6-Amino-5-chloro-2-cyclopyrimidine-4-carboxylic acid

Endpoint summary

Description of key information

Skin irritation (OECD 404, GLP), rabbit: not irritating

 Eye irritation (OECD 405, GLP), rabbit: not irritating

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

Study period:

6 - 16 March 2007

Reliability:

1 (reliable without restriction)

Rationale for reliability incl. deficiencies:

guideline study

Qualifier:

according to guideline

Guideline:

OECD Guideline 404 (Acute Dermal Irritation / Corrosion)

Version / remarks:

adopted: 2002

Deviations:

no

GLP compliance:

yes

Species:

rabbit

Strain:

New Zealand White

Details on test animals or test system and environmental conditions:

TEST ANIMALS
- Source: Covance Research Products, Denver, Pennsylvania, USA
- Age at study initiation: adult
- Weight at study initiation: 3830 g, 3024 g and 2979 g
- Housing: individual housed in stainless steel wire-mesh cages suspended above cage boards
- Diet: approximately 125 g of PMI® Nutrition International, LLC Certified Rabbit LabDiet® 5322 daily during the study
- Water: ad libitum
- Acclimation period: at least 7 days

ENVIRONMENTAL CONDITIONS
- Temperature (°C): 16 - 22
- Humidity (%): 30 - 70
- Photoperiod (hrs dark / hrs light): 12/12

Type of coverage:

semiocclusive

Preparation of test site:

shaved

Vehicle:

unchanged (no vehicle)

Controls:

not required

Amount / concentration applied:

TEST MATERIAL
- Amount applied: 0.5 g moistened with 0.4 mL deionized water (a thick paste was formed)

Duration of treatment / exposure:

4 h

Observation period:

Reading time points: 1, 24, 48 and 72 h

Number of animals:

1 male (initial test)
2 males (confirmatory test)

Details on study design:

TEST SITE
- Area of exposure: 6 cm² on the scapular to the lumbar region of the back
- Type of wrap if used: The treated skin was covered with a 2-ply gauze pad and held in place with with non-irritating tape. The trunk of each rabbit was wrapped with porous tape. The tape was further secured with waterproof tape.

REMOVAL OF TEST SUBSTANCE
- Washing (if done): The test sites were gently washed with warm water and gently patted dry.
- Time after start of exposure: 4 h

OBSERVATION TIME POINTS
1, 24, 48 and 72 h (The rabbit that was initially treated was also examined immediately after test substance removal. )

SCORING SYSTEM:
- Method of calculation: Draize scoring system

Irritation parameter:

erythema score

Basis:

mean

Remarks:

of 3 animals

Time point:

24/48/72 h

Score:

0

Max. score:

4

Reversibility:

other:

Remarks:

reversibility: not applicable

Irritation parameter:

edema score

Basis:

mean

Remarks:

of 3 animals

Time point:

24/48/72 h

Score:

0

Max. score:

4

Reversibility:

other:

Remarks:

reversibility: not applicable

Irritant / corrosive response data:

Erythema (score of 2 and 1) but no edema was observed on the test site of the first rabbit at 0 minutes and 60 minutes after test substance removal, respectively. At later time points, no skin reactions were observed. The remaining animals did not show signs indicative for dermal irritation during the study.

Other effects:

- Other adverse systemic effects: No clinical signs of toxicity were observed and no body weight loss occurred.

Table 1: Results of skin irritation study

Observation time	Rabbit no.
	1 *		2		3
	Erythema	Edema	Erythema	Edema	Erythema	Edema
0 h (Immediately after exposure)	2	0	-	-	-	-
1 h	1	0	0	0	0	0
24 h	0	0	0	0	0	0
48 h	0	0	0	0	0	0
72 h	0	0	0	0	0	0
Mean value 0 + 1 + 24 + 48 + 72 h	0.6	0.00	0.00	0.00	0.00	0.00
Mean value 24 + 48 + 72 h	0.00	0.00	0.00	0.00	0.00	0.00

*Initial rabbit tested

Table 2: Body weight

Rabbit number	sex	Initial weight (g)	Final weight (g)
1	male	2830	2834
2	male	3024	3053
3	male	2976	3046

Interpretation of results:

other: CLP/EU GHS criteria not met, no classification required according to Regulation (EC) No 1272/2008

Conclusions:

A reliable study conducted in accordance with OECD 404 and GLP, found the test material to be non-irritating to the skin of rabbits. Mild erythema were only observed in one animal immediately after the exposure period and 1h after exposure, but were fully reversible within 24 h. No oedema were reported at any time point.

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:

19 - 29 March 2007

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:

adopted: 2002

Deviations:

yes

Remarks:

no use of topical anesthetics and systemic analgesics

GLP compliance:

yes

Species:

rabbit

Strain:

New Zealand White

Details on test animals or tissues and environmental conditions:

TEST ANIMALS
- Source: Covance Research Products, Denver, Pennsylvania, USA
- Age at study initiation: adult
- Weight at study initiation: 3267 g, 3524 g and 3610 g
- Housing: individual housed in stainless steel wire-mesh cages suspended above cage boards
- Diet: approximately 125 g of PMI® Nutrition International, LLC Certified Rabbit LabDiet® 5322 daily during the study
- Water: ad libitum
- Acclimation period: at least 7 days

ENVIRONMENTAL CONDITIONS
- Temperature (°C): 16 - 22
- Humidity (%): 30 - 70
- Photoperiod (hrs dark / hrs light): 12/12

Vehicle:

unchanged (no vehicle)

Controls:

yes, concurrent no treatment

Amount / concentration applied:

TEST MATERIAL
- Amount applied: approximately 86 mg (a weight equal to a 0.1 mL volume)

Duration of treatment / exposure:

The treated and control eyes of all rabbits remained unwashed.

Observation period (in vivo):

1, 24, 48 and 72 h

Number of animals or in vitro replicates:

3

Details on study design:

REMOVAL OF TEST SUBSTANCE
- Washing: unwashed

SCORING SYSTEM: Draize Scale

TOOL USED TO ASSESS SCORE: illumination, magnification, fluorescein

Irritation parameter:

cornea opacity score

Basis:

mean

Remarks:

of 3 animals

Time point:

24/48/72 h

Score:

0

Max. score:

4

Reversibility:

other:

Remarks:

reversibility: not applicable

Irritation parameter:

iris score

Basis:

animal #1

Time point:

24/48/72 h

Score:

0

Max. score:

2

Reversibility:

other:

Remarks:

reversibility: not applicable

Irritation parameter:

iris score

Basis:

animal #2

Time point:

24/48/72 h

Score:

0.67

Max. score:

2

Reversibility:

fully reversible within:

Remarks:

72 h

Irritation parameter:

iris score

Basis:

animal #3

Time point:

24/48/72 h

Score:

0

Max. score:

2

Reversibility:

other:

Remarks:

reversibility: not applicable

Irritation parameter:

conjunctivae score

Basis:

animal #1

Time point:

24/48/72 h

Score:

1

Max. score:

3

Reversibility:

fully reversible within:

Remarks:

72 h

Irritation parameter:

conjunctivae score

Basis:

animal #2

Time point:

24/48/72 h

Score:

0.67

Max. score:

3

Reversibility:

fully reversible within: 72 h

Irritation parameter:

conjunctivae score

Basis:

animal #3

Time point:

24/48/72 h

Score:

0.67

Max. score:

3

Reversibility:

fully reversible within:

Remarks:

72 h

Irritation parameter:

chemosis score

Basis:

animal #1

Time point:

24/48/72 h

Score:

0.33

Max. score:

4

Reversibility:

fully reversible within:

Remarks:

48 h

Irritation parameter:

chemosis score

Basis:

animal #2

Time point:

24/48/72 h

Score:

0

Max. score:

4

Reversibility:

other:

Remarks:

reversibility: not applicable

Irritation parameter:

chemosis score

Basis:

animal #3

Time point:

24/48/72 h

Score:

0

Max. score:

4

Reversibility:

other:

Remarks:

reversibility: not applicable

Irritant / corrosive response data:

Additional time point 1h (for further details please refer to table 1 in the "Any other information on results incl. tables):
Cornea opacity: mean / of 3 animals was 0.
Iris score: mean / of 3 animals was 0.33.
Conjuctivae score: mean / of 3 animals 2.
Chemosis score: mean / of 3 animals 1.33.

Other effects:

- Lesions and clinical observations: Fluorescein stain examinations were negative for corneal injury in the treated eyes of all rabbits. No clinical signs of toxicity were observed, and no body weight loss occurred.
- Other observations: One of the rabbits pawed the treated eye after instillation of the test substance.

Table 1: Summary Occular Irritation Scores

	Rabbitt	Corneal			Conjunctiva			Fluorescein
Reading	Number	Opacity	Area	Iritis	Redness	Chemosis	Discharge	Stain
1 h	44a	0	0	0	2	2	0	NA
	169	0	0	1	2	1	0	NA
	170	0	0	0	2	1	0	NA
24 h	144a	0	0	0	2	1	0	Negative
	169	0	0	1	2	0	0	Negative
	170	0	0	0	1	0	0	Negative
48 h	144a	0	0	0	1	0	0	Negative
	169	0	0	1	0	0	0	Negative
	170	0	0	0	1	0	0	Negative
72 h	144a	0	0	0	0	0	0	Negative
	169	0	0	0	0	0	0	Negative
	170	0	0	0	0	0	0	Negative

NA = not applicable

a)    Initial animal tested

 

Table 2: Mean Scores for Ocular Responses for Individual Rabbits (24/48/72 h)

Rabbit number	Corneal opacity	Iritis	Conjunctival redness	Conjunctival chemosis
144	0	0	1	0.33
169	0	0.67	0.67	0
170	0	0	0.67	0

 

Table 3: Body weight

Rabbit number	sex	Initial weight (g)	Final weight (g)
144	male	3167	3260
169	male	3524	3560
170	male	3610	3658

Interpretation of results:

other: CLP/EU GHS criteria not met, no classification required according to Regulation(EC) No. 1272/2008

Conclusions:

Under the conditions of the acute eye irritation/corrosion test, the test substance did not show irritant properties towards the eyes.
CLP: not classified

Endpoint conclusion

Endpoint conclusion:

no adverse effect observed (not irritating)

Additional information

Skin irritation

In the available skin irritation study performed according to OECD TG 404 and in compliance with GLP three male albino rabbits were exposed to 500 mg test substance moistened with deionized water onto shaved skin for 4 h under semi-occlusive conditions (2011 d). Skin reactions were evaluated according to the Draize scoring system 1, 24, 48 and 72 h post-application. One rabbit revealed erythema (score of 2 and 1), 0 minutes and 60 minutes after test substance removal, respectively. No further skin reactions were observed at later time points in this animal. The remaining animals revealed no erythema or edema at any observation point. No further local or systemic effects were apparent in any animal during the study period. Based on the study results the test substance is not considered to be irritating to the skin.

 

In addition to the skin irritation study, an acute dermal toxicity study and a repeated dose toxicity study following dermal application in rats are available.

 

In the acute dermal toxicity study (performed according to OECD TG 402), the test substance was administered at a dose of 5000 mg/kg bw for 24 hrs to 5 males and females (2011 c). Erythema (score of 1 or 2) were observed in 9 rats and ulceration was observed in 4 rats on the day after application of the test substance. No dermal irritation was observed in the remaining rat. As the skin reactions might be due to the gauze pads adhering to the test sites at the time of washing, a clear link to skin irritating properties of the test substance is not concluded.

 

In the 28-days repeated dose toxicity study in rats dermal administration of 400 and 1000 mg/kg bw/day test substance induced very slight erythema in single animals graded with score 1 (400 mg/kg bw/day: 1/10 males on Day 28; 1000 mg/kg bw/day: 2/10 males on Day 3 and 28, respectively) (2008 a). Females did not reveal signs of local effects on the skin at the application site. Regarding the low severity and low incidence of local reactions, they are not considered as indicative for skin irritating properties of the test substance that required classification.

 

In summary, no local skin reactions have been observed in the standard skin irritation test according to OECD TG 404. Local skin reactions observed in rats either appeared at relatively high doses of 5000 mg/kg bw after application of 24 hrs and/or were relatively mild and only observed in single animals at doses of 400 or 1000 mg/kg bw/day after repeated dermal applications. Thus, if observed dermal reactions were generally of a mild nature, appeared at low incidences at rather high dosages and were not reported to be persistent over 14 days. Thus, the available data do not meet the criteria for classification for skin irritation according to regulation (EC) No 1272/2008 on classification, labelling and packaging (CLP).

 

Eye irritation

In the available eye irritation study performed with the test substance according to OECD TG 405 and in compliance with GLP (2011 e), approximately 86 mg of test substance was instilled in the eye of three rabbits. The untreated eye served as control. The treated and control eyes of all rabbits remained unwashed. Local reactions in the eyes were evaluated according to the Draize scoring system 1, 24, 48 and 72 h after application.

No effects on corneal opacity were noted at any time point in any animal. However, a mean iris score of 0.67 (24/48/72 h) was observed in one animal, which was fully reversible within 72 h. In addition, two animals had a mean conjunctivae score of 0.67 and the other one of 1 (24/48/72 h), which was fully reversible within 72 h in all animals. A chemosis score of 0.33 (24/48/72 h) was observed in one animal and it was fully reversible within 48 h. At an additionally investigated time point 1h after test substance administration, a mean iris score of 0.33, a mean conjunctivae score of 2 and a mean chemosis score of 1.33 was determined (mean of 3 animals, respectively). Fluorescein stain examinations were negative for corneal injury all rabbits. No clinical signs of toxicity were observed, and no body weight loss occurred. No further local or systemic effects were observed in any animal during the study period. Based on the obtained results, the test substance is not considered eye irritating.

Conclusion on skin and eye irritation properties

In conclusion, the test substance exhibited only low skin and eye irritating properties not meeting the classification criteria for skin and eye irritation according to according to regulation (EC) No 1272/2008 on classification, labelling and packaging (CLP).

Justification for classification or non-classification

The test substance does not meet the classification criteria for skin and eye irritation according to regulation (EC) No 1272/2008. Data are therefore conclusive but not sufficient for classification.