Phenol, paraalkylation products with C12-rich branched olefins derived from propene oligomerisation, reaction products with sulphur monochloride and decene, reaction products with Benzoic acid, 2-hydroxy-,C14-18 alkyl dervis., polybutenyl benzenesulphonic acid, carbon dioxide and calcium hydroxide

Endpoint summary

Description of key information

EC 903-162-9 was found to be non irritating in an in vitro skin study conducted to GLP standards and in accordance with OECD guidelines.
EC 903-162-9 was found to be non irritating in both an in vitro eye irritation study conducted to GLP standards and subsequently an in vivo study conducted in accordance with OECD guidelines and to GLP standards. 

Key value for chemical safety assessment

Skin irritation / corrosion

Link to relevant study records

Reference

Endpoint:

skin irritation / corrosion

Remarks:

other: In Vitro study conducted in accordance with OECD 439 guidance

Type of information:

experimental study

Adequacy of study:

key study

Study period:

18 January 2012 to 01 June 2012

Reliability:

1 (reliable without restriction)

Rationale for reliability incl. deficiencies:

other: Study conducted in accordance with current guidelines and GLP compliant.

Qualifier:

according to guideline

Guideline:

other: OECD (2010), In Vitro Skin Irritation: Reconstructed Human Epidermis Test Method, OECD Guidelines for the Testing of Chemicals No. 439, OECD, Paris.

GLP compliance:

yes

Remarks on result:

other: EpiSkin viability of 103.09% ±13.43% (mean of triplicate values). EC 903-162-9 was demonstrated to be non-irritant when tested in the EpiSkin in vitro irritation assay.

Irritant / corrosive response data:

EpiSkin viability of 103.09% ±13.43% (mean of triplicate values)

MTT Direct Reduction Test (Preliminary Assay)

 

The test was scored by visual assessment of the formation of purple-coloured formazan. The negative control (PBS) did not reduce MTT to formazan. The positive control (eugenol) and EC 903-162-9 were reduced MTT to formazan.

 

EpiSkin^®Irritation Test

 

Percentage Viability of EpiSkin^® Tissues After ca 41h Recovery Time.

Treatment	Mean viability per tissue (%)	Mean viability per test item (%)	SD (%)
EC 903-162-9 (corrected)	91.31	103.09	13.43
	100.26
	117.71
Aqueous SDS Solution (5%, w/v) (Positive Control)	12.82	13.22	0.35
	13.39
	13.44
PBS Solution (Negative Control)	105.31	100.00	7.75
	91.10
	103.59

Negative Controls

 

The negative control results were similar for the three viable EpiSkin^®units dosed with

Dulbecco’s PBS. Exposure to Dulbecco’s PBS resulted in a mean EpiSkin^®viability of

100.00% ±7.75%.

 

Positive Controls

 

The positive control results were similar for the three viable EpiSkin^® units dosed with

aqueous SDS solution (5%, w/v). Exposure to aqueous SDS solution (5%, w/v) resulted in amean EpiSkin^® viability of 13.22% ±0.35%.

 

EC 903-162-9

 

Non-Viable Controls

 

The mean absorbance value of the three replicate non-viable tissues dosed with EC 903-162-9 was 0.148 ± 0.034. The mean absorbance value of the three replicate un-dosed non-viable tissues was 0.098±0.016. Therefore, the absorbance value for the effect of the un-removed test item was 0.050. This value was subtracted from the absorbance value of the viable tissues dosed with EC-903-162-9.

 

Viable Tissues

 

The results were similar for the three viable EpiSkin^®units dosed with EC 903-162-9.

Exposure to EC 903-162-9 resulted in a mean EpiSkin^®viability (corrected for effect of un-removed test item) of 103.09% ± 13.43% of the negative control value.

Interpretation of results:

not irritating

Remarks:

Migrated information Criteria used for interpretation of results: OECD GHS

Conclusions:

In conclusion, EC 903-162-9 was demonstrated to be non-irritant when tested in the EpiSkin in vitro irritation assay.

Executive summary:

Evaluation of skin irritation is part of the Human Health Hazard Assessment required for registration of a chemical. In this study, the irritation potential of EC 903-162-9 was evaluated using the SkinEthic EpiSkin in vitro irritation assay. Prior to the conduct of the irritation assay, a preliminary test was conducted to assess the intrinsic ability of the test item to reduce methylthiazoldiphenyl-tetrazolium bromide (MTT) to formazan. The test item was capable of MTT reduction. Therefore, non-viable control tissues were dosed in parallel with the irritation assay to quantify this effect and the results were corrected accordingly. The dermal irritation potential was assessed by applying an aliquot ca 10 µL of EC 903-162-9 to the exposed surface of three EpiSkin reconstructed human epidermis (RhE) units for 15 min. The surface area of the EpiSkin was 0.38 cm², therefore the application rate was 26.3 µL/cm². After the 15 min exposure period, the test item was washed from the surface of the EpiSkin using Dulbecco’s phosphate-buffered saline (PBS) and tissue swabs. The EpiSkin was then incubated for a recovery period of ca 41 h in a humidified incubator set to maintain temperature and CO₂ levels of 37°C and 5%, respectively. Following incubation, the EpiSkin units were transferred to assay medium containing MTT (0.3 mg/mL) and returned to the incubator for 3 h. Biopsies of the EpiSkin membranes were then removed, added to acidified isopropanol, and refrigerated for ca 69 h in order to extract the formazan. The formazan production (cell viability) was assessed by measuring the optical density of the extracts at a wavelength of 550 nm. Three replicates of the positive control, aqueous sodium dodecyl sulphate (SDS) solution (5%, w/v) (10 µL), and the negative control, PBS (10 µL) were tested in parallel to demonstrate the efficacy of the assay. The viability of each individual EpiSkin tissue was calculated as a percentage of the mean negative control viability (defined as 100%). Exposure to EC-903-162-9 resulted in a mean EpiSkin viability of 103.09% ± 13.43% of the negative control value. Exposure to the positive control, aqueous SDS solution (5%, w/v), resulted in a mean EpiSkin viability of 13.22% ± 0.35% of the negative control value. In conclusion, EC 903-162-9 was demonstrated to be non-irritant when tested in the EpiSkin in vitro irritation assay.

Endpoint conclusion

Endpoint conclusion:

no adverse effect observed (not irritating)

Eye irritation

Link to relevant study records

Reference

Endpoint:

eye irritation: in vivo

Type of information:

experimental study

Adequacy of study:

key study

Study period:

29 January to 08 February 2013

Reliability:

1 (reliable without restriction)

Rationale for reliability incl. deficiencies:

other: Compliant with current guidelines and GLP compliant

Qualifier:

according to guideline

Guideline:

OECD Guideline 405 (Acute Eye Irritation / Corrosion)

Deviations:

no

GLP compliance:

yes

Species:

rabbit

Strain:

New Zealand White

Details on test animals or tissues and environmental conditions:

TEST ANIMALS
- Source: Born at Charles River, Edinburgh
- Age at study initiation: Approximately 9 months old
- Weight at study initiation: 4.525 kg and 4.127 kg
- Housing: Singly housed
- Diet (e.g. ad libitum): ad libitum
- Water (e.g. ad libitum): ad libitum
- Acclimation period: At least 5 days

ENVIRONMENTAL CONDITIONS
- Temperature (°C): 18°C on each day
- Humidity (%): 42% to 58%
- Air changes (per hr): 10
- Photoperiod (hrs dark / hrs light): 12 hrs light / 12 hrs dark

IN-LIFE DATES: From: To: 29 January to 08 February 2013

Vehicle:

unchanged (no vehicle)

Controls:

not required

Amount / concentration applied:

TEST MATERIAL
- Amount(s) applied (volume or weight with unit): 0.1 mL
- Concentration (if solution): Not applicable

Duration of treatment / exposure:

Single dose

Observation period (in vivo):

72 h

Number of animals or in vitro replicates:

Two

Details on study design:

REMOVAL OF TEST MATERIAL
- Washing (if done): None

SCORING SYSTEM:
OECD Test Guideline scoring system
TOOL USED TO ASSESS SCORE: Hand held magnifier and pen torch (when necessary). When required only, fluorescein and ultraviolet illumination

Irritation parameter:

overall irritation score

Basis:

mean

Time point:

other: 24, 48 and 72 h

Score:

0

Max. score:

0

Reversibility:

other: Not applicable

Other effects:

Slight ocular discharge at 1 h and 4 h post instillation

Interpretation of results:

not irritating

Remarks:

Migrated information Criteria used for interpretation of results: EU

Conclusions:

Using the criteria described in regulation (EC) 1272/2008, under the conditions of the study, EC 903-162-9 would be considered to be non-irritating to the rabbit eye.

This study is considered to be relevant, reliable and adequate for risk assessment and for classification purposes.

Executive summary:

This study investigated the eye irritation potential of EC-903-162-9 after a single instillation to the rabbit eye.

Two female rabbits received 0.1 mL of EC-903-162-9, instilled, as supplied, into the conjunctival sac of the right eye. The left eye remained untreated and hence it acted as a control. Both eyes were examined for evidence of irritation approximately 1, 4, 24, 48 and 72 h after instillation.

No irritation was noted in the control eyes of either rabbit at any observation timepoint.

There were no reactions in the iris of either animal. Corneal opacity, slight conjunctival redness and a slight discharge were noted 1 h after test item instillation in one animal. However, after the eye was stained with Fluorescein and the eye examined under ultraviolet illumination, no area of opacity was found. Slight discharge was noted 4 h after test item instillation in both animals.

Using the criteria described in the European Union Council Directive 67/548/EEC and Regulation (EC) No 1272/2008 (CLP) under the conditions of the study, EC-903-162-9 would be considered to be non-irritating to the rabbit eye.

Endpoint conclusion

Endpoint conclusion:

no adverse effect observed (not irritating)

Respiratory irritation

Endpoint conclusion

Endpoint conclusion:

no study available

Additional information

Dermal Irritation

The in vitro skin irritation study conducted in accordance with OECD guideline 439, using the EpiSkin test system showed the registration substance to be non irritating, this data combined with lack of irritation seen during an acute dermal study on a structurally similar material (EC 903-161-3) up to the limit dose tested is sufficient to conclude that the material is not a dermal irritant. This testing strategy complies with that set out in REACH Annex VII and VIII.

Eye Irritation

The in vitro eye irritation study showed the registration substance to be non irritating within this test system. From this data it was considered that progression to in vivo eye irritation study was appropriate as there was no indication that the registration material was either corrosive or irritating to the eyes. No eye irritation was seen in the in vivo eye irritation study which was conducted in accordance with OECD 405 guidelines and to GLP standards. It was therefore concluded that the registration substance is not irritating to the eyes. This testing strategy complies with that set out in REACH Annex VII and VIII.

Justification for selection of skin irritation / corrosion endpoint:
Key study conducted in accordance with OECD guideline 439 and GLP.

Justification for selection of eye irritation endpoint:
Key study conducted in accordance with OECD guideline 405 and GLP.

Justification for classification or non-classification

Neither the in vitro Episkin test system that tested EC 903-162-9 or the acute dermal toxicity study on the read-across analogue substance produced any signs of dermal irritation that would warrant classification under Regulation (EC) No 1272/2008 (CLP).

Neither the in vitro eye irritation study nor the in vivo eye irritation study of EC 903-162-9 produced any signs of eye irritation that would warrant classification under Regulation (EC) No 1272/2008 (CLP).