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Toxicological information

Eye irritation

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Administrative data

Endpoint:
eye irritation: in vitro / ex vivo
Type of information:
experimental study
Adequacy of study:
key study
Study period:
2009
Reliability:
1 (reliable without restriction)
Rationale for reliability incl. deficiencies:
other: Guideline study performed according to GLP

Data source

Reference
Reference Type:
study report
Title:
Unnamed
Year:
2009
Report Date:
2009

Materials and methods

Test guideline
Qualifier:
according to
Guideline:
other: ICCVAm Test Method Evaluation Report: Appendix G ICCVAM recommended HET-CAM Method Protocol (Nov. 2006)
Deviations:
no
Principles of method if other than guideline:
Background Review Document Current Status of In Vitro Test Methods for Identifying Ocular Corrosives and Severe Irritants: Hen’s Egg Test - Chorioallantoic Membrane Test Method (March 2006, NIH Publication No. 06-4515)
OECD 405 Draft Guideline “Acute Eye Irritation/Corrosion”, adopted 24. April 2002
GLP compliance:
yes (incl. certificate)

Test material

Reference
Name:
Unnamed
Type:
Constituent
Type:
Constituent
Details on test material:
Name Slags, steelmaking, elec. furnace (stainless/high alloy steel production - EAF S)
Appearance solid
Composition UVCB
CAS-No. 65996-69-2
EINECS-No. 266-002-0
Molecular formula not applicable (UVCB)
Molecular weight not applicable (UVCB
Purity 100 w/w % slag
Homogeneity not stated
Volatility extremely low (melting point > 300 °C)
Stability solid slag is stable at room temperature
Solubility slightly soluble in water
Date of production not stated
Date of expiry Year 2024
Storage conditions Room temperature 20 ± 5°C

Test animals / tissue source

Species:
other: hen
Strain:
other: Lohmann selected Leghorn chicken’s eggs
Details on test animals or tissues and environmental conditions:
in vitro test system HETCAM applied
Freshly laid Lohmann Leghorn chicken eggs were obtained from the LSL Rhine Main Gefluegelvermehrungsbetrieb (Dieburg, Germany) nine days before the start of the test. The eggs were incubated at 38.3 ± 0.2 °C and 58 % ± 2 % humidity in a still air incubator for eight days. During incubation, the eggs were rotated to prevent an attachment of the embryo to one side of the egg.

Test system

Vehicle:
unchanged (no vehicle)
Controls:
not required
Amount / concentration applied:
On the day of the test, the air bubble of a freshly incubated egg was marked. The egg was opened on the air bubble and the inner membrane was moistened with 0.9 % sodium chloride. After 30 min in the incubator, the solution was decanted and the inner membrane (CAM) was carefully removed with a forceps.
300 µL (equalling to 252.5 mg; this is the mean of all slag weights) of the test material were given directly onto the CAM surface in such a manner that 50% of the surface of the membrane were covered with test item.
Duration of treatment / exposure:
The reactions of the CAM were observed during 5 min:
• haemorrhage (bleeding from the vessels)
• vascular lysis (blood vessel disintegration)
• coagulation (intra- and extra-vascular protein denaturation)
The time for the appearance of each of the noted endpoints was monitored and recorded in seconds
Observation period (in vivo):
5 min
Number of animals or in vitro replicates:
0
Details on study design:
Sodium dodecyl sulfate solution C12H25NaO4S, CAS-No. 151-21-3. Solvent is deionised water, 1% solution (10 g/L) and
sodium hydroxide solution 0.1 n NaOH, CAS-No. 1310-73-2, dissolved in deionised water at 4 g/L, were used as positive controls:
Sodium chloride solution, NaCl, 0.9% solution in deionised water, was used as a negative control:
The following instruments and devices were used:
• Autoclave MLS 3020
• Candling light (for determination of the position of the egg bubble)
• Dentist’s rotating saw blade (for opening the egg)
• Tweezers
• Stop clock
• Scales Mettler PB 5001-S02
• Analytical scales MT AB 184 SA
• Heating chamber
• Orbital shaker GFL 3005
• Adjustable pipettes with sterile tips, LAUS No. 8 (0.2 – 2 mL)
• Digital Hygrometer
• Schott-bottles, glass vials, and culture flasks for solutions and media (sterilised before use by heating to 180 °C (two hours) or autoclavation)

Results and discussion

In vitro

Results
Irritation parameter:
in vitro irritation score
Run / experiment:
The HET-CAM test
Negative controls validity:
valid
Positive controls validity:
valid
Remarks on result:
no indication of irritation

In vivo

Resultsopen allclose all
Irritation parameter:
other: haemorrhage
Basis:
mean
Time point:
other: 5 min
Score:
0
Max. score:
0
Reversibility:
other: not relevant as no effect occured
Irritation parameter:
other: lysis
Basis:
mean
Time point:
other: 5 min
Score:
0
Max. score:
0
Reversibility:
other: not relevant as no effect occured
Irritation parameter:
other: coagulation
Basis:
mean
Time point:
other: 5 min
Score:
0
Max. score:
0
Reversibility:
other: not relevant as no effect occured
Irritant / corrosive response data:
see Table 3
Other effects:
no relevant effect

Any other information on results incl. tables

The observations which were made are given in the following table:

Table 3.  Experimental Observations

Test Group

Amount applied

Time until haemorrhage

Time until lysis

Time until coagulation

 

 

seconds

seconds

seconds

Negative Control

300 µL

301

301

301

Negative Control

300 µL

301

301

301

Negative Control

300 µL

301

301

301

Positive Control 0.1-N NaOH

300 µL

8

34

14

Positive Control 0.1-N NaOH

300 µL

7

32

16

Positive Control 0.1-N NaOH

300 µL

7

29

17

Positive Control 1% SDS

300 µL

20

57

301

Positive Control 1% SDS

300 µL

17

58

301

Positive Control 1% SDS

300 µL

10

59

301

Slag, ferrous metal, blast furnace (air cooled - ABS), fine-ground

264.1 mg

301

301

301

Slag, ferrous metal, blast furnace (air cooled - ABS), fine-ground

250.7 mg

301

301

301

Slag, ferrous metal, blast furnace (air cooled - ABS), fine-ground

247.5 mg

301

301

301

Slag, ferrous metal, blast furnace (air cooled - ABS), fine-ground

240.8 mg

301

301

301

Slag, ferrous metal, blast furnace (air cooled - ABS), fine-ground

245.3 mg

301

301

301

Slag, ferrous metal, blast furnace (air cooled - ABS), fine-ground

266.3 mg

301

301

301

Applicant's summary and conclusion

Interpretation of results:
not irritating
Remarks:
Migrated information Criteria used for interpretation of results: EU
Conclusions:
Slags, steelmaking, elec. furnace (stainless/high alloy steel production - EAF S) have no irritation potential
Executive summary:

Fine ground test material of slags, steelmaking, elec. furnace (stainless/high alloy steel production - EAF S) was brought onto the surface of the chorio-allantoic membrane (CAM) of a hen’s egg which had been incubated at 38.3oC and 58 % humidity for nine days.Observation time was five minutes at room temperature (negative controls: physiological sodium chloride solution, positive controls sodium dodecylsulfat (SDS, 1 %) and NaOH (0.1 n))

Slags, steelmaking, elec. furnace (stainless/high alloy steel production - EAF S) caused no effects on the blood vessels of the CAM (no haemorrhage, no lysis, no coagulation). The calculated mean irritation score is 0.0.

Slags, steelmaking, elec. furnace (stainless/high alloy steel production - EAF S) have no irritation potential.