Calcium nitrite

Eye irritation

Administrative data

Endpoint:

eye irritation: in vivo

Type of information:

experimental study

Adequacy of study:

key study

Study period:

18 December 1995 - 6 January 1996

Reliability:

1 (reliable without restriction)

Rationale for reliability incl. deficiencies:

guideline study

Remarks:

Good quality study, conducted to GLP. The test material was a 34% solution of hydrated calcium nitrite.

Data source

Reference

Reference Type:

study report

Title:

Unnamed

Year:

1996

Report date:

1996

Materials and methods

Test guideline

Qualifier:

according to guideline

Guideline:

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

Version / remarks:

29.12.92

Deviations:

no

GLP compliance:

yes

Test material

Test material information

Constituent 1

Reference substance name:

Calcium nitrite

EC Number:

237-424-2

EC Name:

Calcium nitrite

Cas Number:

13780-06-8

Molecular formula:

Ca.2HNO2

IUPAC Name:

calcium nitrite

Constituent 2

Test material form:

liquid

Test animals / tissue source

Species:

rabbit

Strain:

New Zealand White

Details on test animals or tissues and environmental conditions:

TEST ANIMALS
- Source: Froxfield (UK) Ltd., Petersfield, Hampshire, England
- Age at study initiation: 12-13 weeks
- Weight at study initiation: 2.7-3.1 kg
- Housing: Individually housed in metal cages with perforated floors
- Diet (e.g. ad libitum): SDS Stanrab (P) standard laboratory rabbit diet ad libitum
- Water (e.g. ad libitum): drinking water ad libitum
- Acclimation period: not specified

ENVIRONMENTAL CONDITIONS
- Temperature (°C): 19
- Humidity (%): 30-70
- Air changes (per hr): 19
- Photoperiod (hrs dark / hrs light): 12/12

IN-LIFE DATES: No data

Test system

Vehicle:

unchanged (no vehicle)

Controls:

other: contralateral eye of treated animals served as concurrent control

Amount / concentration applied:

TEST MATERIAL
- Amount(s) applied (volume or weight with unit): 0.1 mL [equivalent to about 0.034 mL as calcium nitrite]
- Concentration (if solution): the test material was administered as supplied (34% aqueous solution of calcium nitrite)

Duration of treatment / exposure:

Eyelids were gently held together for one second before releasing; no subsequent washing to remove test substance.

Observation period (in vivo):

17 days. Examination of the eyes was made 1 hour and 1, 2, 3 (24, 48 and 72 hours), 4, 7, 14, 15 and 17 days after instillation.

Number of animals or in vitro replicates:

Two (one female, sex not specified in the other)

Details on study design:

REMOVAL OF TEST SUBSTANCE
- Washing (if done): the treated eye was not rinsed
- Time after start of exposure: not applicable

SCORING SYSTEM: Ocular irritation was assessed using the prescribed numerical system:
Corneal opacity: No ulceration or opacity (0); scattered or diffuse areas of opacity (other than slight dulling of normal lustre), details of iris clearly visible (1); easily discernable translucent areas, details of iris slightly obscured (2); nacreous areas, no details of iris visible, size of pupil barely discernible (3); opaque cornea, iris not discernible through the opacity (4).
Iris: normal (0); markedly deepened rugae, congestion, swelling, moderate circumcorneal hyperaemia or injection (or combinations thereof), iris still reacting to light (sluggish reaction is positive) (1); no reaction to light, haemorrhage or gross destruction (or combinations thereof) (2).
Conjunctivae: blood vessels normal (0); some blood vessels hyperaemic (injected) (1); diffuse, crimson colour, individual vessels not easily discernible (2); diffuse beefy red (3).
Chemosis: no swelling (0); any swelling above normal (includes nictating membranes) (1); obvious swelling with partial eversion of lids (2); swelling with lids about half-closed (3); swelling with lids more than half-closed (4).

Any other lesion not covered by this scoring system was described.


TOOL USED TO ASSESS SCORE: handheld light

Results and discussion

In vivo

Results

Results 1

Irritation parameter:

overall irritation score

Reversibility:

other: reversibility was not fully assessed due to humane sacrifice of the animals on Day 15 and 17 (after instillation of the test substance), respectively

Remarks on result:

other: An irritation index could not be determined because of the corrosive effects

Results 2

Irritation parameter:

cornea opacity score

Basis:

mean

Time point:

24/48/72 h

Score:

0

Max. score:

4

Reversibility:

other: Corneal effects were only apparent 2 weeks after instillation of the test material; reversibility was not fully assessed due to humane sacrifice of the animals on Day 15 and 17 (after instillation of the test substance), respectively

Results 3

Irritation parameter:

iris score

Basis:

mean

Time point:

24/48/72 h

Score:

0

Max. score:

2

Reversibility:

other: Iridial effects were only apparent 2 weeks after instillation of the test material; reversibility was not fully assessed due to humane sacrifice of the animals on Day 15 and 17 (after instillation of the test substance), respectively

Results 4

Irritation parameter:

conjunctivae score

Basis:

mean

Time point:

24/48/72 h

Score:

2.3

Max. score:

3

Reversibility:

other: Conjunctival effects (redness) tended to worsen with time; reversibility was not fully assessed due to humane sacrifice of the animals on Day 15 and 17 (after instillation of the test substance), respectively

Results 5

Irritation parameter:

chemosis score

Basis:

mean

Time point:

24/48/72 h

Score:

2

Max. score:

4

Reversibility:

other: Conjunctival effects (chemosis) tended to worsen with time; reversibility was not fully assessed due to humane sacrifice of the animals on Day 15 and 17 (after instillation of the test substance), respectively

Irritant / corrosive response data:

Conjunctival redness and chemosis scores of 2-3 were observed in the first animal at all time points (scores of 1-3 were observed in the second animal throughout the observation period); in both cases the effects tended to worsen with time. Necrosis of the conjunctiva and nictating membrane was reported for both animals. Corneal and iridial effects were only apparent 2 weeks after instillation of the test material; these also worsened with time (during the third week of the observation period). Due to the severity of the effects the animals were killed on Day 15 and 17, respectively.

Other effects:

- Lesions and clinical observations: No signs of ill health or toxicity were reported.
- Ophthalmoscopic findings: no data
- Histopathological findings: no data
- Effects of rinsing or washing: not applicable

Applicant's summary and conclusion

Interpretation of results:

Category 1 (irreversible effects on the eye) based on GHS criteria

Conclusions:

In a guideline study, to GLP, calcium nitrite (hydrate) (0.1 mL) produced severe irritation and corrosive effects following instillation into one eye of each of two rabbits.

Executive summary:

In an EU Method B.5 study, to GLP, the ocular irritant potential of calcium nitrite (hydrate) was assessed in two New Zealand White rabbits. The test material (DCI) was available as a 34% aqueous solution of calcium nitrite. Initially, a 0.1 mL instillation of the test material was made to the eye of one female and both lids were gently closed for one second before releasing. The other eye remained untreated and acted as the control. Due to the severity of the observed effects only one other animal (sex unspecified) was treated.

 

Following instillation of the test material, conjunctival redness and chemosis were seen in both animals within one hour. In both cases, the effects tended to worsen as the study progressed, leading to the sacrifice of the animals on Day 15 and 17, respectively. Additional effects included necrosis of the conjunctiva and nictating membrane, along with corneal and iridial effects (only apparent 2 weeks after instillation) which also worsened with time. An irritation index could not be determined because of the corrosive effects. No clinical signs of toxicity were reported.

 

As the results were based on only two rabbits, it is not possible to give a definitive interpretation according to EU CLP criteria (EC 1272/2008). However, it is considered appropriate to classify the test material for serious eye damage (Category 1), as the severe effects observed tended to worsen with time and are not expected to be reversible.