Syrups, wheat, hydrolyzed starch

Skin irritation / corrosion

Administrative data

Endpoint:

skin irritation / corrosion

Remarks:

in vitro

Type of information:

experimental study

Adequacy of study:

key study

Study period:

2010

Reliability:

1 (reliable without restriction)

Rationale for reliability incl. deficiencies:

other: Well documented, according to accepted guidelines

Data source

Reference

Reference Type:

study report

Title:

Unnamed

Year:

2010

Materials and methods

Test guideline

Test guideline 1

Qualifier:

according to guideline

Guideline:

other: OECD-Guideline 431, "In Vitro Skin Corrosion: Human Skin Model Test ", 13 April 2004.

Deviations:

yes

Remarks:

-reference to historical data of the model not reported.

Test guideline 2

Qualifier:

according to guideline

Guideline:

other: OECD Guideline 439, “In Vitro Skin Irritation: Reconstructed Human Epidermis (RhE) Test Method”, Paris 22 July 2010.

Deviations:

yes

Remarks:

-missing data on acceptability of the QC data with reference to historical batch data, reference to historical control data

Test guideline 3

Qualifier:

according to guideline

Guideline:

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

Version / remarks:

2008

Deviations:

yes

Remarks:

-reference to historical data of the model not reported.

Test guideline 4

Qualifier:

according to guideline

Guideline:

other: Regulation (EC) 761/2009: B.46 "In vitro skin irritation: Reconstructed Human Epidermis Model Test"

Deviations:

yes

Remarks:

- missing data on acceptability of the QC data with reference to historical batch data, reference to historical control data

GLP compliance:

yes

Test material

Test material information

Constituent 1

Reference substance name:

(3R,4S,5S,6R)-6-(hydroxymethyl)oxane-2,3,4,5-tetrol; (3S,4S,5R,6R)-6-(hydroxymethyl)-4-{[(2R,3R,4S,5S,6R)-3,4,5-trihydroxy-6-(hydroxymethyl)oxan-2-yl]oxy}oxane-2,3,5-triol

EC Number:

931-687-3

Molecular formula:

not applicable

IUPAC Name:

(3R,4S,5S,6R)-6-(hydroxymethyl)oxane-2,3,4,5-tetrol; (3S,4S,5R,6R)-6-(hydroxymethyl)-4-{[(2R,3R,4S,5S,6R)-3,4,5-trihydroxy-6-(hydroxymethyl)oxan-2-yl]oxy}oxane-2,3,5-triol

Details on test material:

- Name of test material (as cited in study report): GLUCOSE SYRUPS WHEAT HYDROLYZED, Dried wheat glucose syrup, Dried Glucose Syrup
- Physical state: Solid (white, slightly hygroscopic powder)
- Analytical purity: 38.2 (DE )
- Lot/batch No.: 5093187
- Expiration date of the lot/batch: June 2011
- Storage condition of test material: Room temperature, no protection from light
- Other: pH: neutral; Solubility: good soluble in water

Test animals

Species:

human

Strain:

other: not applicable (in vitro human skin model)

Details on test animals or test system and environmental conditions:

MatTek´s EpiDerm System consists of normal, human-derived epidermal keratinocytes which have been cultured to form a multilayered, highly differentiated model of the human epidermis. It consists of organized basal, spinous and granular layers, and a multi-layered stratum corneum containing intercellular lamellar lipid layers arranged in patterns analogous to those found in vivo. The EpiDerm™ tissues (surface 0.6 cm²) are cultured on specially prepared cell culture inserts (MILLICELLs®, 10 mm Ø) and shipped as kits, containing 24 tissues on shipping agarose.

Test system

Type of coverage:

other: not applicable (in vitro human skin model)

Preparation of test site:

other: not applicable (in vitro human skin model)

Vehicle:

water

Controls:

yes, concurrent vehicle

Amount / concentration applied:

TEST MATERIAL
- Amount(s) applied (volume or weight with unit): 25 mg; The "spoonful" was levelled by gently scratching the excess material away, avoiding compression. 25 μL H2O were added for wetting of the test substance.

Duration of treatment / exposure:

EpiDerm Skin Corrosivity Test: 3 minutes; 1 hour
EpiDerm Skin Irritation Test: 1 hour

Number of animals:

EpiDerm Skin Corrosivity Test: 2 tissue replicates
EpiDerm Skin Irritation Test: 3 tissue replicates

Details on study design:

1) 3-(4,5-Dimethylthiazol-2-yl)-2,5-diphenyltetrazolium bromide, Thiazolyl blue (MTT)-test
After incubation with the test substance, post-incubation (EpiDerm Skin Irritation Test) and washing with PBS, the tissues were incubated with MTT medium at 37°C and 5 % CO2. After 3 hours, the MTT medium was aspirated from all wells and the tissues were gently rinsed with phosphate buffered saline (PBS) (2 times). For extraction, the tissues were incubated with extractant solution (isopropanol) for 2 hours with shaking.
After the extraction period, the tissues were pierced with an injection needle and the extract (now a blue formazan solution) was allowed to run into the well from which the tissue was taken. The 24-well plates were placed on a shaker for 15 minutes until the solutions were homogeneous in colour.

2) Cell viability measurement
- EpiDerm Skin Corrosivity Test:
Per each tissue 3 × 200 μL aliquots of the blue formazan solution were transferred into a 96-well flat bottom microtiter plate and the optical density (OD) was measured using the extractant solution as blank in a plate spectrophotometer at 570 nm, without reference filter.
- EpiDerm Skin Irritation Test:
Per each tissue 2 × 200 μL aliquots of the blue formazan solution were transferred into a 96-well flat bottom microtiter plate and the OD was measured using the extractant solution as blank in a plate spectrophotometer at 570 nm, without reference filter.

3) Assay acceptance criteria
- EpiDerm Skin Corrosivity Test:
In the protocol INVITTOX n°119 by ECVAM the following assay acceptance criteria are defined:
• The mean OD 570 nm of the two negative control tissues is ≥ 0.8.
• The mean tissue viability of the 3 minutes positive control is ≤ 30 %.
• The inter tissue viability difference is < 30 % between two identically treated tissues.
- EpiDerm Skin Irritation Test:
• The mean OD 570 nm of the negative control tissues is ≥ 1.0 and ≤ 2.5.
• The mean tissue viability of the positive control is ≤ 20 %.
• The standard deviation calculated from individual percentual tissue viabilities of the 3 identically treated replicates is < 18 %.

Results and discussion

In vivo

Results

Results 1

Irritation parameter:

other: Skin Corrosion tissue viability (%)

Basis:

mean

Time point:

other: 3 min

Score:

97.8

Max. score:

100

Reversibility:

no data

Remarks on result:

other: Non-corrosive

Results 2

Irritation parameter:

other: Skin Corrosion tissue viability (%)

Basis:

mean

Time point:

other: 1 h

Score:

98.2

Max. score:

100

Reversibility:

no data

Remarks on result:

other: Non-corrosive

Results 3

Irritation parameter:

other: Skin irritation tissue viability (%)

Basis:

mean

Time point:

other: 1 h

Score:

96.9

Max. score:

100

Reversibility:

no data

Remarks on result:

other: Non-irritant

Irritant / corrosive response data:

See Tables 1 to 4

Other effects:

No unforeseen events occurred which could impede the outcome of this study.

Any other information on results incl. tables

Table 1: EpiDerm Skin Corrosivity Test: Assay acceptance criteria, validity of the test
Sample	Mean tissue viability (%)	Assay acceptance criteria	Acceptable Yes/No
positive control	16.8	≤ 30 %	Yes
	mean OD
negative control	3 minutes 1.949 1 hour 1.907	≥ 0.8	Yes
	Maximum inter tissue viability difference (%)
GLUCOSE SYRUPS WHEAT HYDROLYZED	3 minutes 1.6 1 hour 6.7	< 30 %	Yes

OD = optical density

 

Table 2: EpiDerm Skin Irritation Test: Assay acceptance criteria, validity of the test
Sample	Mean tissue viability (%)	Assay acceptance criteria	Acceptable Yes/No
positive control	6.1	≤ 20 %	Yes
	mean OD
negative control	2.030	≥ 1.0 and ≤ 2.5	Yes
	SD viability (%)
GLUCOSE SYRUPS WHEAT HYDROLYZED	4.6	<18 %	Yes

OD = optical density; SD = standard deviation

 

Table 3: EpiDerm Skin Corrosivity Test: Cell viability measurement
3 minutes test	Substance	Tissue N	OD – blank corrected	Mean of aliquots	OD mean	Viability (%)	Viability mean (%)
	Negative control	1	1.986	1.949	1.949	100.0	100.0
			1.918
			1.942
		2	1.938	1.950		100.0
			1.953
			1.958
	Positive control	1	0.295	0.294	0.327	15.1	16.8
			0.291
			0.296
		2	0.359	0.359		18.4
			0.363
			0.356
	GLUCOSE SYRUPS WHEAT HYDROLYZED	1	1.897	1.891	1.906	97.0	97.8
			1.883
			1.893
		2	1.938	1.921		98.5
			1.940
			1.884
1 hour test	Negative control	1	1.857	1.847	1.907	96.9	100.0
			1.844
			1.839
		2	1.991	1.966		103.1
			1.942
			1.966
	Positive control	1	0.182	0.185	0.212	9.7	11.1
			0.187
			0.187
		2	0.240	0.238		12.5
			0.229
			0.246
	GLUCOSE SYRUPS WHEAT HYDROLYZED	1	1.928	1.935	1.872	101.5	98.2
			1.914
			1.962
		2	1.814	1.809		94.9
			1.801
			1.812

OD = optical density

 

Table 4: EpiDerm Skin Irritation Test: Cell viability measurement
Substance	Tissue N	OD – blank corrected	Mean of aliquots	OD mean	Viability (%)	Viability mean (%)	SD of viability
Negative control	1	2.142	2.145	2.030	105.62	100.0	5.1
		2.147
	2	1.944	1.946		95.85
		1.948
	3	2.000	2.001		98.53
		2.001
Positive control	1	0.124	0.125	0.124	6.13	6.1	0.4
		0.125
	2	0.114	0.115		5.64
		0.115
	3	0.135	0.132		6.50
		0.129
GLUCOSE SYRUPS WHEAT HYDROLYZED	1	1.952	1.951	1.967	96.07	96.9	4.6
		1.949
	2	2.057	2.069		101.90
		2.081
	3	1.884	1.883		92.72
		1.881

OD = optical density; SD = standard deviation

 

Applicant's summary and conclusion

Interpretation of results:

other: Not classified

Remarks:

Criteria used for interpretation of results: other: CLP (EC 1272/2008)