-bis(N,N',N''-trimethyl-1,4,7-triazacyclononane-N,N',N'')dimanganese

Constituent 2

Reference substance name:

Reaction mass of tri-µ-(2-ethylhexanoato-O)-bis(N,N',N''-trimethyl-1,4,7-triazacyclononane-N,N',N'')dimanganese and µ-(acetato-O)-di-µ-(2-ethylhexanoato-O)-bis(N,N',N''-trimethyl-1,4,7-triazacyclononane-N,N',N'')dimanganese

EC Number:

700-806-9

Molecular formula:

C42H87Mn2N6O6 and C36H75Mn2N6O6

IUPAC Name:

Reaction mass of tri-µ-(2-ethylhexanoato-O)-bis(N,N',N''-trimethyl-1,4,7-triazacyclononane-N,N',N'')dimanganese and µ-(acetato-O)-di-µ-(2-ethylhexanoato-O)-bis(N,N',N''-trimethyl-1,4,7-triazacyclononane-N,N',N'')dimanganese

Details on test material:

- Name of test material (as cited in study report): Manganese Octoate/Acetate - MeTACN complex
- Physical state: white solid
- Lot/batch No.: 16D16603
- Storage condition of test material: room temperature under nitrogen in the dark

In vivo test system

Test animals

Species:

mouse

Strain:

other: CBA/Ca (CBA/CaOlaHsd)

Sex:

female

Details on test animals and environmental conditions:

TEST ANIMALS
- Source: Harlan Laboratories UK Ltd., Oxon, UK.
- Age at study initiation: 8 -12 weeks
- Weight at study initiation: 15-23g
- Housing: suspended solid-floor polypropylene cages furnished with softwood woodflakes.
- Diet (e.g. ad libitum): ad libitum 2014C Teklad Global Rodent diet
- Water (e.g. ad libitum): ad libitum mains tap water
- Acclimation period: at least 5 days

ENVIRONMENTAL CONDITIONS
- Temperature (°C): 19 - 25°C
- Humidity (%): 30 - 70%
- Air changes (per hr): 15
- Photoperiod (hrs dark / hrs light): 12/12

Study design: in vivo (LLNA)

Vehicle:

other: ethanol/distilled water 7:3

Concentration:

25%, 10%, 5%

No. of animals per dose:

5

Details on study design:

RANGE FINDING TESTS:
No signs of systemic toxicity, visual local skin irritation or irritation indicated by an equal to or greater than 25% increase in mean ear thickness were noted.

MAIN STUDY
- Criteria used to consider a positive response: The test item will be regarded as a sensitiser if at least one concentration of the test item results in a threefold or greater increase in 3HTdR incorporation compared to control values. Any test item failing to produce a threefold or greater increase in 3HTdR incorporation will be classified as a "non-sensitiser".

TREATMENT PREPARATION AND ADMINISTRATION:
25 µl of test item was applied to the entire dorsal surface of each ear of each mouse. The application was repeated on days 2 and 3). On day 6 an injection of 250 µl phosphate buffered saline (PBS) containing 20 µCi of 3H-methyl thymidine (3H-TdR) was made into the tail vein of each experimental mouse. Five hours later, the draining Auricular lymph node of each ear was excised into PBS. A single cell suspension of the lymph node cells for each individual animal was prepared by gentle mechanical disaggregation through a 200-mesh stainless steel gauze. The lymph node cells were rinsed through the gauze with 4 ml of PBS into a petri dish labelled with the project number and dose concentration. The lymph node cells suspension was transferred to a centrifuge tube. The petri dish was washed with an additional 5 ml of PBS to remove all remaining lymph node cells and these were added to the centrifuge tube. The lymph node cells were pelleted at 1400 rpm (approximately 190 g) for ten minutes. The pellet was resuspended in 10 ml of PBS and re-pelleted. To precipitate out the radioactive material, the pellet was resuspended in 3 ml of 5% Trichloroacetic acid (TCA). Cells were precipitated with 5% trichloroacetic acid at 4 °C for 18 hours.

Positive control substance(s):

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

Statistics:

Data was processed to give group mean values for disintegrations per minute and standard deviations where appropriate. Individual and group mean disintegrations per minute values were assessed for dose response relationships by analysis of homogeneity of variance followed by one way analysis of variance (ANOVA). In the event of a significant result from the ANOVA, pairwise comparisons were performed between control and treated groups. For homogenous datasets Dunnett’s Multiple Comparison test was used and for non-homogenous datasets Dunnett’s T3 Multiple Comparison Method was used.

Probability values (p) are presented as follows:
P<0.001 ***
P<0.01 **
P<0.05 *
P>0.05 (not significant)

Results and discussion

Positive control results:

Stimulation Index: 6.16
α-Hexylcinnamaldehyde, tech., 85% was considered to be a sensitiser under the conditions of the test.

In vivo (LLNA)

Results

Results 1

Parameter:

other: disintegrations per minute (DPM)

Remarks on result:

other: See table below

Results 2

Parameter:

SI

Remarks:

5% concentration

Value:

1.28

Test group / Remarks:

5 animals

Remarks on result:

other: See table below

Results 3

Parameter:

SI

Remarks:

10% concentration

Value:

1.55

Test group / Remarks:

5 animals

Remarks on result:

other: see table below

Results 4

Parameter:

SI

Remarks:

25% concentration

Value:

2.3

Test group / Remarks:

5 animals

Remarks on result:

other: see table below

Any other information on results incl. tables

Concentration(% w/w) in ethanol/distilled water 7:3	Animal Number	dpm/Animal	Mean dpm/Animal (Standard Deviation)	Stimulation Index	Result
Vehicle	1-1	714.22	905.06(±151.44)	na	na
	1-2	840.23
	1-3	1127.84
	1-4	899.77
	1-5	943.25
5%	2-1	1256.18	1159.33 (±307.06)	1.28	Negative
	2-2	1218.28
	2-3	783.47
	2-4	954.06
	2-5	1584.64
10%	3-1	1483.33	1400.02 (±538.84)	1.55	Negative
	3-2	2117.47
	3-3	665.79
	3-4	1588.57
	3-5	1144.93
25%	4-1	2910.76	2082.68 (±955.78)	2.3	Negative
	4-2	1729.3
	4-3	743.71
	4-4	1932.97
	4-5	3096.68

The results of the statistical analysis of the data indicated there was no significant difference between the control group and the test groups

Applicant's summary and conclusion

Interpretation of results:

not sensitising

Remarks:

Migrated information Criteria used for interpretation of results: EU

Conclusions:

The test item is not sensitising.