2-amino-2-(hydroxymethyl)propane-1,3-diol hydrochloride

Skin irritation / corrosion

Administrative data

Endpoint:

skin irritation: in vitro / ex vivo

Type of information:

experimental study

Adequacy of study:

key study

Study period:

November 1, 2016 - December 20, 2016

Reliability:

1 (reliable without restriction)

Rationale for reliability incl. deficiencies:

guideline study

Data source

Reference

Reference Type:

study report

Title:

Unnamed

Year:

2016

Report date:

2016

Materials and methods

Test guideline

Test guideline 1

Qualifier:

according to guideline

Guideline:

EU Method B.46 (In Vitro Skin Irritation: Reconstructed Human Epidermis Model Test)

Test guideline 2

Qualifier:

according to guideline

Guideline:

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

Test guideline 3

Qualifier:

according to guideline

Guideline:

other: Skinethic skin irritation test -42bis Standard operating procedure (SOP) 2009

GLP compliance:

yes (incl. QA statement)

Test material

Test material information

Constituent 1

Reference substance name:

2-amino-2-(hydroxymethyl)propane-1,3-diol hydrochloride

EC Number:

214-684-5

EC Name:

2-amino-2-(hydroxymethyl)propane-1,3-diol hydrochloride

Cas Number:

1185-53-1

Molecular formula:

C4H11NO3.ClH

IUPAC Name:

2-amino-2-(hydroxymethyl)propane-1,3-diol hydrochloride

Test material form:

solid

Specific details on test material used for the study:

Designation: Art. 108219
Synonym: Tris(hydroxymethyl)-aminomethananhydrochlorid
Trade name: TRIS-HCL Emprove Bio Bulk
CAS-No.: 1185-53-1
Batch: TD16007119
Purity: 100.2% (Titration with perchloric acid)
Appearance: Colourless crystals
Expiry date: June 30, 2021
Storage: +15 to +25°C, store in tightly closed containers, protected from moisture

In vitro test system

Test system:

human skin model

Source species:

human

Cell type:

non-transformed keratinocytes

Cell source:

foreskin from a single donor

Justification for test system used:

standard model

Vehicle:

unchanged (no vehicle)

Remarks:

No vehicle used in this study; The test item was applied neat to the tissues.

Details on test system:

CELL CULTURE
- Supplier: EpiSkin/SkinEthic Laboratories, Lyon, France)
- Source: human keratinocytes cultured on a polycarbonate filter in conditions which permit their terminal differentiation
- Format: 24 well plate
- Batch: 16-RHE-122
- Expires: November 28, 2016

TEMPERATURE USED FOR TEST SYSTEM
- Temperature used during treatment: room temperature
- Temperature of post-treatment incubation: 37°C

REMOVAL OF THE TEST MATERIAL AND CONTROL
After the end of the treatment interval, the residual test item was removed immediately by gently rinsing with a minimum volume of 25 mL DPBS using a pipette. Excess DPBS was removed by gently shaking the inserts and blotting the bottom with blotting paper.

Control samples:

yes, concurrent negative control

Amount/concentration applied:

TEST MATERIAL: 16 mg of solid test material
NEGATIVE CONTROL: 16 µL (Dulbecco`s Phosphate-Buffered Saline)
POSITIVE CONTROL: 16 µL (5% aqueous solution of sodium dodecyl sulfate in deionised water)

Duration of treatment / exposure:

42 min (± 1 minute)

Duration of post-treatment incubation (if applicable):

42 hours (± 1 hour)

Number of replicates:

3

Results and discussion

In vitro

Results

Irritation / corrosion parameter:

% tissue viability

Run / experiment:

Experiment 1 / Run 1

Value:

85.95

Vehicle controls validity:

not applicable

Remarks:

The test item was applied neat to the tissues

Negative controls validity:

valid

Positive controls validity:

valid

Remarks on result:

no indication of irritation

Other effects / acceptance of results:

OTHER EFFECTS:
- Visible damage on test system: none
- Direct-MTT reduction: none
- Colour interference with MTT: none

ACCEPTANCE OF RESULTS:
Acceptability of the Quality Control Data of the Skin Model with Reference to Historical Batch Data:
The negative control OD values were 2.450, 2.192 and 2.313 and, thus, in the range of ≥0.8 and ≤3.0.

Acceptability of the Positive and Negative Control:
After treatment with the positive control (5% aqueous solution of sodium dodecyl sulfate) the mean viability value was 1.13% (standard deviation: 2.95%) and, thus, lower than the historically established threshold of 3.12%.
After treatment with the negative control (DPBS-buffer) the mean OD was 2.318 (standard deviation: 5.57%) and, thus, higher than the historically established threshold of 1.436.

Variability of the Data:
The standard deviation between the three tissues replicates treated with the test item was 11.38% and, thus, ≤18%.
The standard deviations between the three tissue replicates of the negative control and the positive control were 5.57% and 2.95%, respectively, and, thus, ≤18%.

The study met all acceptance criteria.

Any other information on results incl. tables

Group	Time / [min]	Mean OD	Mean Relative viability / [%]
Negative Control	42	2.318	100
Positive Control	42	0.026	1.13
Test Material	42	1.993	85.95

Applicant's summary and conclusion

Interpretation of results:

GHS criteria not met

Conclusions:

This study was performed according to GLP and the methods applied are fully compliant with OECD TG 439. Under the conditions of the present study, the test item TRIS-HCL is not considered to possess an irritant potential to skin (UN GHS: No Category).

Executive summary:

This study was performed according to GLP and the methods applied are fully compliant with OECD TG 439. Under the conditions of the present study, the test item TRIS-HCl is not considered to possess an irritant potential to skin.