Ammonium iron(III) citrate

Endpoint summary

Description of key information

Skin irritation:

The dermal irritation potential of target chemical was assessed in various in-vivo experimental studies.Based on the available studies,it can be concluded that the test chemical is unable to cause skin irritation and considered as not irritating. Comparing the above annotations with the criteria of CLP regulation, it can be classified under the category “Not Classified''.

Eye irritation:

The ocular irritation potential of target chemical was assessed in various in-vivo experimental studies.Based on the available studies,it can be concluded that the test chemical is unable to cause eye irritation and considered as not irritating. Comparing the above annotations with the criteria of CLP regulation, it can be classified under the category “Not Classified''.

Key value for chemical safety assessment

Skin irritation / corrosion

Link to relevant study records

Reference

Endpoint:

skin irritation: in vivo

Type of information:

experimental study

Adequacy of study:

key study

Reliability:

1 (reliable without restriction)

Rationale for reliability incl. deficiencies:

comparable to guideline study

Justification for type of information:

Data is from experimental study report.

Qualifier:

according to guideline

Guideline:

OECD Guideline 404 (Acute Dermal Irritation / Corrosion)

Qualifier:

according to guideline

Guideline:

EU Method B.4 (Acute Toxicity: Dermal Irritation / Corrosion)

Principles of method if other than guideline:

The purpose of the study was to evaluate the acute dermal irritation and/or corrosive effects on skin following the application of the test article.

GLP compliance:

no

Species:

rabbit

Strain:

New Zealand White

Details on test animals or test system and environmental conditions:

Details on test animal
TEST ANIMALS
- Source: Conelli S.n,c., Via Milano. 61 - 28041 ARONA (Novara - Italy) and received on January 7, 1998 (shipping slip No. 00001, dated Itinerary 7. 1998).
- Age at study initiation: 2 - 3 months old
- Weight at study initiation: 2.5 - 2.8kg.
- Fasting period before study:
- Housing: housed in room T05C.
- Diet (e.g. ad libitum): The animals were fed a diet coded "2 RB 15 GLP Certificate" produced by the Charles River ltalia's feed licensec Mucedola S.r.l., Seflimo Milanese. ad libitum
- Water (e.g. ad libitum): municipal water, ad libitum
- Acclimation period: Not specified

ENVIRONMENTAL CONDITIONS
- Temperature (°C): 20°C±2
- Humidity (%):50%±15
- Air changes (per hr): approximately 20 air changes per hour (filtered on HEPA 99.97%).
- Photoperiod (hrs dark / hrs light): The room was illuminated by artificial lighting with a 12-hour circadian cycle (7 a.m. - 7 p.m.).


IN-LIFE DATES: From: To: March 2- March 5, 1998

Type of coverage:

occlusive

Preparation of test site:

clipped

Remarks:

intact

Vehicle:

not specified

Controls:

yes

Amount / concentration applied:

0.5 ml/site

Duration of treatment / exposure:

3 minutes, 1 hour and 4 hours

Observation period:

Observations were performed immediately and 72 hours after the 3-minute and the 1-hour exposure period in the first rabbit and at 1, 24. 48 and 72 hours after the 4-hour exposure period (all rabbits).

Number of animals:

3 female

Details on study design:

Details on study design
TEST SITE
- Area of exposure: the trunk
- % coverage: approximately 6 cm2
- Type of wrap if used: The application area was covered with a gauze patch loosely held in contact with the skin by an impermeable, non-irritant, hypoallergenic tape (occlusive patch).

REMOVAL OF TEST SUBSTANCE
- Washing (if done): residual substance was removed with water
- Time after start of exposure: 1, 24. 48 and 72 hours after

Irritation parameter:

overall irritation score

Basis:

mean

Time point:

other: 3 minutes, 1 hour and 4 hours (for single rabbit ) and after 1, 24. 48 and 72 hours (all rabbits)

Score:

0

Max. score:

4

Reversibility:

other: Not applicable

Remarks on result:

no indication of irritation

Irritant / corrosive response data:

No changes were seen at any test article application site. No clinical signs, either general or local (at the application sites), were noted in any rabbit.

Other effects:

not specified

Table 1:

Grading of Skin Reaction

1.           Erythema and Eschar Formation                No erythema....................................................................................... 0                Very slight erythema (barely perceptible)............................................. 1                Well defined erythema........................................................................ 2                Moderate to severe erythema.............................................................3 Severe erythema (beet redness) to slight                Eschar formation (injuries in depth)..................................................... 4                Maximum possible = 4

2.           edema Formation                No oedema......................................................................................... 0                Very slight edema (barely perceptible)................................................. 1                Slight edema (edges of area well defined by definite raising)................. 2                Moderate edema (raised approximately 1 mm...................................... 3                Severe edema (raised more than 1 mm and extending                Beyond area of exposure)....................................................................4                 Maximum possible = 4

Evaluation of results:  the dermal irritation was evaluated in conjunction with the nature and reversibility or otherwiseofthe responsesobserved.

RESULTS:

Table 2: Observation of single rabbit

Exposure period: 3 minutes

Erythema And Eschar

Obs. wade at	42F (animal No.)
3minutes	0
72 hours	0

Edema

3 minutes	0
72 hours	0

Exposure period: 1hour

ErythemaAnd Eschar

Obs. made at	42F (animal No.)
1 hour	0
72 hours	0

Edema

1 hour	0
72 hours	0

Table 3: obervation of all rabbits

Exposure period:4 hours

Erythema And Eschar

Obs. made at	42F	46F	48F
60 minutes	0	0	0
24 hours	0	0	0
48 hours	0	0	0
72 hours	0	0	0

Edema

Obs. made at	42F	46F	48F
60 minutes	0	0	0
24 hours	0	0	0
48 hours	0	0	0
72 hours	0	0	0

Interpretation of results:

other: Not irritating

Conclusions:

Neither mortality nor adverse general clinical modifications were seen during the study. Thus, the test material was considered to be non-irritant when administrated to rabbits by dermal route.

Executive summary:

An acute dermal irritation study of test chemical was conducted in the female New Zealand White rabbit (3 animals) in accordance with EEC Guidelines (B.4) and OECD Guidelines (404).

 

0.5 ml of the test article were applied on the intact skin of the trunk to a small area (approximately 6 cm2) of skin. The application area was covered with a gauze patch loosely held in contact with the skin by an impermeable, non-irritant, hypoallergenic tape (occlusive patch).  At the end of each exposure period, residual substance was removed with water. Adjacent areas of untreated skin of each animal served as control for the test.

 

Observations were performed immediately and 72 hours after the 3-minute and the 1-hour exposure period in the first rabbit and at 1, 24. 48 and 72 hours after the 4-hour exposure period (all rabbits). Inspections for mortality and general clinical signs were made once a day.

 

No changes were seen at any test article application site. No clinical signs, either general or local (at the application sites), were noted in any rabbit. Thus, the test material was considered to be non-irritant when administrated to rabbits by dermal route.

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

Reliability:

1 (reliable without restriction)

Rationale for reliability incl. deficiencies:

comparable to guideline study

Justification for type of information:

Data is from experimental study report.

Qualifier:

according to guideline

Guideline:

other: the OECD [C(81) 30 (final)]

Principles of method if other than guideline:

The purpose of the study was to evaluate the acute eye irritation and/or corrosive effects on the eye following the application of the test article.

GLP compliance:

no

Species:

rabbit

Strain:

New Zealand White

Details on test animals or tissues and environmental conditions:

Details on test animal
TEST ANIMALS
- Source: Conelli S.n,c., Via Milano. 61 - 28041 ARONA (Novara - Italy) and received on January 7,1998 (shipping slip No. 00001, dated Itinerary 7,1998).
- Age at study initiation: 2 - 3 months old
- Weight at study initiation: 2.5 - 2.8 kg.
- Fasting period before study:Not specified
- Housing: housed in room T05C
- Diet (e.g. ad libitum): The animals were fed a diet coded "2 RB 15 GLP Certificate" produced by the Charles River ltalia's feed licensec Mucedola S.r.l., Seflimo Milanese. ad libitum.
- Water (e.g. ad libitum): municipal water, ad libitum
- Acclimation period: Not specified

ENVIRONMENTAL CONDITIONS
- Temperature (°C): 20°C ± 2
- Humidity (%):50% ± 15
- Air changes (per hr): approximately 20 air changes per hour (filtered on HEPA 99.97%).
- Photoperiod (hrs dark / hrs light): The room was illuminated by artificial lighting with a 12-hour circadian cycle (7 a.m. - 7 p.m.).


IN-LIFE DATES: From: To: March 3 - March 6,1998

Vehicle:

not specified

Controls:

yes

Amount / concentration applied:

0.1 ml / animal

Duration of treatment / exposure:

single dose

Observation period (in vivo):

1, 24, 48 and 72 hours after the test article application.

Number of animals or in vitro replicates:

3 males

Details on study design:

Details on study design
TEST SITE
- Area of exposure: the conjunctival sac of the right eye of each animal


REMOVAL OF TEST SUBSTANCE
- Washing (if done): At 24 hours a washout was performed.
- Time after start of exposure: 1, 24, 48 and 72 hours

SCORING SYSTEM: as mentioned in table 1

TOOL USED TO ASSESS SCORE: After the 24-hour reading, the cornea was examined after instillation of one drop of 1% sodium fluorescein and successive washing out with sterile saline solution.

other : Both eyes of each experimental animal selected for the study were examined within 24 hours of testing. Only animals without eye irritation, ocular defects or pre-existing corneal injury were used.

Irritation parameter:

conjunctivae score

Basis:

mean

Time point:

24/48/72 h

Score:

0

Max. score:

3

Reversibility:

not specified

Remarks on result:

no indication of irritation

Irritation parameter:

chemosis score

Basis:

mean

Time point:

24/48/72 h

Score:

0

Max. score:

4

Reversibility:

other: Not applicable

Remarks on result:

no indication of irritation

Irritation parameter:

iris score

Basis:

mean

Time point:

24/48/72 h

Score:

0

Max. score:

2

Reversibility:

other: Not applicable

Remarks on result:

no indication of irritation

Irritation parameter:

cornea opacity score

Basis:

mean

Time point:

24/48/72 h

Score:

0

Max. score:

4

Reversibility:

other: Not applicable

Remarks on result:

no indication of irritation

Irritant / corrosive response data:

Slight redness of the conjunctivae (grade 1) was the only finding in all rabbits at the observation carried out 1 hour after treatment. No signs of ocular irritancy were subsequently observed. Negative results were obtained at the fluorescein staining performed 24 hours after the test article application .

Other effects:

Neither mortality nor adverse general clinical modifications were seen during the study.

Interpretation of results:

other: Not irritating

Conclusions:

Negative results were obtained at the fluorescein staining performed 24 hours after the test article application. Neither mortality nor adverse general clinical modifications were seen during the study. Thus, the test was considered to be non-irritant when administered to New Zealand White rabbits as a single ocular application.

Executive summary:

An acute ocular irritation study of test chemical was conducted in the male New Zealand White rabbit (3 animals) in accordance with the OECD [C(81) 30 (final)].

Both eyes of each experimental animal selected for the study were examined within 24 hours of testing. Only animals without eye irritation, ocular defects or pre-existing corneal injury were used.

About 0.1 ml of the test article was placed in the conjunctival sac of the right eye of each animal, after gently pulling the lower lid away from the eyeball. The lids were then gently held together for about 3-4 seconds in order to prevent loss of the material. The other eye (the left eye), remaining untreated, served as control. At 24 hours a washout was performed.

Observations were performed at 1, 24, 48 and 72 hours after the test article application. After the 24-hour reading the cornea was examined after instillation of one drop of 1% sodium fluorescein and successive washing out with sterile saline solution. Inspections for mortality and general clinical signs were made once a day.

Slight redness of the conjunctivae (grade 1) was the only finding in all rabbits at the observation carried out 1 hour after treatment. No signs of ocular irritancy were subsequently observed.

Negative results were obtained at the fluorescein staining performed 24 hours after the test article application. Neither mortality nor adverse general clinical modifications were seen during the study. Thus, the test was considered to be non-irritant,when administered to New Zealand White rabbits as a single ocular application.

 

Endpoint conclusion

Endpoint conclusion:

no adverse effect observed (not irritating)

Respiratory irritation

Endpoint conclusion

Endpoint conclusion:

no study available

Additional information

Skin Irritation:

In different studies, the test chemical has been investigated for potential for dermal irritation to a greater or lesser extent. The studies are based on in-vivo experimental studies conducted in different subjects which have been summarized as below:

An acute dermal irritation study of test chemical was conducted in the female New Zealand White rabbit (3 animals) in accordance with EEC Guidelines (B.4) and OECD Guidelines (404). 0.5 ml of the test article were applied on the intact skin of the trunk to a small area (approximately 6 cm2) of skin. The application area was covered with a gauze patch loosely held in contact with the skin by an impermeable, non-irritant, hypoallergenic tape (occlusive patch).  At the end of each exposure period, residual substance was removed with water. Adjacent areas of untreated skin of each animal served as control for the test.Observations were performed immediately and 72 hours after the 3-minute and the 1-hour exposure period in the first rabbit and at 1, 24. 48 and 72 hours after the 4-hour exposure period (all rabbits). Inspections for mortality and general clinical signs were made once a day.No changes were seen at any test article application site. No clinical signs, either general or local (at the application sites), were noted in any rabbit. Thus, the test material was considered to be non-irritant when administrated to rabbits by dermal route.

Moreover in another acute skin irritation study which was performed on rabbits to assess the degree of irritation caused by the test chemical. 6 New Zealand White rabbits were used for the study. Animals are held for 14 days after arrival in the facility but prior to use in tests to ensure that they are healthy. Four intact sites of administration , 2 on each side of rabbits back were assigned a code number:

#1:Test substance

#2: Negative control

#3: Positive control

#4: Vehicle control (if required)

A positive control substance (a known skin irritant, 1 % sodium lauryl sulfate in water) and a negative control (plain gauze patch) were applied to the skin along with the test chemical. Since the test chemical was moistened with physiological saline, a vehicle control patch was also applied along with the test, control patches. This vehicle control patch consists of plain gauze patch moistened with vehicle.Each test or control substance was held in place with a 1 x 1 inch2 12-ply surgical gauze patch. The gauze patch is applied to the appropriate skin site and secured with 1-inch wide strips of Blenderm surgical tape (3M Company, Medical Products Division), at the four edges, leaving the center of the gauze patch non-occluded.0.5-g portion was weighed and placed on the gauze patch prior to application of the patch. The test substance and patch are then placed on the appropriate skin site and secured. The patch is subsequently moistened with 0.5 ml physiological saline.The entire trunk of the animal was covered with an impervious material (Saran Wrap) (Dow Chemical Co., IN) for a 24-hr period of exposure. The Saran Wrap was secured by wrapping several long strips of athletic adhesive tape (Quick Strap Tape, Acme Cotton Products, NY) around the trunk of the animal.A Saf-T-Shield (HAY International, Glendale, CA) collar was fitted and fastened around the neck of each test animal. The collar remains in place for a 24-hr exposure period. The collar prevents the animal from removing the wrapping and patches, while allowing it access to food and water ad libitum. Dermal irritation scores for erythema and edema formation are evaluated by the scale proposed by Draize. If irritation persists for 72 hr post application, dermal irritation scores will be continued until all irritation produced by the substance (test site) resolves or until day 14 post application. Persistent irritation scores are also recorded, as well as the occurrence of eschar (+E) and/or necrosis (+N). The primary dermal irritation index (PDII) was then calculated by dividing the sum of total irritation scores by the number of observations, and classifying the test

substance:

PDII of

0.00 = Non-irritant;

> 0.00-0.50 = negligible irritant;

> 0.50-2.00 = mild irritant;

> 2.00-5.00 = moderate irritant;

> 5.00-8.00 = severe (primary) irritant.

  The test chemical when applied onto the intact skin of 6 New Zealand white rabbits at the concentration 0.5 g in 0.5 ml saline produced mild skin irritation effects along with the PDII score was 1.8. Moreover, the study considered two classification system ,i.e. primary dermal irritation (PDI) classifications and the Federal Hazardous Substances Act (FHSA). As the test chemical induced mild skin effects which was not enough to classify the chemical as positive and also considering FHSA classification, the test chemical was considered to be non-irritant to rabbit skin.

All these studies lead to a conclusion that Test chemical is indeed not irritating to skin. Hence, comparing the above annotations with the criteria of CLP regulation, Test chemical can be classified under the category “Not Classified”.

 

Eye Irritation:

In different studies, the test chemical has been investigated for potential for ocular irritation to a greater or lesser extent. The studies are based on in-vivo experimental conducted in rabbits conducted which have been summarized as below:

An acute ocular irritation study of test chemical was conducted in the male New Zealand White rabbit (3 animals) in accordance with the OECD [C(81) 30 (final)]. Both eyes of each experimental animal selected for the study were examined within 24 hours of testing. Only animals without eye irritation, ocular defects or pre-existing corneal injury were used. About 0.1 ml of the test article was placed in the conjunctival sac of the right eye of each animal, after gently pulling the lower lid away from the eyeball. The lids were then gently held together for about 3-4 seconds in order to prevent loss of the material. The other eye (the left eye), remaining untreated, served as control. At 24 hours a washout was performed. Observations were performed at 1, 24, 48 and 72 hours after the test article application. After the 24-hour reading the cornea was examined after instillation of one drop of 1% sodium fluorescein and successive washing out with sterile saline solution. Inspections for mortality and general clinical signs were made once a day. Slight redness of the conjunctivae (grade 1) was the only finding in all rabbits at the observation carried out 1 hour after treatment. No signs of ocular irritancy were subsequently observed. Negative results were obtained at the fluorescein staining performed 24 hours after the test article application. Neither mortality nor adverse general clinical modifications were seen during the study. Thus, the test was considered to be non-irritant,when administered to New Zealand White rabbits as a single ocular application.

Moreover in another ocular irritation study,in which potential of the test chemical was evaluated in rabbits. The study was performed according to Code of Federal Regulations, Title 16: Subchapter C-Federal Hazardous Substances Act. Part 1500.42, Revised January 1, 1981 (test for eye irritants) Guidelines. 9 New Zealand White rabbits were used for the study. The rabbits were acclimatized for 14 days prior to test initiation.One hour prior to instillation of the test substance, both eyes of each of at least 12 rabbits was examined for signs of irritation and corneal defects with a handheld slit lamp (Kowa SL-5). Then all eyes are stained with a 2.0% sodium fluorescein and examined to confirm the absence of corneal lesions. The overall study design uses nine rabbits.The contra lateral eye remained untreated and served as control. The treated eyes of the three rabbits from 9 were irrigated for1min with room temperature tap water using a polyethylene squeeze bottle, starting 20 sec after installation. At least 1 hr after fluorescein staining, the test substance was placed on one eye of each animal by gently pulling the lower lid away from the eyeball to form a cup (conjunctival cul-de-sac) into which the test material was dropped. The eyes are observed for any immediate signs of discomfort after instilling the test substance, with blepharospasm and/or excessive tearing being taken as indications of irritating sensations caused by the test substance. Grading and scoring of ocular irritation are performed in accordance with the standard Draize grades for ocular lesions. The eyes are examined and grades of ocular reactions were recorded at 1 hr, 1, 2, 3, 4, and 7 days after exposure. After all observations were recorded and photographed 1 day after exposure, all corneas are further examined with the aid of sodium fluorescein.The mean Draize scores after 24 hours post-instillation was 0.0. Based on the readings and classification criteria mentioned the test chemical can be considered to be not irritating to eyes.

All these studies lead to a conclusion that Test chemical is indeed not irritating to eye. Hence, comparing the above annotations with the criteria of CLP regulation, Test chemical can be classified under the category “Not Classified”.

Justification for classification or non-classification

The skin and eye irritation potential of test chemical was observed in various studies. The results obtained from these studies indicate that the chemical is not likely to cause skin and eye irritation. Hence, the test chemical can be classified under the category “Not Classified” for skin and eye irritation as per CLP.