Registration Dossier

Administrative data

Endpoint:
skin irritation / corrosion, other
Type of information:
experimental study
Adequacy of study:
key study
Study period:
June-September 2018
Reliability:
1 (reliable without restriction)
Rationale for reliability incl. deficiencies:
guideline study

Data source

Reference
Reference Type:
study report
Title:
Unnamed
Year:
2018
Report Date:
2018

Materials and methods

Test guidelineopen allclose all
Qualifier:
according to
Guideline:
OECD Guideline 439 (In Vitro Skin Irritation: Reconstructed Human Epidermis Test Method)
Deviations:
no
Qualifier:
according to
Guideline:
EU Method B.46 (In Vitro Skin Irritation: Reconstructed Human Epidermis Model Test)
Deviations:
no
GLP compliance:
yes (incl. certificate)

Test material

Reference
Name:
Unnamed
Type:
Constituent
Test material form:
solid: particulate/powder
Details on test material:
Dry powder, white to slightly yellow
Specific details on test material used for the study:
Identification: Riboxyl TM
Appearance: White to slightly yellow powder

In vitro test system

Test system:
human skin model
Source species:
human
Cell type:
other: human-derived epidermal keratinocytes
Justification for test system used:
In the interest of sound science and animal welfare, a sequential testing strategy is recommended to minimize the need of in vivo testing. One of the validated in vitro skin irritation tests is the EPISKIN test, which is recommended in international guidelines (e.g.OECD and EC).
Vehicle:
unchanged (no vehicle)
Details on test system:
EPISKIN Small ModelTM (EPISKIN-SMTM, 0.38 cm2, Batch no.: 18-EKIN-027)
This model is a three-dimensional human epidermis model, which consists of adult human-derived epidermal keratinocytes which have been seeded on a dermal substitute consisting of a collagen type I matrix coated with type IV collagen. The keratinocytes were cultured for 13 days, which results in a highly differentiated and stratified epidermis model comprising the main basal, supra basal, spinous and granular layers and a functional stratum corneum.
Control samples:
yes, concurrent negative control
yes, concurrent positive control
Amount/concentration applied:
The skin was moistened with 5 μL Milli-Q water (Millipore Corp., Bedford, Mass., USA) to ensure close contact of the test item to the tissue and the solid test item (28.1 to 39.9 mg) was added into 12-well plates on top of the skin tissues.
Three tissues were treated, with 25 μL PBS (negative control) and 3 tissues with 25 μL 5% SDS (positive control) respectively.
Duration of treatment / exposure:
The positive control was re-spread after 7 minutes contact time.
After the exposure period of 15 ± 0.5 minutes at room temperature, the tissues were washed with phosphate buffered saline to remove residual test item.
Duration of post-treatment incubation (if applicable):
After rinsing, the cell culture inserts were each dried carefully and moved to a new well on 2 mL pre-warmed maintenance medium until all tissues were dosed and rinsed. Subsequently the skin tissues were incubated for 42 hours at 37°C.
Number of replicates:
The test was performed on a total of 3 tissues per test item together with negative and positive
controls.

Results and discussion

In vitro

Results
Irritation / corrosion parameter:
% tissue viability
Value:
ca. 111
Vehicle controls validity:
valid
Negative controls validity:
valid
Positive controls validity:
valid
Remarks on result:
no indication of irritation
Other effects / acceptance of results:
The positive control had a mean cell viability after 15 ± 0.5 minutes exposure of 7.8%. The absolute mean OD570 of the negative control tissues was within the laboratory historical control data range (See Appendix 3). The standard deviation value of the percentage viability of three tissues treated identically was < 10%, indicating that the test system functioned properly.

Any other information on results incl. tables

Table 2

Mean Tissue Viability in the In Vitro Skin Irritation Test with D-Ribose      

   Mean tissue viability (% control)  Standard deviation (%)
 Negative control  100  9.9
 D-Ribose  111  6.2
 Positive control  7.8  3.0

Table1. Mean absorption in the in vitro skin irritation test with D-Ribose

 OD570  A  B  C  Mean    SD
 Negative Control  1.293  1.234  1.065  1.198  +/-  0.118
 D-Ribose  1.417  1.278  1.301  1.332  +/-  0.074
 Positive Control  0.061  0.087  0.133  0.093  +/-

 0.036

Applicant's summary and conclusion

Interpretation of results:
GHS criteria not met
Conclusions:
D-Ribose is non-irritant in the in vitro skin irritation test under the experimental conditions described in this report and should not be classified according to the Globally Harmonized System of Classification and Labeling of Chemicals (GHS) of the United Nations.
Executive summary:

The objective of this study was to evaluate D-Ribose for its ability to induce skin irritation on a human three dimensional epidermal model (EPISKIN Small model (EPISKIN-SMTM)).

The possible skin irritation potential of

D-Ribose

was tested through topical application for 15 minutes.

The study procedures described in this report were based on the most recent OECD and EC guidelines.

Batch Q0926501 of D-Ribose was a white to slightly yellow powder. Skin tissue was moistened with 5 μL of Milli-Q water and at least 10 mg of D-Ribose was applied directly on top of the skin tissue for 15 ± 0.5 minutes. After a 42 hour post-incubation period, determination of the cytotoxic (irritancy) effect was performed. Cytotoxicity is expressed as the reduction of mitochondrial dehydrogenase activity measured by formazan production from MTT at the end of the treatment.

Skin irritation is expressed as the remaining cell viability after exposure to the test item. The relative mean tissue viability obtained after 15 ± 0.5 minutes treatment with D-Ribose compared to the negative control tissues was 111%. Since the mean relative tissue viability for D-Ribose was above 50% after 15 ± 0.5 minutes treatment Riboxyl TM is considered to be non-irritant.

The positive control had a mean cell viability of 7.8% after 15 ± 0.5 minutes exposure. The absolute mean OD570 (optical density at 570 nm) of the negative control tissues was within the laboratory historical control data range. The standard deviation value of the percentage viability of three tissues treated identically was < 10%, indicating that the test system functioned properly.

In conclusion, D-Ribose is non-irritant in the in vitro skin irritation test under the experimental conditions described in this report and should not be classified according to the Globally Harmonized System of Classification and Labeling of Chemicals (GHS) of the United Nations.