1,4-α-glucan branching enzyme

Endpoint summary

Key value for chemical safety assessment

Skin irritation / corrosion

Link to relevant study records

Reference

Reference 1

Endpoint:

skin irritation: in vivo

Type of information:

experimental study

Adequacy of study:

key study

Study period:

07-09-2007 to 08-11-2007

Reliability:

1 (reliable without restriction)

Rationale for reliability incl. deficiencies:

guideline study

Qualifier:

according to guideline

Guideline:

OECD Guideline 404 (Acute Dermal Irritation / Corrosion)

Version / remarks:

Adopted on 24 April 2002

Deviations:

no

GLP compliance:

yes (incl. QA statement)

Species:

rabbit

Strain:

New Zealand White

Type of coverage:

semiocclusive

Preparation of test site:

other: Not necessary since they are reconstructed skin membranes

Remarks:

SPF bred New Zealand White albino rabbits.

Vehicle:

unchanged (no vehicle)

Controls:

yes, concurrent positive control

yes, concurrent negative control

Amount / concentration applied:

TEST MATERIAL
- Amount(s) applied (volume or weight with unit): 0.5 mL
- Concentration (if solution): used as is

Duration of treatment / exposure:

4 hours.

Observation period:

1, 24, 48, 72 hours and at 7 and 14 days after treatment.

Details on study design:

TEST SITE
- Area of exposure: 2.5 x 2.5 cm
- % coverage: 100%
- Type of wrap if used: The test substance was distributed over a cotton gauze patch. The patch loaded with the test substance was fixed to the selected application site by means of semi-occlusive, adhesive tape. Subsequently, the entire trunk of the rabbit was wrapped with a self-adhesive-gauze to maintain the patch in position and to retard evaporation of volatile substances.

REMOVAL OF TEST SUBSTANCE
- Washing (if done): The test site was cleaned with a tissue moistened with water.
- Time after start of exposure: Exposure time 4 hours. Observation period post-exposure: 1, 24, 48, 72 hours and at 7 and 14 days.

SCORING SYSTEM:
- Method of calculation: The resulting skin reactions were evaluated by the method of Draize et al. (J. Pharrnacol. Exp. Ther. 82 (1944) 377-390.

Irritation parameter:

erythema score

Basis:

animal #1

Time point:

24/48/72 h

Score:

0.66

Max. score:

1

Reversibility:

fully reversible within: 72 hours

Remarks on result:

no indication of irritation

Irritation parameter:

erythema score

Basis:

animal #3

Time point:

24/48/72 h

Score:

0.66

Max. score:

1

Reversibility:

fully reversible within: 72 hours

Remarks on result:

no indication of irritation

Irritation parameter:

erythema score

Basis:

animal #5

Time point:

24/48/72 h

Score:

0.66

Max. score:

1

Reversibility:

fully reversible within: 72 hours

Remarks on result:

no indication of irritation

Irritation parameter:

edema score

Basis:

animal #1

Time point:

24/48/72 h

Score:

0

Max. score:

1

Reversibility:

fully reversible within: 24 hours

Remarks on result:

no indication of irritation

Irritation parameter:

edema score

Basis:

animal #3

Time point:

24/48/72 h

Score:

0

Max. score:

1

Reversibility:

fully reversible within: 24 hours

Remarks on result:

no indication of irritation

Irritation parameter:

edema score

Basis:

animal #5

Time point:

24/48/72 h

Score:

0

Max. score:

1

Reversibility:

fully reversible within: 24 hours

Remarks on result:

no indication of irritation

Interpretation of results:

Category 2 (irritant) based on GHS criteria

Conclusions:

According to the EC-standards (as published in the Official Journal of the European Communities, L 110 A, Volume 36, 4 May 1993), 1,4-alpha-glucan branching enzyme, batch PPY27209 is not irritating to (human) skin. According to the Globally Harmonised System of Classification and Labelling of Chemicals (GHS; UN/ECE 2003) 1,4-alpha-glucan branching enzyme, batch PPY27209 is also not irritating to (human) skin.

Executive summary:

A sample of 1,4-alpha-glucan branching enzyme, batch PPY27209 was tested for acute dermal irritating properties in an experiment with three albino rabbits, according to EC Directive 92/69/EC, method B.4 and OECD Guideline no. 404. 1,4-alpha-glucan branching enzyme, batch PPY27209 caused very slight erythema and very slight oedema. At 24 and 48 hours after treatment, only very slight erythema was observed in the three rabbits. At 72 hours after removal, no signs of skin irritation were observed in any of the three rabbits.

According to the EC-standards (as published in the Official Journal of the European Communities, L 110 A, Volume 36, 4 May 1993), 1,4-alpha-glucan branching enzyme, batch PPY27209 is not irritating to (human) skin. According to the Globally Harmonised System of Classification and Labelling of Chemicals (GHS; UN/ECE 2003) 1,4-alpha-glucan branching enzyme, batch PPY27209 is also not irritating to (human) skin.

Reference 2

Endpoint:

skin irritation: in vitro / ex vivo

Remarks:

in vitro

Type of information:

experimental study

Adequacy of study:

key study

Study period:

09-06-2008 to 04-07-2008

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)

Version / remarks:

Not stated.

Deviations:

no

GLP compliance:

no

Test system:

human skin model

Remarks:

Episkin Standard Model (Episkin-SM)

Source species:

human

Cell type:

non-transformed keratinocytes

Cell source:

foreskin from multiple donors

Vehicle:

unchanged (no vehicle)

Details on test system:

SKIN PREPARATION
- Procedure used: The ECVAM Skin Irritation Validation Study protocol was followed. Episkin Standard Model (Episkin-SM), 12 tissue inserts of each 0.38cm^2, including maintainace medium provided by SkinEthic.

TEMPERATURE USED FOR TEST SYSTEM
- Temperature used during treatment / exposure: 37ºC and 5% CO2
- Temperature of post-treatment incubation (if applicable): Same

REMOVAL OF TEST MATERIAL AND CONTROLS
- Number of washing steps: each skin model was removed from the well, rinsed with 25 mL PBS to remove the study substance, and blotted dry.
- Observable damage in the tissue due to washing: None
- Modifications to validated SOP: None

DYE BINDING METHOD
- Dye used in the dye-binding assay: MTT
- Spectrophotometer: Yes, but not specified.
- Wavelength: 575 nm
- Filter: Not specified
- Filter bandwidth: Not specified

NUMBER OF INDEPENDENT TESTING RUNS / EXPERIMENTS TO DERIVE FINAL PREDICTION: 1

PREDICTION MODEL / DECISION CRITERIA
The acceptance criteria for a valid test;
* The mean OD value for the three NC tissue is >= 0.6 and the standard deviation value (SD) of the viability is<= 18
* The mean viability for the positive control (SDS treated) expressed as % of the NC, is below or 30% and the SD is <= 18

Amount/concentration applied:

TEST MATERIAL
- Amount(s) applied (volume or weight with unit): 10 μL, 26.3 μL/cm^2
- Concentration (if solution): used as is

Duration of treatment / exposure:

15 min.

Duration of post-treatment incubation (if applicable):

42 hours.

Number of replicates:

Triplicate tissues each for test substance, negative control Phosphate Buffered Saline and positive control 5% Sodium Dodecyl Sulphate.

Irritation / corrosion parameter:

% tissue viability

Run / experiment:

1

Value:

87.9

Vehicle controls validity:

not applicable

Negative controls validity:

valid

Positive controls validity:

valid

Remarks on result:

no indication of irritation

Other effects / acceptance of results:

- OTHER EFFECTS:
- Direct-MTT reduction: No reduction was observed.
- Colour interference with MTT: None observed.

The acceptance criteria for a valid test:
* The mean OD value for the three NC tissue is >= 0.6 and the standard deviation value (SD) of the viability is <= 18
* The mean viability for the positive control (SDS treated) expressed as % of the NC, is below or 30% and the SD is <= 18

Interpretation of results:

GHS criteria not met

Conclusions:

1,4-alpha-glucan branching enzyme, batch PPY27209 was examined for skin irritation in the Episkin in vitro assay for skin irritation and it was found to be a non-skin irritant.

Executive summary:

1,4-alpha-glucan branching enzyme, batch PPY27209 was examined for skin irritation in the Episkin in vitro assay for skin irritation. The ECVAM Skin Irritation Validation Study protocol was followed. The results of the present test were within the acceptance criteria for a valid test. 1,4-alpha-glucan branching enzyme, batch PPY27209 was found to be a non-skin irritant.

In conclusion, 1,4-alpha-glucan branching enzyme is not classified.

Endpoint conclusion

Endpoint conclusion:

no adverse effect observed (not irritating)

Eye irritation

Link to relevant study records

Reference

Endpoint:

eye irritation: in vivo

Remarks:

in vitro

Type of information:

experimental study

Adequacy of study:

key study

Study period:

07-09-2007 to 13-11-2007

Reliability:

1 (reliable without restriction)

Rationale for reliability incl. deficiencies:

guideline study

Qualifier:

according to guideline

Guideline:

OECD Guideline 405 (Acute Eye Irritation / Corrosion)

Version / remarks:

Adopted on 24 April 2002.

Deviations:

no

GLP compliance:

yes (incl. QA statement)

Species:

rabbit

Strain:

New Zealand White

Remarks:

SPF bred New Zealand White albino rabbits.

Details on test animals or tissues and environmental conditions:

TEST ANIMALS
- Source: Charles River, Germany
- Age at study initiation: Males, young adult
- Weight at study initiation: 1966, 1936 and 2106 g
- Housing: Individually in stainless steel cages with perforated floor.

Vehicle:

unchanged (no vehicle)

Controls:

not required

Amount / concentration applied:

TEST MATERIAL
- Amount(s) applied (volume or weight with unit): The chicken eye cornea was treated with 0.1 mL.
- Concentration (if solution): Undiluted test sample, 9.5% TOS.

Duration of treatment / exposure:

After administration, the upper and lower eye lid were carefully closed and subsequently held together for at least one second before releasing, to prevent loss of material.

Observation period (in vivo):

1, 24, 48, 72 hours

Details on study design:

REMOVAL OF TEST SUBSTANCE
- Washing (if done): None
- Time after start of exposure: few seconds

SCORING SYSTEM: The acute irritation of the test sample was classified according to the EC criteria for the labelling and classification of dangerous substances, as published in the Official Journal of the European Communities, L 110 A, Volume 36, 4 May 1993 and to the Globally Harmonised System of Classification and Labelling of Chemicals (GHS; UN/ECE 2003).

Irritation parameter:

cornea opacity score

Basis:

mean

Time point:

24/48/72 h

Score:

0

Remarks on result:

no indication of irritation

Irritation parameter:

iris score

Basis:

mean

Time point:

24/48/72 h

Score:

0

Remarks on result:

no indication of irritation

Irritation parameter:

conjunctivae score

Remarks:

redness

Basis:

mean

Time point:

24/48/72 h

Score:

0.3

Max. score:

1

Reversibility:

fully reversible within: 48 hours

Remarks on result:

no indication of irritation

Irritation parameter:

chemosis score

Remarks:

swelling

Basis:

mean

Time point:

24/48/72 h

Score:

0

Remarks on result:

no indication of irritation

Irritation parameter:

other: discharge

Basis:

mean

Time point:

24/48/72 h

Score:

0

Remarks on result:

no indication of irritation

Interpretation of results:

GHS criteria not met

Conclusions:

1,4-alpha-glucan branching enzyme, batch PPY27209 was tested for acute eye irritating properties in an experiment with three albino rabbits. At 48 h after treatment, the eye effects had cleared completely.
According to the EC-standards (as published in the Official Journal of the European Communities, L 110 A, Volume 36, 4 May 1993) 1,4-alpha-glucan branching enzyme, batch PPY27209 was not irritating to eyes. According to the Globally Harmonised System of Classification and Labelling of Chemicals (GHS; UN/ECE 2003) 1,4-alpha-glucan branching enzyme, batch PPY27209 was also not irritating to eyes.

Executive summary:

1,4-alpha-glucan branching enzyme, batch PPY27209 was tested for acute eye irritating properties in an experiment with three albino rabbits, according to EC Directive 92/69/EC, method B.5 and OECD Guideline no. 405.

The test item caused slight redness of the conjunctivae at the 1 hour and 24 hours observations. At 48 h after treatment, all eye effects had cleared completely.

According to the EC-standards (as published in the Official Journal of the European Communities, L 110 A, Volume 36, 4 May 1993), 1,4-alpha-glucan branching enzyme, batch PPY27209 was not irritating to (human) eyes.

According to the Globally Harmonised System of Classification and Labelling of Chemicals (GHS; UN/ECE 2003) 1,4-alpha-glucan branching enzyme, batch PPY27209 was also not irritating to (human) eyes.

Endpoint conclusion

Endpoint conclusion:

no adverse effect observed (not irritating)

Justification for classification or non-classification

Based on the results from the skin- and eye irritation tests,1,4-alpha-glucan branching enzyme is not classified