Charcoal

Skin sensitisation

Administrative data

Endpoint:

skin sensitisation: in vivo (LLNA)

Type of information:

experimental study

Adequacy of study:

key study

Study period:

2010-06-18 to 2010-08-09

Reliability:

1 (reliable without restriction)

Rationale for reliability incl. deficiencies:

other: Study was conducted as per OECD 429 and EU B.42 test method guidelines

Data source

Reference

Reference Type:

study report

Title:

Unnamed

Year:

2010

Report date:

2010

Materials and methods

Test guideline

Qualifier:

according to guideline

Guideline:

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

Deviations:

no

Principles of method if other than guideline:

NA

GLP compliance:

no

Type of study:

mouse local lymph node assay (LLNA)

Test material

Test material information

Constituent 1

Reference substance name:

Charcoal

EC Number:

240-383-3

EC Name:

Charcoal

Cas Number:

16291-96-6

IUPAC Name:

Carbón vegetal

Details on test material:

Identity: Charcoal (Probe 1: C-Fix = 73.3%)
CAS No.: 16291-96-6
Batch No.: 19062009 1PP
Purity: C-Fix: 73.3%; dose calculation not adjusted to purity
Storage: At room temperature, moisture protected
Expiration Date: December 2030

In vivo test system

Test animals

Species:

mouse

Strain:

other: CBA/CaOlaHsd

Sex:

female

Details on test animals and environmental conditions:

Animals: Female mice of 8-12 weeks of age and with 19-23 g of body weight were included in the study; Housing: Animals were individually housed; Lighting: 12 h light/12 h dark; Temperature: 22 ± 2°C; Relative humidity: 35 to 85%; Food: Animals had access to pelleted standard diet, ad libitum; Water: Tap water, ad libitum; Bedding: Granulated soft wood bedding; Acclimatisation: Study animals were acclimated to their housing for 5 days prior to their first day of dosing.

Study design: in vivo (LLNA)

Vehicle:

propylene glycol

Remarks:

99%

Concentration:

3 groups of 4 female mice were treated on the dorsal surface of both ears once per day for 3 days with the test article, charcoal (Probe 1: C-Fix = 73.3%) at 2.5, 5, and 10% (w/w) in the vehicle propylene glycol.

No. of animals per dose:

4 female mice

Details on study design:

ANIMAL ASSIGNMENT AND TREATMENT
- Name of test method: Local Lymph Node Assay
- Criteria used to consider a positive response: A 3-fold or greater increase in proliferative activity relative to the concurrent vehicle control is considered a positive response.

TREATMENT PREPARATION AND ADMINISTRATION:
On each day of dosing, the test article was prepared at the appropriate concentrations (w/w) by suspending the appropriate amount of test article in propylene glycol. The application volume of 25 µL was used.

4 groups of 4 female mice were treated on the dorsal surface of both ears once per day for 3 consecutive days as follows:
Group 1: vehicle, propylene glycol; Group 2: Charcoal, 2.5% (w/w); Group 3: Charcoal, 5% (w/w); and Group 4: Charcoal 10% (w/w)

On day 6, the mice were injected, i.v., with 20.4 µCi of 3H-Methyl thymidine (3HTdR) per mouse. Five hours later, the mice were euthanized and the draining auricular lymph nodes were removed. The lymph node cells were treated with 5% trichloroacetic acid (TCA) to precipitate the DNA. The resulting pellets were counted in a β-scintillation counter to determine incorporation of 3HTdR.

Positive control substance(s):

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

Statistics:

The mean values and standard deviations were calculated for the body weight parameter.

Results and discussion

In vivo (LLNA)

Results

Results 1

Parameter:

SI

Remarks on result:

other: SI: Vehicle control = 1.00; Charcoal (2.5% w/v) = 0.65; Charcoal (5% w/v) = 0.72; and Charcoal (10% w/v) = 1.11

Results 2

Parameter:

other: disintegrations per minute (DPM)

Remarks on result:

other: DPM per lymph node: Vehicle control = 295.6; Charcoal (2.5% w/v) = 192.7; Charcoal (5% w/v) = 211.4; and Charcoal (10% w/v) = 329.3

Any other information on results incl. tables

DPM and SI:

 

Test item concentration % (w/w)	Group	Measurement DPM	Calculation			Result
			DPM-BG^a)	number of lymph nodes	DPM per lymph node^b)	S.I.
---	BG I	20	---	---	---	---
---	BG II	17	---	---	---	---
0	1	2383	2365	8	295.6	1.00
2.5	2	1560	1542	8	192.7	0.65
5	3	1710	1692	8	211.4	0.72
10	4	2653	2635	8	329.3	1.11

BG=    Background (1 ml 5% trichloroacetic acid) in duplicate

1      =    Control Group

2-4=    Test Groups

S.I.=    Stimulation Index

^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 estimated concentration of the test item required to produce a 3-fold increase in draining lymph node cell proliferative activity (termed the EC3 value) can be calculated according to the equation [EC3=(a-c) [(3-d)/(b-d)] + c]. Here, (a, b) and (c, d) are respectively the co-ordinates of the two pair of data lying immediately above and below the S.I. value of 3 on the local lymph node assay dose response plot.

The EC3 value could not be calculated, since all S.I.´s are below 3.

Viability / Mortality

No deaths occurred during the study period.

Clinical Signs

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

Body Weights

The body weight of the animals, recorded prior to the first application and prior to treatment with 3HTdR, was within the range commonly recorded for animals of this strain and age.

Applicant's summary and conclusion

Interpretation of results:

not sensitising

Remarks:

Migrated information up to 10% (w/w) of charcoal (Probe 1: C-Fix 73.3%) tested

Conclusions:

The test item, charcoal (Probe 1: C-Fix = 73.3%) was not a skin sensitiser under the test conditions of this study.