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Toxicological information

Skin sensitisation

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Administrative data

Endpoint:
skin sensitisation: in vivo (LLNA)
Type of information:
experimental study
Adequacy of study:
key study
Reliability:
1 (reliable without restriction)
Rationale for reliability incl. deficiencies:
guideline study

Data source

Reference
Reference Type:
study report
Title:
Unnamed
Year:
2010
Report Date:
2010

Materials and methods

Test guidelineopen allclose all
Qualifier:
according to
Guideline:
OECD Guideline 429 (Skin Sensitisation: Local Lymph Node Assay)
Qualifier:
according to
Guideline:
EU Method B.42 (Skin Sensitisation: Local Lymph Node Assay)
Principles of method if other than guideline:
Test material is 'Naphtha (petroleum), steam-cracked, C8-10 aromatic hydrocarbon fraction, alkylated and oligomerised' in form of the technical product Novares L 100.
GLP compliance:
yes (incl. certificate)
Type of study:
mouse local lymph node assay (LLNA)

Test material

Reference
Name:
Unnamed
Test material form:
liquid: viscous
Details on test material:
- Manufacturers identification: Novares TL 10, Novares L 100, Novares L 800, Novares C 10 (different types/technical products of the substance)
- Substance type: organic
- Test material is 'Naphtha (petroleum), steam-cracked, C8-10 aromatic hydrocarbon fraction, alkylated and oligomerised', EC list number 701-299-7 (assigned by ECHA after request for change of EC number/EC name). Originally, the substance Hydrocarbons, C9-unsaturated, polymerised, CAS No. 71302- 83-5, was submitted for registration. Subsequent to substance validation, the identity of the substance was changed by ECHA (EC number 615-276-2 and new name 'Distillates (petroleum), steam-cracked, C8-12 fraction", mainly C8 - C10 unsaturated aromatic and alkylaromatic hydrocarbons, predominantly styrene- and indene-derivatives, obtained by Lewis acid-initiated alkylation and polymerisation').
- for additional information see respective study records
Specific details on test material used for the study:
- Name of test material (as cited in study report): Novares L 100; CAS-no. 71302-83-5 (Hydrocarbons, C9- unsaturated, polymerized)
- Composition of test material, percentage of components: see Section 1.2 Composition
- Lot/batch No.: 28738
- Stability under test conditions: no measured data; based on chemical structure assumed to be stable
- Storage condition of test material: room temperature, exclusion of light

In vivo test system

Test animals

Species:
mouse
Strain:
Balb/c
Sex:
female
Details on test animals and environmental conditions:
TEST ANIMALS
- Source: Breeding farm VELAZ s.r.o., Koleč u Kladna, Czech Republic
- Age at study initiation: 8 - 10 wks
- Weight at study initiation: 17 - 19 g
- Housing: macrolon cages
- Diet: ad libitum
- Water: ad libitum
- Acclimation period: at least 5 days

ENVIRONMENTAL CONDITIONS
- Temperature (°C): 22 ± 3
- Humidity (%): 30 – 70
- Photoperiod (hrs dark / hrs light): 12 / 12

Study design: in vivo (LLNA)

Vehicle:
other: DAE 433 - mixture of 40% dimethylacetamide, 30% acetone, and 30% ethanol
Concentration:
0.3, 3.0 and 30% test item in vehicle (DAE 433)
No. of animals per dose:
5
Details on study design:
RANGE FINDING TESTS:
- Compound solubility: dispersion
- Irritation: no
- Lymph node proliferation response: not examined

MAIN STUDY

ANIMAL ASSIGNMENT AND TREATMENT
- Criteria used to consider a positive response:
Stimulation index (SI) is ≥ 3
Response increases in dose-related manner
- Criteria for irritation potential: individual and mean ear weight

TREATMENT PREPARATION AND ADMINISTRATION:
Days 1-3: application of 25 µL test suspension (test item in DAE 433), 1x on day 1, 2, and 3
Days 4 and 5: no treatment
Day 6: Injection of 250 μL of phosphate-buffered saline (PBS) containing 7.5E+5 Bq (ca. 21 µCi) of 3H-methyl thymidine into all test and control mice via the tail vein. Five hours later, the animals were killed.
Positive control substance(s):
other: DNCB (dinitrochlorbenzene, 0.5% (w/v) solution)
Statistics:
Non-parametric Kruskal-Wallis test and then the non-parametric two-group Mann-Whitney rank test (probability level 0.05) were applied to all two-group comparisons [using software Statgraphic ® Centurion (version XV, USA)].

Results and discussion

Positive control results:
The positive control substance produced the intended effect. A SI of 18.49 was obtained.

In vivo (LLNA)

Resultsopen allclose all
Key result
Parameter:
EC3
Value:
0.585
Key result
Parameter:
SI
Value:
1.54
Test group / Remarks:
0.3% test item in vehicle
Key result
Parameter:
SI
Value:
15.35
Test group / Remarks:
3% test item in vehicle
Key result
Parameter:
SI
Value:
17.06
Test group / Remarks:
30% test item in vehicle
Cellular proliferation data / Observations:
CELLULAR PROLIFERATION DATA
Measured radioactivity data (DPM) are reported in Table 1 under 'Any other information on results incl. tables'

DETAILS ON STIMULATION INDEX CALCULATION
-Group means were calculated from the radioactivity measurements (DPM) of the individual test animals. Stimulation indices were calculated by dividing the group means of the positive control group and the test groups by the group mean of the negative control group.

EC3 CALCULATION
Equation for EC3 calculation
EC3 = c + [(3 - d) / (b - d] * (a - c),
with a = the lowest concentration in % giving a stimulation index >3;
b = the actual stimulation index caused by a;
c = the highest concentration in % failing to produce a stimulation index of 3;
d = the actual stimulation index caused by c.
Note: The vehicle control data (at SI = 1) should not be used for c and d.

Any other information on results incl. tables

Table 1: Individual and mean radioactivity in cell suspensions (Report Table 4)

Activity (DPM)

Animal No.

NC

(1-5)

PC

(6-10)

30%

(11-15)

3%

(16-20)

0.3%

(21-25)

1

550.98

11374.69

9833.62

7176.23

1061.30

2

485.39

10401.94

9768.19

10503.12

539.22

3

568.93

10347.82

8392.34

6681.56

404.66

4

526.30

8837.38

10204.93

6586.68

1621.48

5

596.74

9493.34

8333.70

10939.31

570.46

 Group mean

545.67

10091.03

9306.56

8377.38

839.42

SI

1.00

18.49

17.06

15.35

1.54

NC = vehicle control; PC = positive control

Table 2: Individual and average ear weights (Report Table 5)

Animal No.

Weight of ear biopsies (milligrams)

NC

(1-6)

PC

(7-12)

30%

(13-18)

3%

(19-24)

0.3%

(25-30)

1

24.5

27.4

29.3

30.3

22.2

2

25.2

27.5

28.8

31.5

23.7

3

23.8

26.5

27.9

28.8

23.6

4

23.4

26.7

28.4

29.2

24.1

5

24.7

27.2

33.6

29.7

23.2

 Group mean

24.32

27.06*

29.60*

29.90*

23.36

SD

0.72

0.44

2.29

1.06

0.72

NC = vehicle control; PC = positive control

* statistically significant with p =< 0.05, Mann-Whitney test

Table 3:  Summary table (Report Table 6)

Group

Radioisotope incorporation
in lymph nodes

Ear weight

Mean DPM

SI

Mean (mg)

NC

545.67

1.0

24.32

PC

10091.03

18.49+

27.06

30%

9306.56

17.06+

29.60

3%

8377.38

15.35+

29.90

0.3%

839.42

1.54

23.36

Bold figures with + = SI values 3

Applicant's summary and conclusion

Interpretation of results:
Category 1A (indication of significant skin sensitising potential) based on GHS criteria
Conclusions:
In a LLNA test according to OECD TG 429, test material concentrations of 3% and 30% in vehicle resulted in stimulation indices above 3 (15.35 and 17.06). The EC3 was calculated to 0.585 indicating that the test substance has a high potential for skin sensitisation.