Reaction mass of Phenol, dodecyl-, sulfurized, calcium salts and benzene and butene and calcium carbonate and calcium dihydroxide and sulphur trioxide

Endpoint summary

Description of key information

Dermal Irritation- No irritation or any other effects
Eye Irritation- No irritation or any other effects

Key value for chemical safety assessment

Skin irritation / corrosion

Link to relevant study records

Reference

Endpoint:

skin corrosion: in vitro / ex vivo

Type of information:

experimental study

Adequacy of study:

key study

Study period:

20 January 2012 to 04 June 2012

Reliability:

1 (reliable without restriction)

Rationale for reliability incl. deficiencies:

guideline study

Qualifier:

according to guideline

Guideline:

OECD Guideline 439 (In Vitro Skin Irritation: Reconstructed Human Epidermis Test Method)

Deviations:

no

GLP compliance:

yes (incl. QA statement)

Test system:

human skin model

Type of coverage:

open

Vehicle:

unchanged (no vehicle)

Amount / concentration applied:

10 µL

Duration of treatment / exposure:

15 minutes

Details on study design:

TEST SYSTEM
- SkinEthic EpiSkin

TEST SITE
- Area of exposure: 0.38 cm2

REMOVAL OF TEST SUBSTANCE
- Washing (if done): washed with >25 mL PBS and tissue paper
- Time after start of exposure: 15 minutes

Irritation / corrosion parameter:

% tissue viability

Run / experiment:

Rep 3, mean of 2 runs

Value:

106.51

Vehicle controls validity:

valid

Negative controls validity:

valid

Positive controls validity:

valid

Remarks on result:

no indication of irritation

Irritation / corrosion parameter:

% tissue viability

Run / experiment:

Rep 2, mean of 2 runs

Value:

100.29

Vehicle controls validity:

valid

Negative controls validity:

valid

Positive controls validity:

valid

Remarks on result:

no indication of irritation

Irritation / corrosion parameter:

% tissue viability

Run / experiment:

mean of 3 Reps, positive control

Value:

16.79

Vehicle controls validity:

not applicable

Negative controls validity:

not applicable

Positive controls validity:

not applicable

Remarks on result:

positive indication of irritation

Irritation / corrosion parameter:

% tissue viability

Run / experiment:

mean of 3 reps, negative control

Value:

100

Vehicle controls validity:

not applicable

Negative controls validity:

not applicable

Positive controls validity:

not applicable

Remarks on result:

no indication of irritation

Other effects / acceptance of results:

- OTHER EFFECTS:
- Direct-MTT reduction: EC-903-161-3 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 co
rrected accordingly.
DEMONSTRATION OF TECHNICAL PROFICIENCY:
ACCEPTANCE OF RESULTS:
- Acceptance criteria met for negative control: met
- Acceptance criteria met for positive control: met
- Acceptance criteria met for variability between replicate measurements:
Three replicates of the positive control, aqueous 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%).
The assay was deemed acceptable if the following criteria were met:
The non-specific MTT reduction was =30% relative to the negative control optical density.
The mean OD value of the 3 negative control tissues was >0.6 and <1.5 and the standard deviation
value (SD) of the % viability was <18%.
The mean % viability of the 3 positive control tissues was <30% and the SD was <18%.
The mean % viability SD of the 3 treated tissues was <18%

Non-Viable Controls
The mean absorbance value of the three replicate non-viable tissues dosed with EC-903-161-3 was
0.184 ± 0.057. The mean absorbance value of the three replicate undosed non-viable tissues was 0.125
± 0.014. Therefore, the absorbance value for the effect of the unremoved test item was 0.060. This
value was subtracted from the absorbance value of the viable tissues dosed with EC-903-161-3.
Viable Tissues
The absorbance data for one of the tissues dosed with EC-903-161-3 (tissue Replicate 1) were rejected,
due to abnormally high values. Therefore, the mean viability was calculated from the results of the two
accepted tissues. Exposure to EC-903-161-3 resulted in a mean EpiSkin® viability (corrected for the
effect of unremoved test item) of 103.40% of the negative control value.
The standard deviation could not be calculated and the acceptability of the results could not be assessed
on the basis of the acceptance criteria described in Section 7.8 of the report. The results have been
accepted on the following basis:
• The results of the two accepted tissues were similar. The mean viability of the tissues differed by ca 6%.
• The rejection of the results of tissue Replicate 1 represents a “worst case” scenario (i.e. rejection
reduces the mean absorbance value and therefore the mean tissue viability).
Negative Controls
The negative control results were similar for the three viable EpiSkin® tissues dosed with Dulbecco’s
PBS. Exposure to Dulbecco’s PBS resulted in a mean EpiSkin® viability of 100.00% ± 3.57%

Positive Controls
The positive control results were similar for the three viable EpiSkin® tissues dosed with aqueous SDS
solution (5%, w/v). Exposure to aqueous SDS solution (5%, w/v) resulted in a mean EpiSkin® viability
of 16.79% ± 6.15%

Interpretation of results:

GHS criteria not met

Conclusions:

In conclusion, EC-903-161-3 was demonstrated to be non-irritant (no category) when tested in the EpiSkin® in vitro irritation assay.

Executive summary:

In this study, the irritation potential of EC-903-161-3 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.
EC-903-161-3 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 (10 µL) of the EC-903-161-3 to the
exposed surface of three EpiSkin tissues 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 tissues using Dulbecco’s phosphate-buffered saline (PBS)
and tissue swabs. The EpiSkin tissues were then incubated for a recovery period of 42 h ± 1 h in a
humidified incubator set to maintain temperature and CO2 levels of 37°C and 5%, respectively. Following
incubation, the EpiSkin tissues 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 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-903-161-3 resulted in a mean EpiSkin viability of 103.40%
of the negative control value. Exposure to the positive control, aqueous sodium dodecyl sulphate (SDS)
solution (5%, w/v), resulted in a mean EpiSkin viability of 16.79% ± 6.15% of the negative control value.
In conclusion, EC-903-161-3 was demonstrated to be non-irritant (no category) when tested in the
EpiSkin in vitro irritation assay.

Endpoint conclusion

Endpoint conclusion:

no adverse effect observed (not irritating)

Eye irritation

Endpoint conclusion

Endpoint conclusion:

no adverse effect observed (not irritating)

Respiratory irritation

Endpoint conclusion

Endpoint conclusion:

no study available

Additional information

Dermal Irritation -A study performed on a Registration substance in accordance with the OECD 439 guideline & the ECVAM guidelines, reported no irritancy or any other effects.

 

Eye Irritation -An in vitro eye irritation study (the HCE in vitro ocular irritation test) that

has undergone ECVAM pre-validation testing was performed on a test material and reported no irritancy or any other effects.

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

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

Justification for classification or non-classification

Dermal Irritation-A study performed on a test material in accordance with the OECD 439 guideline & the ECVAM guidelines, reported no irritancy or any other effects. No classification is therefore warranted under CLP Regulation (EC) 1272/2008.

 

Eye Irritation- An in vitro skin irritation study (SkinEthic) that has undergone ECVAM pre-validation testing was performed on a test material and reported no irritancy or any other effects. No classification is therefore warranted under CLP Regulation (EC) 1272/2008.