Registration Dossier

Administrative data

Description of key information

Irritating potential of calcium hydrogen phosphonate was evaluated by in vitro tests.Negative results were obtained for skin irritation in the vitro study (OECD 439, with a mean cell viability of 100%).

Calcium hydrogen phosphonate, applied to rabbit eye mucosa, caused conjunctival effects at one hour after application which were fully reversible within 24 hours.

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:
11 January 2016 to 22 June 2016
Reliability:
1 (reliable without restriction)
Rationale for reliability incl. deficiencies:
guideline study
Qualifier:
according to
Guideline:
OECD Guideline 439 (In Vitro Skin Irritation: Reconstructed Human Epidermis Test Method)
Version / remarks:
2015
Deviations:
yes
Remarks:
Tp° in the incubator > 0.2°C.The SD value of viability (%) was 37.4% instead of 18% max for negative control tissues but homogenous results were obtained in test item treated tissues and historical data. No impact on the conclusion of the study.
Qualifier:
according to
Guideline:
EU Method B.46 (In Vitro Skin Irritation: Reconstructed Human Epidermis Model Test)
Deviations:
yes
Remarks:
see above
GLP compliance:
yes (incl. certificate)
Test system:
human skin model
Source species:
human
Cell type:
other: reconstructed human epidermis
Details on test system:
The 0.50 cm2 reconstructed epidermis (Episkin SA, RHE/S17 Batch No. 16-RHE-008)
Medium and Incubation T°C: 37°C
Control samples:
yes, concurrent no treatment
yes, concurrent positive control
Amount/concentration applied:
Amount applied per tissue : 16 mg
Duration of treatment / exposure:
Exposure period of 42 minutes, followed by rinsing. The rinsed tissues were checked for any coloration. There were incubated for a 41 hours and 05 minutes post-treatment incubation-period in fresh medium at 37°C, 5% CO2. Then the epidermis were put in contact with MTT solution. Observation of MTT-> formazan transformation by viable cells.
Number of replicates:
triplicates
Irritation / corrosion parameter:
% tissue viability
Value:
100
Vehicle controls validity:
not applicable
Negative controls validity:
valid
Positive controls validity:
valid
Remarks on result:
no indication of irritation
Remarks:
the exact % viability value was 118 considered 100
Other effects / acceptance of results:
In the preliminary tests, the test item was found not to have direct MTT reducing properties or significant colouring potential.
Main test: All acceptance criteria for the negative and positive controls were fulfilled except that the standard deviation value of viability percent of the tissues treated with the negative control was 37.4% instead of 18% (maximal value) as initially scheduled in the study plan. Considering the study results obtained in the tissues treated with the test item and the historical data, this deviation is considered as without impact on the conclusion of the study. Indeed, the results of the tissues treated with the test item are homogenous and all markedly above the cut-off of 50%. The study was therefore considered to be valid.
Following a 42 minutes exposure and 42 hours of recovery period, the relative mean viability of the tissues treated with the test item was 118% (considered 100%) with a standard deviation of 13%.
As the mean viability was > 50% after the MTT reduction, the IL-1 a concentrations in culture media samples retained were analyzed by ELISA
The mean IL-1a concentration for treated tissues was 7.08 pg/mL vs 169.54 pg/mL in the positive control. Due to this value being below 60 pg/mL, the results met the criteria for an in vitro classification as non-irritant to skin.
Interpretation of results:
GHS criteria not met
Conclusions:
In vitro, the test item is considered to be non-irritant to the skin.
Executive summary:

The objective of this study was to evaluate the skin irritation potential of calcium hydrogen phosphonate using the Episkin reconstructed human epidermis model. The study design was based upon international guidelines (OECD Guideline No. 439 and Commission Regulation (EC) No. 761/2009, B.46).

 Preliminary tests were performed to detect the ability of the test item to directly reduce MTT as well as its colouring potential. Following the preliminary tests, the skin irritation potential of the test item was tested in the main test. The test item and both the negative and positive controls were applied topically on triplicate tissues and incubated at room temperature for 42 minutes. At the end of the treatment period, each tissue was rinsed and incubated for 42 hours at 37.5°C, 5% CO2 in a humidified incubator. The cell viability was then assessed by means of the colourimetric MTT reduction assay.

Relative viability values were calculated for each tissue and expressed as a percentage of the mean viability of the negative control tissues which was set at 100% (reference viability).

In addition, the concentration of the inflammatory mediator IL-1awas evaluated in the culture medium retained following the 42-hour recovery period. This quantification, based on an ELISA assay, was performed since the mean relative viability of the test item-treated tissues was > 50% following the MTT reduction assay.

In the preliminary tests, the test item was found not to have direct MTT reducing properties or colouring potential.

 In the main test, all acceptance criteria for the negative and positive controls were fulfilled except that the standard deviation value of viability percent of the tissues treated with the negative control was 37.4% instead of 18% (maximal value) as initially scheduled in the study plan. Considering the study results obtained in the tissues treated with the test item and the historical data, this deviation is considered as without impact on the conclusion of the study. Indeed, the results of the tissues treated with the test item are homogenous and all markedly above the cut-off of 50%. The study was therefore considered to be valid.

Following a 42 minutes exposure and 42 hours of recovery period, the relative mean viability of the tissues treated with the test item was 118% (considered 100%) with a standard deviation of 13%.

As the mean viability was > 50% after the MTT reduction, the IL-1aconcentrations in culture media samples retained were analyzed by ELISA

The mean IL-1aconcentration for treated tissues was 7.08 pg/mL vs 169.54 pg/mL in the positive control. Due to this value being below 60 pg/mL, the results met the criteria for an in vitro classification as non-irritant to skin.

 Under the experimental conditions of this study, the test item is considered to be non-irritant to skin.

According to the results of this study, the classification of the test item should be:

.            not classified (Directive 67/548/EEC) and no category (Regulation (EC) No. 1272/2008).

Endpoint conclusion
Endpoint conclusion:
no adverse effect observed (not irritating)

Eye irritation

Link to relevant study records
Reference
Endpoint:
eye irritation: in vivo
Type of information:
experimental study
Adequacy of study:
key study
Study period:
12 May 2016 to 30 June 2016
Reliability:
1 (reliable without restriction)
Rationale for reliability incl. deficiencies:
guideline study
Qualifier:
according to
Guideline:
OECD Guideline 405 (Acute Eye Irritation / Corrosion)
Version / remarks:
OECD Guidelines for Testing of Chemicals 405 (2nd October 2012)
Deviations:
yes
Remarks:
see "Any other information" for details
Qualifier:
according to
Guideline:
EU Method B.5 (Acute Toxicity: Eye Irritation / Corrosion)
Version / remarks:
Commission Regulation (EC) No 440/2008, B.5 (L 142, 30 May 2008)
Deviations:
yes
Remarks:
See "Any other information" for details
Qualifier:
according to
Guideline:
EPA OPPTS 870.2400 (Acute Eye Irritation)
Version / remarks:
OPPTS 870.2400 (EPA 712-C-98-195) August 1998
Deviations:
yes
Remarks:
See "Any other information" for details
GLP compliance:
yes (incl. certificate)
Specific details on test material used for the study:
No further details specified in the study report.
Species:
rabbit
Strain:
New Zealand White
Details on test animals or tissues and environmental conditions:
Species and strain: New Zealand White rabbits
Source: S&K-LAP Kft., 2173 Kartal, Császár út 135, Hungary
Justification of strain: The New Zealand White rabbit is one of the standard strains used for acute irritation toxicity studies.
Animal health: Only animals in acceptable health condition were used for the test. Both eyes of each animal provisionally selected for testing were examined prior to starting the study. Animals showing eye irritation, ocular defects or pre-existing corneal injury were not used.
Number of animals: 3 animals
Age of animals at treatment: 11 weeks old
Sex: Male
Body weight range on the day of treatment: 2857 g – 3134 g before euthanasia: 2901 g – 3255 g
Date of receipt: 11 May 2016
Acclimatization time: 6-7 days
Animal identification: The animals were identified by engraved ear tags. The cages were marked with individual identity cards with information about study code, sex, cage number, dose and individual animal number.
HUSBANDRY
Animal health: Only healthy animals were used for the test. The veterinarian certified health status.
Housing/Enrichment: Rabbits were individually housed in AAALAC approved metal wire rabbit cages. Cages were of an open wire structure and cages were placed together to allow some social interaction with rabbit(s) in adjoining cages.
Number of animal room: 610
Light: 12 hours daily, from 6.00 a.m. to 6.00 p.m.
Temperature: 20.1 – 23.2 °C
Relative humidity: 25 – 69%
Ventilation: 15-20 air exchanges/hour
The temperature and relative humidity values were measured continuously. The measured range was checked at least daily during the acclimatisation and experimental phases.
FOOD AND FEEDING
Animals received UNI diet for rabbits produced by Cargill Takarmány Zrt., H-5300 Karcag, Madarasi út 0399, Hungary, ad libitum.
WATER SUPPLY
The animals received municipal tap water, as for human consumption, ad libitum, from an automatic system.
Vehicle:
unchanged (no vehicle)
Controls:
yes, concurrent no treatment
Amount / concentration applied:
An amount of 0.1 g of the test item was administered to the animals.
Duration of treatment / exposure:
As the solid test item remained in the eye sac in all animals at the one hour observation time point, the treated eye of test animals was rinsed with physiological saline solution.
Observation period (in vivo):
The eyes were examined at 1, 24, 48, 72 hours after treatment. The duration of the observation period was sufficient to identify reversibility or irreversibility of changes.
Number of animals or in vitro replicates:
Initially only one rabbit was treated with test item. The local effects were non-irritant (all scores were zero) at 24 hours, then two further rabbits were treated with the test item.
Details on study design:
In vitro study results
An in vitro eye irritation study was performed by PHYCHER Bio développement (study code: ICE-PH-15/0794) in the Isolated Chicken Eye model with test item prior to this study.The results from the in vitro eye irritation study indicated that the test item was not predicted as causing serious eye damage. However the score obtained did not allow a prediction in terms of eye irritation. Thus, an additional testing was required to establish a definitive classification.

Identification of pH
The pH of the test item was measured as pH 5.5, permitting the test item to be used in the animal studies.

Pre-study examination
Three male animals in acceptable health condition were selected for the test. Care was taken to select only those animals that had a normal eye condition and any with ocular lesions were rejected.

Analgesic and anaesthetic treatment
Sixty minutes (60 ±10 min) prior to test substance application, a systemic opiate analgesic was administered by subcutaneous injection (SC) under direct Veterinary supervision.Five minutes (5 ±1.5 min) prior to test substance application, a topical ocular anaesthetic was applied to each eye (including the control eye to ensure direct comparison of any ocular observations). Eight hours (8 to 9 hr) after test substance application, a systemic opiate analgesic and a nonsteroidal anti-inflammatory drug (NSAID) were administered by subcutaneous injection (SC) under direct Veterinary supervision. The systemic opiate analgesic was again injected ~12 hours after the post-treatment analgesic.
Systemic opiate analgesic: Buprenorphine 0.01 mg/kg.
Topical ocular anaesthetic: Humacain (oxybuprocaine) one-two drops per eye.
Nonsteroidal anti-inflammatory drug: Meloxicam 0.5 mg/kg.

Application of the Test Item
The test substance was placed in the conjunctival sac of the left eye of each animal after gently pulling the lower lid away from the eyeball. The lids were then gently held together for at least one second in order to prevent loss of the material.The test item was supplied as a fine dust therefore it was applied as supplied, without grinding.

OBSERVATIONS AND SCORING
Clinical Observations and Evaluation of Ocular Irritation
The eyes were examined at 1, 24, 48, 72 hours after treatment. The duration of the observation period was sufficient to identify reversibility or irreversibility of changes. Any clinical signs of toxicity or signs of ill-health during the study were recorded. At the end of the observation period, the animal was sacrificed by intramuscular injections of Ketamin 10% (Ketamidor) and Xylazin 2% (Primazin 2%) followed by i.v. pentobarbital sodium (Euthanimal 40%). Death was verified by checking pupil and corneal reflex and the absence of respiration.All rabbits were examined for distress at least twice daily, with observations at least 6 hours apart. Clinical observations or signs of ill-health were recorded.

Scoring and Assessment of Local Reaction
The eye irritation scores were evaluated according to the scoring system by Draize (1977) and OECD 405 (02 October 2012). See “Any other information” for scoring system.

Measurement of Body Weight
Individual body weight was recorded on the day of treatment and at the end of observation period.
Irritation parameter:
cornea opacity score
Basis:
mean
Time point:
24/48/72 h
Score:
0
Max. score:
8
Irritation parameter:
iris score
Basis:
mean
Time point:
24/48/72 h
Score:
0
Max. score:
2
Irritation parameter:
chemosis score
Basis:
mean
Time point:
24/48/72 h
Score:
0
Max. score:
4
Irritation parameter:
conjunctivae score
Basis:
mean
Time point:
24/48/72 h
Score:
0
Max. score:
6
Irritant / corrosive response data:
Examination of eye-irritancy
No Initial Pain Reaction (IPR) or any Pain Reaction (PR) was observed during the experimental period.

Animal 1 (No: 960) clinical observation: At one hour after the application, conjunctival redness (score 2), chemosis (score 2) and discharge (score 1) were noted in the rabbit. Test item remained in the eye sac.At 24, 48 and 72 hours after the application, no clinical signs, and no conjunctival or corneal effects were observed.

Animal 2 (No: 949) clinical observation: At one hour after the application, conjunctival redness (score 2), chemosis (score 2) and discharge (score 1) were noted in the rabbit. Test item remained in the eye sac.At 24, 48 and 72 hours after the application, no clinical signs, and no conjunctival or corneal effects were observed.

Animal 3 (No: 954) clinical observation: At one hour after the application, conjunctival redness (score 2), chemosis (score 2) and discharge (score 1) were noted in the rabbit. Test item remained in the eye sac.At 24, 48 and 72 hours after the application, no clinical signs, and no conjunctival or corneal effects were observed.
Other effects:
MORTALITY
There was no mortality observed during the study.

BODY WEIGHTS
The body weight of the animals was considered to be within the normal range of variability.

CLINICAL OBSERVATION
General daily examinationThere were no clinical signs observed that could be related to treatment.As no clinical signs were observed, the study was terminated after 72 hours observation.During the study, the control eye of each animal was symptom-free.The general state and behaviour of animals were normal throughout the study period.The animals’ individual mean scores (considering readings at 24, 48 and 72 hours after the treatment) are as detailed above.

INDIVIDUAL SCORES FOR OCULAR IRRITATION

Abbreviations:

R

=

Redness

OD

=

Opacity degree of density

CH

=

Chemosis

OE

=

Extent of opaque area

D

=

Discharge

IPR/PR

=

Initial or any pain reaction

0

=

Normal (in case of control eye and other lesions)

 

Animal No.: 960

Time

Score of irritation

IPR/PR

Other sign

Conjunctivae

Cornea

Iris

Control eye

R

CH

D

OD

OE

R

Pre-treatment

0

0

0

0

0

0

0

0

0

Post-treatment (h = hour)

1 h

2

2

1

0

0

0

0

0

*

24 h

0

0

0

0

0

0

0

0

0

48 h

0

0

0

0

0

0

0

0

0

72 h

0

0

0

0

0

0

0

0

0

* Test item remained in the eye sac

 

Animal No.: 949

Time

Score of irritation

IPR/PR

Other sign

Conjunctivae

Cornea

Iris

Control eye

R

CH

D

OD

OE

R

Pre-treatment

0

0

0

0

0

0

0

0

0

Post-treatment (h = hour)

1 h

2

2

1

0

0

0

0

0

*

24 h

0

0

0

0

0

0

0

0

0

48 h

0

0

0

0

0

0

0

0

0

72 h

0

0

0

0

0

0

0

0

0

* Test item remained in the eye sac

 

Animal No.: 954

Time

Score of irritation

IPR/PR

Other sign

Conjunctivae

Cornea

Iris

Control eye

R

CH

D

OD

OE

R

Pre-treatment

0

0

0

0

0

0

0

0

0

Post-treatment (h = hour)

1 h

2

2

1

0

0

0

0

0

*

24 h

0

0

0

0

0

0

0

0

0

48 h

0

0

0

0

0

0

0

0

0

72 h

0

0

0

0

0

0

0

0

0

* Test item remained in the eye sac

 

MEAN VALUES OF EYE IRRITATION (24, 48, 72 hour reading)

Animal Number

Sex

Cornea Opacity

Iris

Conjunctive

Redness

Chemosis

Discharge

960

Male

0.00

0.00

0.00

0.00

0.00

949

Male

0.00

0.00

0.00

0.00

0.00

954

Male

0.00

0.00

0.00

0.00

0.00

 

BODY WEIGHT DATA

Animal Number

Before treatment (g)

Before euthanasia (g)

Body weight gain (g)

960

2857

2901

44

949

3007

2156

149

954

3134

3255

121

 

Interpretation of results:
GHS criteria not met
Conclusions:
The test item Calcium hydrogen phosphonate, applied to rabbit eye mucosa, caused conjunctival effects at one hour after application which were fully reversible within 24 hours.According to Regulation (EC) No 1272/2008, Calcium hydrogen phosphonate does not require classification as an eye irritant.According to the UN Globally Harmonised System of Classification and Labelling of Chemicals, Calcium hydrogen phosphonatedoes not require classification as an eye irritant.
Executive summary:

An acute eye irritation study of the test item, Calcium hydrogen phosphonate was performed in New Zealand White rabbits. The irritation effects of the test item were evaluated according to the Draize method (OECD No.: 405, 2012). Rabbits were treated with analgesic and anaesthetic as per the regulatory guideline. Three animals were used to make the classification.

The test item was placed into the conjunctival sac of the left eye of each animal. The untreated right eye served as control. A single amount of 0.1 g of the test item was administered as a single dose.

 The eyes were examined at 1, 24, 48 and 72 hours after application.

 No Initial Pain Reaction (IPR) or any Pain Reaction (PR) was observed during the experimental period.

 

Animal 1 (No: 960) clinical observation

At one hour after the application, conjunctival redness (score 2), chemosis (score 2) and discharge (score 1) were noted in the rabbit. Test item remained in the eye sac.

At 24, 48 and 72 hours after the application, no clinical signs, and no conjunctival or corneal effects were observed.

 

Animal 2 (No: 949) clinical observation

At one hour after the application, conjunctival redness (score 2), chemosis (score 2) and discharge (score 1) were noted in the rabbit. Test item remained in the eye sac.

At 24, 48 and 72 hours after the application, no clinical signs, and no conjunctival or corneal effects were observed.

 

Animal 3 (No: 954) clinical observation

At one hour after the application, conjunctival redness (score 2), chemosis (score 2) and discharge (score 1) were noted in the rabbit. Test item remained in the eye sac.

At 24, 48 and 72 hours after the application, no clinical signs, and no conjunctival or corneal effects were observed.

 

As no clinical signs were observed, the experiment was terminated after 72 hours observation.

 During the experiment, the control eye of each animal was symptom-free.

 The general state and behaviour of animals were normal throughout the study period.

 No mortality occurred during the study. The bodyweights of all rabbits were considered to be within the normal range of variability.

 The animals’ individual mean scores (considering readings at 24, 48 and 72 hours after the treatment) were as follows:

Animal 1             Animal 2             Animal 3

Chemosis          0.00                      0.00                      0.00

Discharge          0.00                      0.00                     0.00

Redness            0.00                      0.00                      0.00

Cornea              0.00                      0.00                      0.00

Iris                     0.00                      0.00                      0.00

 

The test item, Calcium hydrogen phosphonate, applied to rabbit eye mucosa, caused conjunctival effects at one hour after application which were fully reversible within 24 hours.

 According to Regulation (EC) No 1272/2008, Calcium hydrogen phosphonate does not require classification as an eye irritant.

 According to the UN Globally Harmonised System of Classification and Labelling of Chemicals, Calcium hydrogen phosphonate does not require classification as an eye irritant.

Endpoint conclusion
Endpoint conclusion:
no adverse effect observed (not irritating)

Respiratory irritation

Endpoint conclusion
Endpoint conclusion:
no study available

Additional information

In vitro skin irritation

The objective of this study was to evaluate the skin irritation potential of calcium hydrogen phosphonate using the Episkin reconstructed human epidermis model. The study design was based upon international guidelines (OECD Guideline No. 439 and Commission Regulation (EC) No. 761/2009, B.46).

Preliminary tests were performed to detect the ability of the test item to directly reduce MTT as well as its colouring potential. Following the preliminary tests, the skin irritation potential of the test item was tested in the main test. The test item and both the negative and positive controls were applied topically on triplicate tissues and incubated at room temperature for 42 minutes. At the end of the treatment period, each tissue was rinsed and incubated for 42 hours at 37.5°C, 5% CO2 in a humidified incubator. The cell viability was then assessed by means of the colourimetric MTT reduction assay.

Relative viability values were calculated for each tissue and expressed as a percentage of the mean viability of the negative control tissues which was set at 100% (reference viability).

In addition, the concentration of the inflammatory mediator IL-1awas evaluated in the culture medium retained following the 42-hour recovery period. This quantification, based on an ELISA assay, was performed since the mean relative viability of the test item-treated tissues was > 50% following the MTT reduction assay.

In the preliminary tests, the test item was found not to have direct MTT reducing properties or colouring potential.

 In the main test, all acceptance criteria for the negative and positive controls were fulfilled except that the standard deviation value of viability percent of the tissues treated with the negative control was 37.4% instead of 18% (maximal value) as initially scheduled in the study plan. Considering the study results obtained in the tissues treated with the test item and the historical data, this deviation is considered as without impact on the conclusion of the study. Indeed, the results of the tissues treated with the test item are homogenous and all markedly above the cut-off of 50%. The study was therefore considered to be valid.

Following a 42 minutes exposure and 42 hours of recovery period, the relative mean viability of the tissues treated with the test item was 118% (considered 100%) with a standard deviation of 13%.

As the mean viability was > 50% after the MTT reduction, the IL-1aconcentrations in culture media samples retained were analyzed by ELISA

The mean IL-1aconcentration for treated tissues was 7.08 pg/mL vs 169.54 pg/mL in the positive control. Due to this value being below 60 pg/mL, the results met the criteria for an in vitro classification as non-irritant to skin.

 Under the experimental conditions of this study, the test item is considered to be non-irritant to skin.

According to the results of this study, the classification of the test item should be:

.  not classified (Directive 67/548/EEC) and no category (Regulation (EC) No. 1272/2008).

In vitro eye irritation

The potential irritant and corrosive properties of calcium hydrogen phosphonate to the eye was assessed following the Isolated Chicken Eye method that can identify chemicals inducing serious eye damage and chemicals not requiring classification for eye irritation or serious eye damage. The design of this study was based on the guideline OECD Guideline 438 and the study was performed in compliance with the OECD Principles of Good Laboratory Practice.

The ocular reactions observed in eyes treated with the test item were:

- maximal mean score of corneal opacity : 0.5, corresponding to ICE class I;

- mean score of fluorescein retention: 2.7, corresponding to ICE class IV;

- maximal mean corneal swelling: +1% at 75 minutes post dose, corresponding to ICE class I.

Therefore the combination of the three endpoints for the test item calcium hydrogen phosphonate was 1 x IV, 2 x 1. The combination of the three endpoints for the positive control, Sodium hydroxide, lead to a classification "Corrosive/Severe Irritant", as expected and the combination of the three endpoints for the negative control, physiological saline, to a "No Category" classification, as expected.

In accordance with the Regulation (EC) No. 1272/2008, the results obtained under these experimental conditions do not enable to predict the classification of the test item calcium hydrogen phosphonate.

Therefore an acute eye irritation study with calcium hydrogen phosphonate was performed in New Zealand White rabbits. The irritation effects of the test item were evaluated according to the Draize method (OECD No.: 405, 2012). Rabbits were treated with analgesic and anaesthetic as per the regulatory guideline. Three animals were used to make the classification.

The test item was placed into the conjunctival sac of the left eye of each animal. The untreated right eye served as control. A single amount of 0.1 g of the test item was administered as a single dose.

 The eyes were examined at 1, 24, 48 and 72 hours after application.

 No Initial Pain Reaction (IPR) or any Pain Reaction (PR) was observed during the experimental period.

 Animal 1 clinical observation

At one hour after the application, conjunctival redness (score 2), chemosis (score 2) and discharge (score 1) were noted in the rabbit. Test item remained in the eye sac.

At 24, 48 and 72 hours after the application, no clinical signs, and no conjunctival or corneal effects were observed.

 Animal 2 clinical observation

At one hour after the application, conjunctival redness (score 2), chemosis (score 2) and discharge (score 1) were noted in the rabbit. Test item remained in the eye sac.

At 24, 48 and 72 hours after the application, no clinical signs, and no conjunctival or corneal effects were observed.

 Animal 3 clinical observation

At one hour after the application, conjunctival redness (score 2), chemosis (score 2) and discharge (score 1) were noted in the rabbit. Test item remained in the eye sac.

At 24, 48 and 72 hours after the application, no clinical signs, and no conjunctival or corneal effects were observed.

 As no clinical signs were observed, the experiment was terminated after 72 hours observation.

During the experiment, the control eye of each animal was symptom-free.

The test item, Calcium hydrogen phosphonate, applied to rabbit eye mucosa, caused conjunctival effects at one hour after application which were fully reversible within 24 hours.

According to Regulation (EC) No 1272/2008, Calcium hydrogen phosphonate does not require classification as an eye irritant.

Justification for classification or non-classification

According to Regulation (EC) No 1272/2008, Calcium hydrogen phosphonate does not require classification as a skin or eye irritant.

According to the UN Globally Harmonised System of Classification and Labelling of Chemicals, Calcium hydrogen phosphonate does not require classification as a skin or eye irritant.