Short Chain Fructo Oligosaccharides

Skin irritation / corrosion

Administrative data

Endpoint:

skin corrosion: in vitro / ex vivo

Type of information:

experimental study

Adequacy of study:

key study

Study period:

June 2011

Reliability:

1 (reliable without restriction)

Rationale for reliability incl. deficiencies:

guideline study

Data source

Reference

Reference Type:

study report

Title:

Unnamed

Year:

2011

Report date:

2011

Materials and methods

Test guideline

Test guideline 1

Qualifier:

according to guideline

Guideline:

OECD Guideline 431 (In Vitro Skin Corrosion: Human Skin Model Test)

Deviations:

no

Test guideline 2

Qualifier:

according to guideline

Guideline:

other: EU method B.40bis

Deviations:

no

GLP compliance:

yes (incl. QA statement)

Test material

Test material information

Constituent 1

Reference substance name:

short chain Fructo-Oligosaccharides

EC Number:

908-300-1

Molecular formula:

C6H11O5(C6H10O5)nOH

IUPAC Name:

short chain Fructo-Oligosaccharides

Test material form:

solid: particulate/powder

Details on test material:

Identification: Actilight 950P
Substance: Short-chain fructo-oligosaccharides
EC No.: 908-300-1
Appearance: White powder
Purity: 96%

COMPOSITION:
scFOS is short chains of fructose molecules linked to a molecule of sucrose (glucose-fructose disaccharide). Thus, scFOS is a multiconstituent substance composed of three oligosaccharides: 1-kestose (GF2), nystose (GF3) and fructosyl-nystose (GF4).
With:
- TOTAL FOS: 96% expressed as dry matter
- Kestose (GF2): 37.1% FOS
- Nystose (GF3): 50.5% FOS
- 1F-Fructofuranosylnystose (GF4:) 12.4% FOS
- Glucose +Fructose + Saccharose : 4.0% expressed as dry matter

Specific details on test material used for the study:

• Sponsor’s identification: ACTILIGHT® 950P
• Form: powder
• Colour: White
• Storage : room temperature

In vitro test system

Test system:

human skin model

Source species:

human

Vehicle:

unchanged (no vehicle)

Details on test system:

RECONSTRUCTED HUMAN EPIDERMIS (RHE) TISSUE
- Model used: Epidermal Skin Test EST-1000, Cell Systems
- Tissue batch number(s): - Batch No. EST- 110502-001)

The insert (filter + epidermis) was gently removed from the agarose while avoiding leaving agarose on the polycarbonate filter. The insert was placed in a 6 wells culture plate which has been previously filled with 1 mL of maintenance medium (Cell Systems MM-160511). The culture dishes were incubated at 37°C, 5% CO2 during 20 hours and 25 minutes before treatment.

TEMPERATURE USED FOR TEST SYSTEM
- Temperature used during treatment / exposure: room température
- Temperature of post-treatment incubation (if applicable):

REMOVAL OF TEST MATERIAL AND CONTROLS
3 minutes and 1 hour after the test item application, the human epidermis was washed 20 times with 1 mL of PBS (PAN BIOTECH GmbH, Batch No. 4721010).

MTT DYE USED TO MEASURE TISSUE VIABILITY AFTER TREATMENT / EXPOSURE
- MTT concentration / Incubation time: The skin sample was placed in MTT solution of 1 mg/mL concentration for 3 hours at 37°C, 5% CO2.
The precipitated blue formazan product is then extracted using isopropanol during 2 hours under agitation in the dark, and the concentration of formazan was measured by determining the Optical Density (OD) at 570 nm, just after dilution of the extraction in isopropanol (1:3).
The absorbance was measured in triplicate of MTT extract.
The measured absorbances are proportional to the number of live cells.

The direct interaction of MTT with the test item was checked by adding 25 mg of ACTILIGHT® 950P to 0.3 mL of the solution of MTT at 1 mg/mL. No blue or purple coloration was noted after 3 hours of incubation at 37°C, 5% CO2. Therefore, there is no direct interaction between the test item and MTT.

PREDICTION MODEL / DECISION CRITERIA:
- The test substance is considered to be corrosive to skin if the viability after 3 minutes exposure is striçtly less than 50%, or if the viability after 3 minutes exposure is greater than or equal to 50 % and the viability after 1 hour exposure is less than 15%.]
- The test substance is considered to be non-corrosive to skin if the viability after 3 minutes exposure is greater than or equal to 50% and the viability after 1 hour exposure is greater than or equal to 15%.]

Control samples:

yes, concurrent negative control

yes, concurrent positive control

Amount/concentration applied:

TEST MATERIAL:
The test item has been applied, as supplied, at the dose of 25 mg, to the epidermal surface of 2 human skin models, then 25 µL pf PBS was added.


NEGATIVE CONTROL
In the same experimental conditions, a negative control (PBS ) has been carried out.

POSITIVE CONTROL
In the same experimental conditions, a positive control (8N KOH) has been carried out.

Duration of treatment / exposure:

The test item remained on the epidermis during 3 minutes and during 1 hour at room température

Results and discussion

In vitro

Results

Results 1

Irritation / corrosion parameter:

% tissue viability

Run / experiment:

3 min

Value:

115.6

Vehicle controls validity:

not applicable

Negative controls validity:

valid

Positive controls validity:

valid

Results 2

Irritation / corrosion parameter:

% tissue viability

Run / experiment:

1hour

Value:

97.3

Vehicle controls validity:

not applicable

Negative controls validity:

valid

Positive controls validity:

valid

Other effects / acceptance of results:

- Direct-MTT reduction:
The direct interaction of MTT with the test item was checked by adding 25 mg of ACTILIGHT® 950P to 0.3 mL of the solution of MTT at 1 mg/mL. No blue or purple coloration was noted after 3 hours of incubation at 37°C, 5% CO2.
Therefore, there is no direct interaction between the test item and MTT

Applicant's summary and conclusion

Interpretation of results:

GHS criteria not met

Conclusions:

under the conditions of the test, 3 minutes and 1 hour after the test item application, the mean viability of the epidermis skins treated with the test item was 115.6% (considered as 100%) and 97.3%, versus 2.7% and 1.0%, respectively, in the positive control (potassium hydroxide 8N). The substance is considered as not corrosive to the skin in accordance with the Regulation EC No. 1272/2008, the test item must not be classified in category 1 “Corrosive”.

Executive summary:

The aim of the study was to evaluate the possible corrosive effects of the test item after topical administration on in vitro human reconstituted epidermis (Epidermal Skin Test EST-1000, Cell Systems). The experimental protocol was established in accordance with the OECDguideline No 431 dated April 13th, 2004 and the method B.40bis of the Council regulation No. 440/2008.

The test item ACTILIGHT® 950P was applied, as supplied, at the dose of 25 mg, to 2 Human skin model surfaces (Epidermal Skin Test EST-1000, Cell Systems), then 25 µL of PBS was added to ensure a good contact with the epidermis during 3 minutes and 1 hour.

 

3 minutes and 1 hour after the test item application, the mean viability of the epidermis skins treated with the test item was 115.6% (considered as 100%) and 97.3%, versus 2.7% and 1.0%, respectively, in the positive control (potassium hydroxide 8N).

 

The results obtained, under these experimental conditions, enable to conclude that the test item must not be classified in category 1 “Corrosive” in accordance with the Regulation EC No. 1272/2008. The substance is considered a not corrosive to the skin.

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