Titanium carbide

Endpoint summary

Description of key information

The test item titanium carbide was assessed for dermal irritation in vitro according to OECD 439. Based on the results of this test, the test item is not considered a dermal irritant.

Eye irritation was assessed in vitro (BOCP; OECD 437) and confirmed in vivo (OECD 405). No effects relevant for classification of the test item were observed.

Key value for chemical safety assessment

Skin irritation / corrosion

Link to relevant study records

Reference

Endpoint:

skin irritation: in vitro / ex vivo

Type of information:

experimental study

Adequacy of study:

key study

Study period:

2012-08-22 to 2012-10-18

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:

22 July 2010

Deviations:

no

Qualifier:

according to guideline

Guideline:

EU Method B.46 (In Vitro Skin Irritation: Reconstructed Human Epidermis Model Test)

Version / remarks:

06 July 2012

GLP compliance:

yes (incl. QA statement)

Specific details on test material used for the study:

SOURCE OF TEST MATERIAL
- batch number of test material: 82496
- Purity: ≥99%

TREATMENT OF TEST MATERIAL PRIOR TO TESTING
- 5 µl distilled water were aplied by a pipette to the epidermal surface in order to improve further contact between the powder and the epidermis. The water was gently spread with the pipette. Afterwards, approximately 10 mg (26.3 mg/cm³) of the powder were applied to the epidermis surface.

Test system:

human skin model

Source species:

human

Cell type:

non-transformed keratinocytes

Justification for test system used:

This test uses the EPISKIN-SM(TM) reconstructed human epidermis model (Skin Ethic) which consists of human keratinocytes (NHEK) and therefore represents in vitro the target organ of the species of interest and closely mimics the biochemical and physiological properties of the upper parts of the human skin, i.e. the epidermis.

Vehicle:

water

Details on test system:

RECONSTRUCTED HUMAN EPIDERMIS (RHE) TISSUE
- Model used: EPISKIN-SM(TM)
- Tissue batch number(s): 12-EKIN-030
- Production date: August 21, 2012
- Shipping date: not specified
- Delivery date: not specified
- Date of initiation of testing: 22 August 2012

TEMPERATURE USED FOR TEST SYSTEM
- Temperature used during treatment / exposure: room temperature
- Temperature of post-treatment incubation (if applicable): 37 ± 1 °C

REMOVAL OF TEST MATERIAL AND CONTROLS
- Volume and number of washing steps: not specified
- Observable damage in the tissue due to washing: not specified
- Modifications to validated SOP: not specified

MTT DYE USED TO MEASURE TISSUE VIABILITY AFTER TREATMENT / EXPOSURE
- MTT concentration: final concentration 0.3 mg/ml
- Incubation time: 3h ± 5 min
- Wavelength: 550 nm

FUNCTIONAL MODEL CONDITIONS WITH REFERENCE TO HISTORICAL DATA
- Barrier function: exceeded specifications
- Morphology: Well-differentiated epidermis consisting of a basal layer, several spinous and granular layers and a thick stratum corneum. Exceeded specifications.

NUMBER OF REPLICATE TISSUES: The test was performed on a total of 3 tissues per dose group.

CONTROL TISSUES USED IN CASE OF MTT DIRECT INTERFERENCE
- The test item showed no reduction of MTT compared to the solvent and showed no colouring detectable by unaided eye-assessment

NUMBER OF INDEPENDENT TEST SEQUENCES / EXPERIMENTS TO DERIVE FINAL PREDICTION: One

PREDICTION MODEL / DECISION CRITERIA
- The test substance is considered to be irritant to skin if the viability after 15 minutes exposure and 42 hours of post-incubation is less than or equal to 50%. The test substance may be considered non-irritant to skin if the tissue viability after exposure and post-treatment incubation is higher than 50%.

Control samples:

yes, concurrent negative control

yes, concurrent positive control

Amount/concentration applied:

TEST MATERIAL
- Amount(s) applied: 10 mg (26.3 mg/cm²)

VEHICLE
- Amount(s) applied: 5 µl
- Purity: distilled water

NEGATIVE CONTROL
- Amount(s) applied: 10 µl

POSITIVE CONTROL
- Amount(s) applied: 10 µl
- Concentration: 5% SDS

Duration of treatment / exposure:

15 ± 0.5 min

Duration of post-treatment incubation (if applicable):

42 hours

Number of replicates:

3

Details on study design:

TEST SITE
- Area of exposure: 0.38 cm²
- % coverage: 100 %

REMOVAL OF TEST SUBSTANCE
- Washing (if done): phosphate buffered saline
- Time after start of exposure: 15 ± 0.5 min

PRE-EXPERIMENT
To check the non specific MTT-reducing capability of the test item 10 mg of the test item were mixed per 2 mL MTT medium and incubated for 3 h at 37 ± 1 °C in the dark. If the mixture turns blue/purple, the test item is presumed to have reduced MTT. To check the colouring potential of the test item 10 mg of the test item were mixed per 90 μL Aqua dest. in a transparent recipient for 15 min.

EXPERIMENTAL PROCEDURE
Upon receipt of the EPISKIN-SM, the tissues were transferred into 12-well plates containing 2 mL prewarmed maintenance medium per well. The 12-well plates were incubated in a humidified incubator at 37 ± 1 °C, 5.0% CO2 for at least 24 h.
After this pre-incubation the tissues were treated with each dose group in triplicate, starting with the negative control. Start time was recorded with dosing of the first tissue. Then the tissues were incubated at room temperature for 15 ± 0.5 min. Afterwards, the tissues were washed with PBS to remove any residual test item. Excess PBS was removed by blotting bottom with blotting paper. The inserts were placed in a prepared 12-well plate containing 2 mL prewarmed fresh maintenance medium and post-incubated at 37 ± 1 °C, 5.0% CO2 for 42 ± 1 h.
After this incubation period the plates were placed for 15 ± 2 min. on a plate shaker. Then the inserts were transferred in a prepared 12-well plate containing 2 mL prewarmed MTT medium and further incubated for 3 h ± 5 min. at 37 ± 1 °C, 5.0% CO2.
After the 3 h MTT incubation period the tissues were placed on blotting paper to dry the tissues. Afterwards a total biopsy of the epidermis by using the special biopsy punch was performed and the epidermis was separated from the collagen matrix with the aid of forceps. Both parts (epidermis and collagen matrix) were transferred into suitable tubes and 500 μL of acidic isopropanol were added. Extraction was carried out protected from light over the weekend at 2 - 8°C.
At the end of the formazan extraction period the tubes were mixed by vortexing until solution colour became homogeneous. If any visible cell/tissue fragments were in suspension, the tubes were centrifuged at 500 rpm to eliminate the fragments and avoid further possible interference with the absorbance readings. Per tissue 2 x 200 μL aliquots of the extract were transferred into a 96-well plate and OD was measured at 550 nm without reference wavelength in a plate spectrophotometer.

SCORING SYSTEM:
Irritant potential of the test item was predicted from the relative mean tissue viabilities compared to the negative control tissues concurrently treated with PBS. The test item is considered to be irritant to skin in accordance with regulation EC1272/2008 and UN GHS "Category 2", if the tissue viability after 15 min of exposure and 42 h of post-incubation is less or equal to 50%. The test substance may be considered as non-irritant to skin in accordance with UN GHS "No category" if the tissue viability after exposure and post-treatment incubation is higher than 50%.

Barrier function and morphology were provided by the Rhe model supplier to assure the proper functioning of the tissues.

Irritation / corrosion parameter:

% tissue viability

Run / experiment:

Test item (mean of 3 replicates)

Value:

> 50

Vehicle controls validity:

not applicable

Negative controls validity:

valid

Positive controls validity:

valid

Remarks on result:

other:

Remarks:

Remarks: The results are presented in the table below.

Table 1: Experimental results of the EPISKIN-SM test after application of Titanium carbide powder.

Name	Negative Control			Positive Control			Test Item
Tissue	1	2	3	1	2	3	1	2	3
Mean OD550of 3 replicate tissues (blank-corrected	0.742			0.184			0.740
SD OD550	0.052			0.080			0.077
Relative tissue viabilities (%)	100.2	104.6	95.2	37.6	21.6	15.1	105.8	105.6	87.9
Mean relative tissue viability (%)	100			25			100
SD tissue viability (%)	4.7			11.6			10.3
CV (% viability)	4.7			46.8			10.3

Interpretation of results:

GHS criteria not met

Conclusions:

Titanium carbide is no irritant in the human skin model EPISKIN-SM.

Executive summary:

In this in vitro dermal irritation study with the three-dimensional human skin model EPISKIN-SM conducted according to OECD TG 439, the irritation potential of Titanium carbide was examined. The test substance (10 mg) was applied topically on the tissue for 15 min, and thereafter the exposed tissue culture was incubated for 42 h. A positive and a negative control were used. The determination of cytotoxic effects was performed with the MTT assay. Irritation potential of the test susbstance was determined based on the cytotoxicity observed (mean tissue viability). All acceptability criteria were fulfilled. The results revealed a mean relative tissue viability higher than 50%, and hence, the test substance is not a skin irritant under the conditions of this test.

Endpoint conclusion

Endpoint conclusion:

no adverse effect observed (not irritating)

Eye irritation

Link to relevant study records

Reference

Reference 1

Endpoint:

eye irritation: in vivo

Type of information:

experimental study

Adequacy of study:

key study

Study period:

2012-11-23 to 2012-12-14

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:

24 April 2002

Deviations:

no

Qualifier:

according to guideline

Guideline:

EU Method B.5 (Acute Toxicity: Eye Irritation / Corrosion)

Version / remarks:

30 May 2008

Qualifier:

according to guideline

Guideline:

EPA OPPTS 870.2400 (Acute Eye Irritation)

Version / remarks:

August 1998

GLP compliance:

yes (incl. QA statement)

Specific details on test material used for the study:

TREATMENT OF TEST MATERIAL PRIOR TO TESTING
- The test item was used as provided by the sponsor

Species:

rabbit

Strain:

New Zealand White

Details on test animals or tissues and environmental conditions:

TEST ANIMALS
- Source: Charles River Deutschland, 97633 Sulzfeld, Germany
- Age at study initiation: approximately 13 weeks old
- Weight at study initiation: > 2 kg
- Housing: ABS - plastic rabbit cages, floor 4200 cm²
- Diet: Free access to autoclaved hay and to Altromin 2123 maintenance diet for rabbits (lot no. 1516), rich in crude fibre
- Water: Free access to tap water (drinking water, municipal residue control, microbiological controls at regular intervals)
- Certificates of food, water and bedding are filed at BSL BIOSERVICE
- Acclimation period: at least 5 days under laboratory conditions

ENVIRONMENTAL CONDITIONS
- Temperature (°C): 18 ± 3 °C (recommendations of TVT, GV-SOLAS)
- Humidity (%): 55 ± 10 °C
- Air changes (per hr): 10
- Photoperiod (hrs dark / hrs light): 12 h : 12 h

TVT = TVT Tierärztliche Vereinigung für Tierschutz, Merkblatt 127 zur Tiergerechten Haltung von Versuchskaninchen, 2010
GV-SOLAS = Planung, Struktur von Versuchstierbereichen tierexperimentell tätiger Institutionen; Veröffentlichung GV-SOLAS (Gesellschaft für Versuchstierkunde, Society for Laboratory Animal Science), Mai 1988

Vehicle:

unchanged (no vehicle)

Controls:

other: untreated eye of each animal served as control

Amount / concentration applied:

TEST MATERIAL
- Amount(s) applied (volume or weight with unit): 0.1 g

Duration of treatment / exposure:

72 h (the treated eye was not rinsed 24 hours after the application.)

Observation period (in vivo):

72 h

Number of animals or in vitro replicates:

3

Details on study design:

REMOVAL OF TEST SUBSTANCE
- Washing: none

SCORING SYSTEM: see "Any other information on materials and methods incl. tables"

TOOL USED TO ASSESS SCORE: fluorescein

Irritation parameter:

cornea opacity score

Basis:

animal #1

Time point:

24/48/72 h

Score:

0

Max. score:

4

Remarks on result:

no indication of irritation

Irritation parameter:

cornea opacity score

Basis:

animal #2

Time point:

24/48/72 h

Score:

0

Max. score:

4

Remarks on result:

no indication of irritation

Irritation parameter:

cornea opacity score

Basis:

animal #3

Time point:

24/48/72 h

Score:

0

Max. score:

4

Remarks on result:

no indication of irritation

Irritation parameter:

iris score

Basis:

animal #1

Time point:

24/48/72 h

Score:

0

Max. score:

2

Remarks on result:

no indication of irritation

Irritation parameter:

iris score

Basis:

animal #2

Time point:

24/48/72 h

Score:

0

Max. score:

2

Remarks on result:

no indication of irritation

Irritation parameter:

iris score

Basis:

animal #3

Time point:

24/48/72 h

Score:

0

Max. score:

2

Remarks on result:

no indication of irritation

Irritation parameter:

conjunctivae score

Basis:

animal #1

Time point:

24/48/72 h

Score:

0.67

Max. score:

3

Reversibility:

fully reversible within: 72 h

Remarks on result:

no indication of irritation

Irritation parameter:

conjunctivae score

Basis:

animal #2

Time point:

24/48/72 h

Score:

0.33

Max. score:

3

Reversibility:

fully reversible within: 48 h

Remarks on result:

no indication of irritation

Irritation parameter:

conjunctivae score

Basis:

animal #3

Time point:

24/48/72 h

Score:

0.33

Max. score:

3

Reversibility:

fully reversible within: 48 h

Remarks on result:

no indication of irritation

Irritation parameter:

chemosis score

Basis:

animal #1

Time point:

24/48/72 h

Score:

0

Max. score:

4

Remarks on result:

no indication of irritation

Irritation parameter:

chemosis score

Basis:

animal #2

Time point:

24/48/72 h

Score:

0

Max. score:

4

Remarks on result:

no indication of irritation

Irritation parameter:

chemosis score

Basis:

animal #3

Time point:

24/48/72 h

Score:

0

Max. score:

4

Remarks on result:

no indication of irritation

Irritant / corrosive response data:

- First time of effects, all animals: 1 hour post-application redness grade 1
- Last time of effects:
* animal no. 1: 48 hours post-application redness grade 1
* animal no. 2 and no. 3: 24 hours post-application redness grade 1
- Reversibility of the observed effects:
* animal no. 1: the changes were fully reversible within 72 hours post-application
* animal no. 2 and no. 3: the changes were fully reversible within 48 hours post-application

Other effects:

After the application into the eyes of three female NZW rabbits the test item produced irritant but no corrosive effects (see Tables 3 to 5 in section "Any other information on results incl. tables").
Neither mortalities nor significant clinical signs of toxicity were observed.
The eyes were not rinsed after the application.
Upon fluorescein examinations at the end of the observation period of 72 hours no corneal lesions were found in any animal.
Conjunctival redness was observed in all animals and are individually described in Tables 3 to 5 in section "Any other information on results incl. tables".

Table 2: Mean Values of Eye Irritation Scores – (24, 48, 72 Hour Reading)

Animal No.	Sex	Conjunctival Redness	Conjunctival Chemosis	Iris Cornea	Opacity
1	female	0.67	0.00	0.00	0.00
2	female	0.33	0.00	0.00	0.00
3	female	0.33	0.00	0.00	0.00

 

Table 3: Eye Irritation Scores – Animal No. 1

Single Data
	Time Post-Application				Average Score (24, 48 and 72 hours)
	1 hour	24 hours	48 hours	72 hours
	T/C	T/C	T/C	T/C
Conjunctival Redness	1/0	1/0	1/0	0/0	0.67
Conjunctival Chemosis	0/0	0/0	0/0	0/0	0
Iris	0/0	0/0	0/0	0/0	0
Cornea	0/0	0/0	0/0	0/0	0

T = Test item, C = control

Table 4: Eye Irritation Scores – Animal No. 2 

Single Data
	Time Post-Application				Average Score (24, 48 and 72 hours)
	1 hour	24 hours	48 hours	72 hours
	T/C	T/C	T/C	T/C
Conjunctival Redness	1/0	1/0	0/0	0/0	0.33
Conjunctival Chemosis	0/0	0/0	0/0	0/0	0
Iris	0/0	0/0	0/0	0/0	0
Cornea	0/0	0/0	0/0	0/0	0

T = test item, C = control

Table 5: Eye Irritation Scores – Animal No. 3

Single Data
	Time Post-Application				Average Score (24, 48 and 72 hours)
	1 hour	24 hours	48 hours	72 hours
	T/C	T/C	T/C	T/C
Conjunctival Redness	1/0	1/0	0/0	0/0	0.33
Conjunctival Chemosis	0/0	0/0	0/0	0/0	0
Iris	0/0	0/0	0/0	0/0	0
Cornea	0/0	0/0	0/0	0/0	0

T = test item, C = control

Interpretation of results:

GHS criteria not met

Conclusions:

In this study, titanium carbide is not irritant to the eye.

Executive summary:

In a primary eye irritation study according to OECD 405, 0.1 g of titanium carbide was instilled into the conjunctival sac of one eye each of three female New Zealand White rabbits (age approximately 13 weeks). Eyes were not washed. Animals were observed for a total of 72 hours. Irritation was scored by evaluating conjunctival redness, conjunctival chemosis, iris, and corneal opacity.

Only slight effects visible as conjunctival redness have been observed which were fully reversible within 72 hours. Neither mortalities nor significant clinical signs of toxicity were observed.

In this study, titanium carbide is not an eye irritant.

Reference 2

Endpoint:

eye irritation: in vitro / ex vivo

Type of information:

experimental study

Adequacy of study:

key study

Study period:

2012-07-12 to 2012-12-03

Reliability:

1 (reliable without restriction)

Rationale for reliability incl. deficiencies:

guideline study

Qualifier:

according to guideline

Guideline:

OECD Guideline 437 (Bovine Corneal Opacity and Permeability Test Method for Identifying Ocular Corrosives and Severe Irritants)

Version / remarks:

07 September 2009

Deviations:

no

GLP compliance:

yes (incl. QA statement)

Specific details on test material used for the study:

TREATMENT OF TEST MATERIAL PRIOR TO TESTING
- The test item was suspended with physiological saline 0.9% NaCl to gain a 20% concentration.

Species:

cattle

Strain:

not specified

Details on test animals or tissues and environmental conditions:

SOURCE OF COLLECTED EYES
- Source: The assay uses isolated corneas obtained as a by-product from an abattoir from freshly slaughtered animals, e.g. from a local slaughterhose.
- Number of animals: not specified
- Characteristics of donor animals (e.g. age, sex, weight): not specified
- Storage, temperature and transport conditions of ocular tissue (e.g. transport time, transport media and temperature, and other conditions): On the test day, fresh eyes were collected from the slautherhouse and were transported in HBSS containing Pen/Strep on ice to the laboratories. Immediately after arrival of the eyes, cornea preparation was initiated.
- Time interval prior to initiating testing: Immediately after arrival
- Indication of any existing defects or lesions in ocular tissue samples: The eyes were carefully examined for defects and any defective eyes were discarded.
- Indication of any antibiotics used: 5% penicillin/ streptomycin (Gibco, lot no. 109259L, expiry date: 12/2012)
- Selection and preparation of corneas: The tissue surrounding the eyeball was carefully pulled away and the cornea was excised leaving a 2 to 3 mm rim of sclera. The isolated corneas were stored in a petri dish containing HBSS.
- Quality check of the isolated corneas: Before the corneas were mounted in corneal holders (MC2, Clermont, France) with the endothelial side against the O-ring of the posterior chamber, they had been visually examined for defects and any defective cornea had been discarded.

Vehicle:

physiological saline

Remarks:

0.9 % NaCl

Controls:

yes, concurrent positive control

yes, concurrent negative control

Amount / concentration applied:

TEST MATERIAL
- Amount(s) applied (volume or weight with unit): 750 µL
- Concentration (if solution): 20 %

VEHICLE
- Amount(s) applied (volume or weight with unit): physiol. saline
- Concentration (if solution): 0.9 %
- Lot/batch no. (if required): 111214

Duration of treatment / exposure:

After 4 h +/- 5 min exposure with TiC or control substance in 0.9 % physiol. saline;
After washing subsequent incubation for 90 minutes with fresh complete RPMI and 1 mL of a 5 mg/mL sodium fluorescein solution

Observation period (in vivo):

Not applicable

Duration of post- treatment incubation (in vitro):

90 minutes at 32 ± 1 °C.

Number of animals or in vitro replicates:

- 3 corneas for the test item
- 3 corneas as negative controls treated with physiological saline 0.9% NaCl
- 3 corneas as positive control treated with imidazole 20% in physiological saline 0.9% NaCl

The BCOP assay is considered to be valid if the in vitro irritation score obtained with the positive control falls within the two standard deviations of the current historical mean.

Details on study design:

NUMBER OF REPLICATES: Three

NEGATIVE CONTROL USED: physiological saline (0.9% NaCl)

POSITIVE CONTROL USED: 20% imidazole in physiological saline (0.9% NaCl)

APPLICATION DOSE AND EXPOSURE TIME: 750 µl for 4h ± 5 minutes at 32 ± 1 °C

TREATMENT METHOD: closed chamber

REMOVAL OF TEST SUBSTANCE
- Number of washing steps after exposure period: at least 3 times with minimum essential medium (MEM)
- POST-EXPOSURE INCUBATION: yes. 90 minutes at 32 ± 1 °C

METHODS FOR MEASURED ENDPOINTS:
- Corneal opacity: The opacitometer (MC2, Clermont, France) had been switched on 15 min before the calibration procedure was started. Empty cornea holders were placed into the opacitometer and the readout was adjusted to zero using the “BAL”-turning knob. For calibration the polyester foil no. 1 was introduced into the test chamber and the readout was adjusted to 75 using the “CAL”-turning knob. To test the linearity of the measurement, two additional calibration foils, polyester foil no. 2 and polyester foil no. 3, were measured. For these, the opacitometer was supposed to display 150 and 225, respectively (± 3%). If this had not been the case, the calibration procedure would have had to be repeated. The calibration procedure was performed before each test and was documented in the raw data.
An initial opacity measurement was performed after the equilibration period when the medium was removed from both chambers and replaced with fresh complete RPMI. Following the incubation period, the epithelium was washed at least three times with MEM (containing phenol red). Once the medium was free of test substance, the cornea was finally rinsed with complete RPMI (without phenol red). The anterior chamber was refilled with complete RPMI and an opacity measurement was performed.
The change in opacity for each cornea was calculated by subtracting the initial opacity reading from the final opacity reading. These values were corrected by subtracting from each the average change in opacity observed for the negative-control corneas. The mean opacity value for each treatment was calculated by averaging the corrected opacity values of each cornea for a given treatment. The mean OD490 for the blank wells were calculated.
- Corneal permeability: After the opacity measurement the medium was removed from both chambers of the holder. The posterior chamber was refilled with fresh complete RPMI. 1 ml of a 5 mg/ml sodium fluorescein solution was added to the anterior chamber and the corneas were incubated for 90 minutes at 32 ± 1 °C. Then the medium from the posterior chamber was removed and its optical density at 490 nm (OD490) was determined, using a spectrophotometer.
The mean blank OD490 was subtracted from the OD490 of each well (corrected OD490). Any dilutions that were made to bring the OD490 values into the linear range of the spectrophotometer (OD490 should be less than 1.500), were taken into account by multiplying the OD490 value of the dilution by the dilution factor. The final-corrected OD490 of the test article and the positive control were calculated by subtracting the average corrected OD490 of the negative control corneas from the corrected OD490 value of each treated cornea:

Final-corrected OD490 = (OD490 – mean blank OD490) – average-corrected negative control OD490

The mean OD490 value of each treatment group was calculated by averaging the final corrected OD490 values of the treated corneas for that treatment condition.

SCORING SYSTEM: In Vitro Irritancy Score (IVIS)
= mean opacity value + (15x mean permeability OD490 value)

DECISION CRITERIA:
The IVIS cut-off values for identifiying test substances as inducing serious eye damage (UN GHS Category 1) and test substances not requiring classification for eye irritation of serious eye damage (UN GHS No Category):
IVIS 0 – 3: non irritant
IVIS 3.1 - 25: mild irritant
IVIS 25.1 - 55: moderate irritant
IVIS 55.1 - 80: severe irritant (Category 1)
IVIS >80.1: very severe irritant

The assay is considered to be valid if the in vitro irritation score obtained with the positive control falls within the two standard deviations of the current historical mean.

Irritation parameter:

in vitro irritation score

Run / experiment:

4 hrs incubation with test item

Value:

2.19

Vehicle controls validity:

not applicable

Negative controls validity:

valid

Positive controls validity:

valid

Table 1: Evaluation of the BCOP Assay

Mean score	Evaluation
0 – 3	Non irritant
3.1 – 25	Mild irritant
25.1 – 55	Moderare irritant
55 – 80	Severe irritant
> 80.1	Very severe irritant

 

Table 2: Opacity

Cornea No.	Test item	Initial Opacity	Final Opacity	Change of Opacity Value	Corrected Opacity Value
1	Negative Control	5	6	1
2		5	5	0
3		5	3	-2
MV		5.00	4.67	-0.33
1	Positive Control	6	209	203	203.33
2		6	184	178	178.33
3		6	188	182	182.33
MV		6.00	193.67	187.67	188.00
1	Test Item	3	5	2	2.33
2		4	8	4	4.33
3		6	8	2	2.33
MV		4.33	7.00	2.67	3.00

 

Table 3: Permeability

Cornea No.	Test item	OD490	Corrected OD490 Value
1	Negative Control	0.087
2		0.089
3		0.110
MV		0.095
1	Positive Control	2.102	2.007
2		1.805	1.710
3		2.081	1.986
MV		1.996	1.901
1	Test Item	0.035	-0.060
2		0.018	-0.077
3		0.071	-0.024
MV		0.041	-0.054

 

 

Table 4: In Vitro Irritation Score

Cornea No.	Test item	Corrected Opacity Value	Corrected OD490 Value	IVIS
1	Negative Control	1.00	0.087
2		0.00	0.089
3		-2.00	0.110
MV		-0.33	0.095	1.10
1	Positive Control	203.33	2.007
2		178.33	1.710
3		182.33	1.986
MV		188.00	1.901	216.51
1	Test Item	2.33	-0.060
2		4.33	-0.077
3		2.33	-0.024
MV		3.00	-0.054	2.19

 

Interpretation of results:

GHS criteria not met

Conclusions:

According to the evaluation criteria the test item Titanium carbide is considered non-irritant to the eye.

Executive summary:

The eye irritancy potential of Titanium carbide was investigated in the bovine corneal opacity and permeability assay according to OECD 437. The Bovine Corneal Opacity and Permeability (BCOP) test method is an in vitro test method that can be used to classify substances as “ocular corrosives and severe irritants”.

Three corneas each were treated with the test item, imidazole 20 % in physiol. saline 0.9 % (positive control), and with physiol. saline 0.9 % (negaive control), respectively, for 4 hours at 32 °C. Optical density was measured at 490 nm (OD490).

The mean in vitro irritation score was 2.19. Thus, titanium carbide is not classified a severe irritant. The in vitro irritation score obtained with the positive control fell within the standard deviation of the historical mean.

Endpoint conclusion

Endpoint conclusion:

no adverse effect observed (not irritating)

Respiratory irritation

Endpoint conclusion

Endpoint conclusion:

no study available

Additional information

Titanium carbide was tested negative in an in vitro dermal irritation study with the three-dimensional human skin model EPISKIN-SM (OECD 439). The determination of cytotoxic effects was performed with the MTT assay. Irritation potential of the test substance was determined based on the cytotoxicity observed (mean tissue viability). The results revealed mean relative tissue viability higher than 50%, and hence, the test substance is not a skin irritant under the conditions of this test.

The eye irritancy potential was investigated in the bovine corneal opacity and permeability assay (BCOP) according to OECD 437. This in vitro test method can be used to classify substances as “ocular corrosives and severe irritants” only. The mean in vitro irritation score was 2.19. Thus, titanium carbide is not classified as severe eye irritant.

The irritation potential was further evaluated in a primary eye irritation study according to OECD 405. Titanium carbide was instilled into the conjunctival sac of one eye each of three female New Zealand White rabbits. Only slight effects have been observed which were fully reversible within 72 hours. Neither mortalities nor significant clinical signs of toxicity were observed. In this study, titanium carbide is not an eye irritant.

Justification for classification or non-classification

Titanium carbide is not considered an irritant to the skin or to the eye in any of the assays of a standard testing battery. Therefore classification according to the criteria set out in regulation (EC) No. 1272/2008 is not warranted.