Heavy wax residue fraction, waste plastic derived, hydrothermal conversion

Skin sensitisation

Administrative data

Endpoint:

skin sensitisation: in vivo (LLNA)

Type of information:

experimental study

Adequacy of study:

key study

Study period:

From 11 April 2022 to 9 May 2022

Reliability:

1 (reliable without restriction)

Data source

Reference

Reference Type:

study report

Title:

Unnamed

Year:

2022

Report date:

2022

Materials and methods

Test guideline

Qualifier:

according to guideline

Guideline:

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

GLP compliance:

yes

Type of study:

mouse local lymph node assay (LLNA)

Test material

Test material information

Constituent 1

Reference substance name:

Heavy wax residue fraction, waste plastic derived, hydrothermal conversion

Molecular formula:

Not applicable - UVCB

IUPAC Name:

Heavy wax residue fraction, waste plastic derived, hydrothermal conversion

Test material form:

other: solid - lumps

Details on test material:

- Stability: stable under storage conditions
- Storage: room temperature (20 ± 5 °C)
- Appearance: large, brown, solid lumps
- Purity: not applicable, UVCB
- Homogeneity: homogeneous
- EC no.: 950-445-8

In vivo test system

Test animals

Species:

mouse

Strain:

CBA/Ca

Remarks:

CBA/CaOlaHsd

Sex:

female

Details on test animals and environmental conditions:

- Source: Anlab, Praha, Czech Republic
- Females (if applicable) nulliparous and non-pregnant: yes, 32 animals
- Age at study initiation: 9 weeks for Pre-screen test, 11 weeks for main study
- Housing conditions: The animals were housed in TECNIPLAST cages from the Tecniplast Company, in Conventional animal room No. B2-609. Maximum six females were housed per cage. The room temperature was within the range of 20-24 oC, relative humidity was within the range of 40-60 %. The animals were subjected to a 12-hour light/ 12-hour dark cycle
- Diet (e.g. ad libitum): A laboratory food Teklad Global 16% Protein Rodent Diet was served ad libitum
- Water (e.g. ad libitum): The animals received tap water for human consumption ad libitum. The water from the local mains was monitored for quality by testing for the microbiological and chemical quality by Waterworks Bratislava quarterly. Bottles were exchanged and cleaned once a week. The quality of drinking water is periodical monitored (including microbiological control) and recorded
- Acclimation period: 5 days
- Animal Health: The health condition of animals and skin integrity was examined by a veterinarian before initiation of the study (Pre-screen test and Main study). Animals were healthy, without visible symptoms of disease.

Study design: in vivo (LLNA)

Vehicle:

acetone/olive oil (4:1 v/v)

Concentration:

25 %, 10 % and 5 % (w/v) based on solubility and pre-screen test results.

No. of animals per dose:

5 animals per treatment group for main study

Details on study design:

Preliminary laboratory tests showed that test item is partially soluble in acetone/olive oil (AOO, 4:1 v/v) and methyl ethyl ketone and creates non-homogenous suspension. Selection of the doses was limited by suspensibility of the test item in recommended vehicles. The Pre-screen test was performed using the concentrations of 25 %, 10 % and 5 % (w/v). Based on absence of clinical symptoms, either of local irritation at the application site or systemic toxicity in the Pre-screen test, the same concentrations of 5 % (w/v), 10 % (w/v) and 25 % (w/v) were selected for the Main study.

Day 1: Each animal was identified, and the body weight was recorded. To the dorsum of each ear 25 µL of the appropriate dilution of the test item, positive control, or the vehicle alone was applied.
Days 2 and 3: The application procedure carried out on day 1 was repeated.
Days 4 and 5: No treatment.
Day 6: The body weight of each animal was recorded. 250 μL of sterile phosphate-buffered saline (PBS) containing 20 μCi (7.4×105 Bq) of tritiated (3H)-methyl thymidine was injected into all test and control mice via the tail vein.
Five hours later, the animals were sacrificed. The draining auricular lymph nodes from each ear were excised and pooled in PBS for each experimental group (pooled treatment group approach).

Positive control substance(s):

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

Results and discussion

Positive control results:

Stimulation index SI = 7.82 for positive control was in the middle of the range of historical positive controls

In vivo (LLNA)

Results

Results 1

Key result

Parameter:

SI

Value:

0.51

Test group / Remarks:

5 % test item

Results 2

Key result

Parameter:

SI

Value:

1.16

Test group / Remarks:

10 % test item

Results 3

Key result

Parameter:

SI

Value:

0.66

Test group / Remarks:

25 % test item

Applicant's summary and conclusion

Interpretation of results:

GHS criteria not met

Conclusions:

Based on the results of this study, HEAVY WAX RESIDUE FRACTION, WASTE PLASTIC DERIVED, HYDROTHERMAL CONVERSION is not considered a skin sensitizer under the conditions of this LLNA study.

Executive summary:

The sensitization potential of HEAVY WAX RESIDUE FRACTION, WASTE PLASTIC DERIVED, HYDROTHERMAL CONVERSION was evaluated using Local Lymph Node Assay (LLNA). The LLNA has been developed to determine the allergic contact sensitization potential of chemicals.

Based on the recommendations of the OECD Guideline No. 429, the test item was dissolved in acetone/olive oil (AOO, 4:1 v/v). The positive control (a-Hexylcinnamic aldehyde) was dissolved in AOO. Selection of the doses was limited by suspensibility of the test item in recommended vehicles. The Pre-screen test was performed using the concentrations of 25 %, 10 % and 5 % (w/v). Based on absence of clinical symptoms, either of local irritation at the application site or systemic toxicity in the Pre-screen test, the same concentrations of 5 % (w/v), 10 % (w/v) and 25 % (w/v) were selected for the Main study.

In the Main study, five female mice (CBA/CaOlaHsd) per group were topically exposed (dorsum of both ears) to the test item at concentrations of 5 % (w/v), 10 % (w/v) and 25 % (w/v) in AOO. Positive control was exposed to the 25 % solution of a-Hexylcinnamic aldehyde in AOO and negative control to the vehicle (AOO), only. Lymphocyte proliferation was measured using incorporation of radioactive ³H-methyl-thymidine in the draining lymph nodes. The radioactive incorporation was measured as disintegrations per minute (DPM). Results were expressed as Stimulation Index (SI) calculated as DPM/pooled treatment group divided by DPM/vehicle control group.

Regardless of the dose applied, daily clinical observation of animals did not show visible clinical signs of systemic toxicity in all HEAVY WAX RESIDUE FRACTION, WASTE PLASTIC DERIVED, HYDROTHERMAL CONVERSION treated mice. No signs of significant local irritation were observed in test item treated groups.

In this study, the Stimulation Indices (SI) of 0.51, 1.16 and 0.66 were determined with the test item at concentrations of 5 % (w/v), 10 % (w/v) and 25 % (w/v) HEAVY WAX RESIDUE FRACTION, WASTE PLASTIC DERIVED, HYDROTHERMAL CONVERSION in AOO, respectively. The EC3 value could not be calculated since none of the tested concentrations induced a SI greater than the threshold value of 3.

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