Benzenesulfonic acid, 4-C10-13-sec-alkyl derivs., ammonium salts

Skin irritation / corrosion

Administrative data

Endpoint:

skin corrosion: in vitro / ex vivo

Type of information:

experimental study

Adequacy of study:

key study

Study period:

27 March 2017 to 06 September 2017

Reliability:

1 (reliable without restriction)

Rationale for reliability incl. deficiencies:

guideline study

Remarks:

Study was conducted in accordance with international guidelines and in accordance with GLP. All guideline validity criteria were met.

Data source

Reference

Reference Type:

study report

Title:

Unnamed

Year:

2017

Materials and methods

Test guideline

Test guideline 1

Qualifier:

according to guideline

Guideline:

OECD Guideline 431 (In Vitro Skin Corrosion: Reconstructed Human Epidermis (RHE) Test Method)

Deviations:

no

Test guideline 2

Qualifier:

according to guideline

Guideline:

EU Method B.40 (In Vitro Skin Corrosion: Transcutaneous Electrical Resistance Test (TER))

Deviations:

no

GLP compliance:

yes (incl. QA statement)

Test material

Test material information

Constituent 1

Reference substance name:

Benzenesulfonic acid, 4-sec-decyl, ammonium salts

Molecular formula:

C16H29NO3S

IUPAC Name:

Benzenesulfonic acid, 4-sec-decyl, ammonium salts

Constituent 2

Reference substance name:

Benzenesulfonic acid, 4-sec-undecyl, ammonium salts

Molecular formula:

C17H31NO3S

IUPAC Name:

Benzenesulfonic acid, 4-sec-undecyl, ammonium salts

Constituent 3

Reference substance name:

Benzenesulfonic acid, 4-sec-dodecyl, ammonium salts

Molecular formula:

C18H33NO3S

IUPAC Name:

Benzenesulfonic acid, 4-sec-dodecyl, ammonium salts

Constituent 4

Reference substance name:

Benzenesulfonic acid, 4-sec-tridecyl, ammonium salts

Molecular formula:

C19H35NO3S

IUPAC Name:

Benzenesulfonic acid, 4-sec-tridecyl, ammonium salts

Test material form:

other: Highly viscous light yellow liquid

Details on test material:

Storage Conditions: Highly viscous light yellow liquid

Specific details on test material used for the study:

RADIOLABELLING INFORMATION (if applicable)
- Radiochemical purity: n/a
- Specific activity: n/a
- Locations of the label: n/a
- Expiration date of radiochemical substance: n/a

STABILITY AND STORAGE CONDITIONS OF TEST MATERIAL
- Storage condition of test material: Room temperature
- Stability under test conditions: Assumed stable
- Solubility and stability of the test substance in the solvent/vehicle: N/A - test item applied uniluted
- Reactivity of the test substance with the solvent/vehicle of the cell culture medium: No

TREATMENT OF TEST MATERIAL PRIOR TO TESTING
- Treatment of test material prior to testing: No, applied as supplied.
- Preliminary purification step (if any): n/a
- Final dilution of a dissolved solid, stock liquid or gel: n/a
- Final preparation of a solid: n/a

FORM AS APPLIED IN THE TEST (if different from that of starting material): n/a

OTHER SPECIFICS: n/a

In vitro test system

Test system:

human skin model

Source species:

human

Cell type:

non-transformed keratinocytes

Cell source:

other: Not reported

Source strain:

other: n/a

Details on animal used as source of test system:

n/a

Vehicle:

unchanged (no vehicle)

Details on test system:

RECONSTRUCTED HUMAN EPIDERMIS (RHE) TISSUE
- Model used: EpiDerm Reconstructed Human Epidermis
- Tissue batch number(s): 00267; EpiDerm kit lot # = 25802
- Production date: not reported
- Shipping date: not reported
- Delivery date: not reported
- Date of initiation of testing: 29 March 2017

TEMPERATURE USED FOR TEST SYSTEM
- Temperature used during treatment / exposure: 37 ± 1 ºC
- Temperature of post-treatment incubation (if applicable): 37 ºC

REMOVAL OF TEST MATERIAL AND CONTROLS
-Volume and number of washing steps: Rinsed with PBS (volume and number of rinsings not reported)
- Observable damage in the tissue due to washing: No
- Modifications to validated SOP: no

MTT DYE USED TO MEASURE TISSUE VIABILITY AFTER TREATMENT / EXPOSURE
- MTT concentration: 1 mg/mL
- Incubation time: 3 hours
- Spectrophotometer: not reported
- Wavelength: 570 nm
- Filter: Not reported
- Filter bandwidth: Not reported
- Linear OD range of spectrophotometer: Not reported

FUNCTIONAL MODEL CONDITIONS WITH REFERENCE TO HISTORICAL DATA
- Viability: acceptable (provided in CoA)
- Barrier function: acceptable (provided in CoA)
- Morphology: acceptable (provided in CoA)
- Contamination: acceptable (provided in CoA)
- Reproducibility: The standard deviation (SD) for tissues should be ≤18 %.

NUMBER OF REPLICATE TISSUES: 3

CONTROL TISSUES USED IN CASE OF MTT DIRECT INTERFERENCE
- n/a

NUMBER OF INDEPENDENT TEST SEQUENCES / EXPERIMENTS TO DERIVE FINAL PREDICTION: 1

PREDICTION MODEL / DECISION CRITERIA (choose relevant statement)
- The test substance is considered to be corrosive to skin if the viability after 3 min exposure is less than 50 %, relative to the negative control.
- The test substance is considered to be corrosive to skin if the viability after exposure is greater than or equal to 50 % after 3 min exposre and < 15 % after 60 min exposure.
- The test substance is considered to be non-corrosive to skin if the viability after exposure is greater than or equal to 50 % after 3 min exposre and ≥ 15 % after 60 min exposure.
- Justification for the selection of the cut-off point(s) if different than recommended in TG 431 and 439: n/a

Control samples:

yes, concurrent negative control

yes, concurrent positive control

Amount/concentration applied:

TEST MATERIAL
- Amount(s) applied (volume or weight with unit): 50 µL
- Concentration (if solution): n/a

VEHICLE
- Amount(s) applied (volume or weight with unit): n/a
- Concentration (if solution): n/a
- Lot/batch no. (if required): n/a
- Purity: n/a

NEGATIVE CONTROL
- Amount(s) applied (volume or weight): 50 µL
- Concentration (if solution): n/a

POSITIVE CONTROL
- Amount(s) applied (volume or weight): 50 µL
- Concentration (if solution): 8 N

Duration of treatment / exposure:

3 minutes and 60 minutes

Duration of post-treatment incubation (if applicable):

Incubated in MTT medium for 3 hours and followed by an overnight extraction in isopropanol.

Number of replicates:

2 for each timepoint

Test animals

Species:

other: n/a

Strain:

other: n/a

Details on test animals or test system and environmental conditions:

n/a

Test system

Type of coverage:

other: n/a

Preparation of test site:

other: n/a

Vehicle:

other: n/a

Amount / concentration applied:

n/a

Duration of treatment / exposure:

n/a

Observation period:

n/a

Number of animals:

n/a

Details on study design:

n/a

Results and discussion

In vitro

Results

Results 1

Irritation / corrosion parameter:

% tissue viability

Run / experiment:

3 minute

Value:

61

Vehicle controls validity:

not examined

Negative controls validity:

valid

Remarks:

Viability = 100 % (OD 1.503)

Positive controls validity:

valid

Remarks:

Viability = 3 % (OD 0.320)

Remarks on result:

no indication of irritation

Results 2

Irritation / corrosion parameter:

% tissue viability

Run / experiment:

60 minute

Value:

64.2

Vehicle controls validity:

not examined

Negative controls validity:

valid

Remarks:

Viability = 100 % (OD 1.086)

Positive controls validity:

valid

Remarks:

Viability = -0.4 % (OD 0.121)

Remarks on result:

no indication of irritation

Other effects / acceptance of results:

- OTHER EFFECTS:
- Visible damage on test system: No
- Direct-MTT reduction: No
- Colour interference with MTT: No

DEMONSTRATION OF TECHNICAL PROFICIENCY: Yes

ACCEPTANCE OF RESULTS:
- Acceptance criteria met for negative control: Yes
- Acceptance criteria met for positive control: Yes
- Acceptance criteria met for variability between replicate measurements: Yes
- Range of historical values if different from the ones specified in the test guideline: n/a

Any other information on results incl. tables

Table 2       Mean OD₅₇₀Values and Viabilities for the Negative Control Item, Positive Control Item and Test Item

 

Tissue	Exposure Period	Mean OD570 of individual tissues	Mean OD570 of duplicate tissues	Standard Deviation	Coefficient of Variation (%)	Relative Mean Viability (%)
Negative Control	3 Minutes	1.744	1.503	0.266	17.7	100
		1.262
	60 Minutes	0.999	1.086	0.097	8.9
		1.173
Positive Control	3 Minutes	0.328	0.320	0.049	13.3	3
		0.311
	60 Minutes	0.113	0.121	0.014	12.3	-0.4
		0.128
Test Item	3 Minutes	0.788	0.916	0.142	15.5	61
		1.044
	60 Minutes	0.697	0.698	0.030	4.3	64.2
		0.699

 OD = Optical density

*=  The mean % viability of the negative control tissue is set at 100 %

nd= Not determined

Positive control values determined from freeze killed cells

 

Applicant's summary and conclusion

Interpretation of results:

GHS criteria not met

Conclusions:

Under the conditions of this study the test substance is not considered to be corrosive in the in vitro skin corrosion test using the EpiDerm Skin Model.

Executive summary:

OECD 431 (2017) - The skin corrosivity potential of reaction mass of Benzenesulfonic acid, 4 -C10 -13 -sec-alkyl derivs., ammonium salts was assessed using an EpiDerm Reconstructed Human Epidermis (RHE) Model Kit in accordance with OECD guidance 431 and GLP.

Duplicate tissues were treated with the test item for exposure periods of 3 and 60 mins. At the end of the exposure period the test item was rinsed from each tissue before being loaded with MTT. After MTT loading each tissue was placed in 2 mL isopropanol for MTT extraction. After extraction, each tissue was pierced and the extraction solution aliquoted for absorbance measurements. Absorbency at 570 nm of each well was measured using a spectrophotometer.

Mean viability of tissues exposed to the test substance after 3 and 60 minutes were 61 % and 64.2 %, respectively. The quality criteria required for acceptance of the resuts was met.

Under the conditions of this study the test substance is not considered to be corrosive to the skin.