4,6-Dichloro-5-fluoropyrimidine

Basic toxicokinetics

Administrative data

Endpoint:

basic toxicokinetics, other

Remarks:

expert statement

Type of information:

other: expert statement

Adequacy of study:

other information

Reliability:

2 (reliable with restrictions)

Rationale for reliability incl. deficiencies:

data from handbook or collection of data

Cross-reference

Cross-reference 1

Reason / purpose for cross-reference:

reference to same study

Reference

Endpoint:

short-term repeated dose toxicity: oral

Type of information:

experimental study

Adequacy of study:

key study

Study period:

2005-05-20 to 2005-08-31

Reliability:

1 (reliable without restriction)

Rationale for reliability incl. deficiencies:

guideline study

Qualifier:

according to guideline

Guideline:

OECD Guideline 407 (Repeated Dose 28-Day Oral Toxicity Study in Rodents)

Version / remarks:

27 July 1995

GLP compliance:

yes

Limit test:

no

Species:

rat

Strain:

Wistar

Remarks:

(Hsd Cpb:WU)

Sex:

male/female

Details on test animals or test system and environmental conditions:

TEST ANIMALS
- Source: Harlan Winkelmann GmbH, Borchen, Germany
- Females (if applicable) nulliparous and non-pregnant: yes
- Age at study initiation: 6 weeks
- Weight at study initiation: males: 139.8 g (mean); females: 122.5 g (mean)
- Fasting period before study:
- Housing: during the experimental period two or 3 animals each were placed
together in Makrolon® cages Type IV under controlled environmental conditions on low-dust wood granules (supplier: Ssniff Spezialdiaten Inc. Soest/Westfalen, Germany; manufacturer: J. Rettenmeier, Ellwangen-Holzmuhle, Germany). Deviating therefrom animals were kept individually in cages Type IIa during the FOB/MA investigations.
- Diet (e.g. ad libitum): ad libitum, fixed-formula standard diet (Provimiv Kliba 3883.0.15 supplied by Provimi Kliba SA, CH-4303 Kaiseraugst, Switzerland)
- Water (e.g. ad libitum): ad libitum, tap water
- Acclimation period: 5 days

ENVIRONMENTAL CONDITIONS
- Temperature (°C): 22+/-2
- Humidity (%): 55+/-5
- Air changes (per hr): 10
- Photoperiod (hrs dark / hrs light): 12/12

IN-LIFE DATES: From: May 20, 2005 To: End of in Life-Phase: Jun. 24, 2005

Route of administration:

oral: gavage

Vehicle:

other: demineralized water / Cremophor EL (98% / 2%; v/v)

Details on oral exposure:

PREPARATION OF DOSING SOLUTIONS: The test item was formulated with demineralized water / Cremophor EL (98% / 2%; v/v) at the appropriate concentrations at room temperature and maximally used over the stability period determined by chemical analyses.

VEHICLE
- Concentration in vehicle: 0.1, 0.5, 2.5 mg/mL
- Amount of vehicle (if gavage): 10mL/kg bw

Analytical verification of doses or concentrations:

yes

Details on analytical verification of doses or concentrations:

A UV-spectrometric method for quantifying the test item in liquid formulations was developed. For analytical investigations, representative samples from the test item formulation covering the test item concentration range used in the study, were taken.
These samples were diluted with a mixture of methanol and 0.4 % aqueous perchloric acid (4:1) and subsequently quantified by a spectrometric method with UV-detection; wave length: 262 nm. Standard solutions of the authentic test item were used for calibration. Linearity, Precision, Specificity and Accuracy of the analytical method were evaluated apart from this GLP-study and fulfil the predefined acceptance criteria. Additional system suitability tests in terms of specificity, precision and linearity indicated, that qualified analytical procedures were followed during the study.
Analysis of blank samples (0 mg/ml) revealed no measurable traces of test item.
In order to generate and process the analytical data a commercial available computerised system was used.
Hardware and operating system:
• UV 2401-PC Photospectrometer; Fa. Shimadzu
• Windows NT computer with UV-Probe software version 1.10

Duration of treatment / exposure:

30 days

Frequency of treatment:

daily

Dose / conc.:

25 mg/kg bw/day (nominal)

Dose / conc.:

5 mg/kg bw/day (nominal)

Dose / conc.:

1 mg/kg bw/day (nominal)

Dose / conc.:

0 mg/kg bw/day (nominal)

No. of animals per sex per dose:

5

Control animals:

yes, concurrent vehicle

Details on study design:

- Dose selection rationale: The dose levels used in this study were based on the results of a pilot study. During this study Wistar rats (2 males and 2 females per dose group) received the test substance in doses of 0, 5, 25,150 mg/kg for a period of two weeks. Animals of the high dose group died or had to be sacrificed in moribund condition. These animals showed clinical findings like piloerection, apathy, laboured breathing. No clinical findings were recorded for the animals of the other groups.
At 5 mg/kg body weight development was comparable to that of the control groups.
At 25 mg/kg body weight development was affected by the treatment (mainly during the first part of the study). At necropsy at the end of the study the stomach of males and females at 25 mg/kg was discolored and appeared thickened. Based on these results the following dose scheme was used for the present study (in agreement with the sponsor representative): 0,1, 5, 25 mg/kg.
Food intake at 25 mg/kg was lower in males and females during the first week.
- Rationale for animal assignment (if not random): Before the start of the study, male and female animals were assigned to the dose
groups. For that purpose they were placed singly at their arrival in cages in the order they were taken out of the boxes. Thereafter, the animal weights were determined and recorded as well as the position of the animal on the shelf. Animals with extremely high or low body weights as well as surplus animals were removed. Based on evenly distributed chance numbers especially generated for this study, the animals were allocated to their final cage number specified by the random list (whereby the first random number on the list gave the actual position of the animal to be moved into cage No. 1, and so on). The cages were then placed one after the other in shelves in the order of increasing numbers.

Observations and examinations performed and frequency:

CAGE SIDE OBSERVATIONS: Yes
- Time schedule: daily

DETAILED CLINICAL OBSERVATIONS: Yes
- Time schedule: weekly

BODY WEIGHT: Yes
- Time schedule for examinations: daily

FOOD CONSUMPTION AND COMPOUND INTAKE (if feeding study):
- Food consumption for each animal determined and mean daily diet consumption calculated as g food/kg body weight/day: Yes
- Compound intake calculated as time-weighted averages from the consumption and body weight gain data: No

FOOD EFFICIENCY:
- Body weight gain in kg/food consumption in kg per unit time X 100 calculated as time-weighted averages from the consumption and body weight gain data: No

WATER CONSUMPTION AND COMPOUND INTAKE (if drinking water study): No

OPHTHALMOSCOPIC EXAMINATION: Yes
- Time schedule for examinations: before start and near the end i
- Dose groups that were examined: n the high dose group and the control

HAEMATOLOGY: Yes
- Time schedule for collection of blood: once at the experimental end
- Anaesthetic used for blood collection: No
- Animals fasted: No
- How many animals: All

CLINICAL CHEMISTRY: Yes
- Time schedule for collection of blood: once at the experimental end
- Animals fasted: No
- How many animals: All

PLASMA/SERUM HORMONES/LIPIDS: No
-
URINALYSIS: Yes
- Time schedule for collection of urine: once day 23
- Metabolism cages used for collection of urine: Not specified
- Animals fasted: yes, during urine collection


NEUROBEHAVIOURAL EXAMINATION: Yes / No / Not specified
- Time schedule for examinations: once day 29 or 30
- Dose groups that were examined: all
- Battery of functions tested: sensory activity / grip strength / motor activity / other:
- home cage observation: posture, piloerection, gait abnormalities, involuntary motor
movements, vocalization, others.
- observations during handling: ease of removing, reaction to being handled, muscle
tone, palpebral closure, lacrimation, nasal discharge, salivation, stains, others.
- open field observations: piloerection, respiratory abnormalities, posture,
involuntary motor movements, stereotypy, bizarre behavior, gait abnormalities, vocalization, arousal, rearing, defecation, urination, others.
- reflex/physiologic observations and measurements: approach response, touch response, auditory response, tail pinch response, righting reflex, pupil size, body temperature, grip strength, landing foot splay.

IMMUNOLOGY: No

Sacrifice and pathology:

GROSS PATHOLOGY: Yes

HISTOPATHOLOGY: Yes

Clinical signs:

effects observed, treatment-related

Description (incidence and severity):

Clinical observations of the animals revealed no treatment-related findings at 1 or 5 mg/kg. At 25 mg/kg 3 males and 2 females showed increased salivation.
Furthermore, piloerection, emaciation, sunken flanks, poor general condition, reduced motility, hunched back and breathing sounds were observed for the male (No. 16) of this dose group, which had to be sacrificed in moribund condition. Tremor was observed once in the 25 mg/kg male No. 18.

Mortality:

mortality observed, non-treatment-related

Description (incidence):

Survival was not affected by the treatment up to 25 mg/kg in males and females.
Male No. 16 (25 mg/kg group) had to be sacrificed on day 21 of the study due to its moribund condition. Histopathology revealed that the lung showed granulocytic infiltration and foreign body granuloma formation. In the wall of the esophagus severe inflammation and necroses accompanied by acanthosis of the epithelium was observed. As these findings are indicative of gavage error the death of this animals is not considered to be test substance-related.

Body weight and weight changes:

effects observed, treatment-related

Description (incidence and severity):

Body weight development was not retarded by the treatment with the test substance up to 5 mg/kg. At 25 mg/kg mean body weights of males were (on most days statistically) significantly lower compared to the control group. The differences reached up to 22% and were 7% at the end of the study. Although the differences in body weights between the 25 mg/kg female group and the control group were not statistically significant it can be seen, that the body weight development was retarded also in this group. The difference to control reached up to 9% and were 8% at the end of the study. The transient decrease in body weights on day 23 in male and female groups was caused by the urinary collection period during which animals received no food.

Food consumption and compound intake (if feeding study):

effects observed, non-treatment-related

Description (incidence and severity):

At 25 mg/kg the food intake per animal was slightly lower compared to that of the
control group. Related to the body weight it was comparable to that of the control
group.

Ophthalmological findings:

no effects observed

Description (incidence and severity):

These examinations gave no evidence for treatment-related effects on the eyes.

Haematological findings:

effects observed, treatment-related

Description (incidence and severity):

The hematological investigations resulted in a few values, which were statistically significantly different to control. However, these differences were not dose related and/or the difference to the respective control value was slight. A toxicological relevance is therefore not assumed. The hematological investigations gave no indication of a test substance related effect on blood coagulation up to 25 mg/kg.
Differential blood count revealed an increase in the number of neutrophils at 25
mg/kg in males and females.

Clinical biochemistry findings:

effects observed, treatment-related

Description (incidence and severity):

Clinical laboratory tests [determination of enzyme activities, concentrations of substrates and electrolytes in peripheral blood revealed in males a slight decrease in concentrations of protein and albumin at 25 mg/kg. The significantly decreased creatinine (25 mg/kg) and glucose (all dose groups) concentrations in females were not considered to be of toxicological relevance as the differences were relatively slight, no individual value did exceed the -2s range of historical control values, a strict dose dependence was absent (glucose) and correlating findings (e.g. histopathological findings) were not observed.

Urinalysis findings:

no effects observed

Description (incidence and severity):

Quantitative analyses of excreted volume, density, protein and creatinine showed that the protein excretion and the protein/creatinine quotient were slightly decreased at 25 mg/kg in males. The semi-quantitatively determined parameters (pH, blood, bilirubin, glucose, protein, ketone bodies and urobilinogen) were not relevantly changed by the treatment. The microscopy of sediment produced no remarkable findings.

Behaviour (functional findings):

no effects observed

Description (incidence and severity):

In summary, the investigations in the home cage, during handling and in the open
field as well as the determination of grip strength and landing foot splay showed no relevant signs or symptoms indicating evidence for a neurotoxic potential in rats exposed to the test substance. The activity determination over the entire 60 minute observation period did not reveal a relevant effect on motor (MA) and locomotor activity (LMA).
The results of the 10-minute intervals indicate that treated and control groups
reacted in a similar way and that there were no relevant differences in the first 10-minute intervals, which are regarded as best indicator of increases or decreases in activity before the animals habituate to a minimal level of activity.

Organ weight findings including organ / body weight ratios:

effects observed, treatment-related

Description (incidence and severity):

Compared to the respective control group relative weights of livers at 25 mg/kg were 14% higher in males and 16% in females.
In 25 mg/kg females relative weights of hearts were significantly increased. The difference to control was 21%.

Gross pathological findings:

effects observed, treatment-related

Description (incidence and severity):

Gross findings related to the treatment were observed in the forestomach in male and female rats at 25 mg/kg. These were white coverings of the forestomach mucosa in four males and five females of the high dose group.
Furthermore, prostate and seminal vesicle were diminished in size in two high dose males. No further findings were observed which were regarded to be related to the treatment with the test substance.

Histopathological findings: non-neoplastic:

effects observed, treatment-related

Description (incidence and severity):

At histopathology, squamous cell hyperplasia with hyperkeratosis in the forestomach was observed with incidences of 0/0/5/4 in males and 0/1/5/5 in
females. The severity score increased with the dose. At 25 mg/kg, hyperplasia and hyperkeratosis were accompanied by ulceration of the forestomach epithelium and chronic inflammation with giant cells of the underlying submucosa. These lesions were scored as grade 2 or 3 (slight to moderate).
Occasionally foreign bodies namely hair fragments were detectable within the
inflammatory lesions. The number of adipocytes was reduced in the femoral (males and females) and sternal (only females) bone marrow at 25 mg/kg. Concomitantly, granulopoiesis appeared to be increased in both femoral and sternal bone marrow in rats of either sex dosed at 25 mg/kg.
Corresponding to the reduced size of accessory sex glands seen in individual males at 25 mg/kg, prostate and seminal vesicles/coagulating glands were atrophic in males of this group. Testes and epididymides revealed no changes influenced by the treatment.
A variety of spontaneous changes was noted in control and treated animals with no indication of an effect of treatment. The spectrum of changes is mainly consistent with changes commonly encountered in laboratory rats of this age and strain.

Key result

Dose descriptor:

NOAEL

Remarks:

local

Effect level:

1 mg/kg bw/day (nominal)

Based on:

test mat.

Sex:

female

Basis for effect level:

other:

Remarks on result:

other: local NOAEL was determined based on local irritative effects on the forestomach mucosa

Key result

Dose descriptor:

NOAEL

Remarks:

systemic

Effect level:

5 mg/kg bw/day (nominal)

Based on:

test mat.

Sex:

male/female

Basis for effect level:

body weight and weight gain

clinical signs

histopathology: non-neoplastic

organ weights and organ / body weight ratios

Key result

Critical effects observed:

yes

Lowest effective dose / conc.:

5 mg/kg bw/day (nominal)

System:

gastrointestinal tract

Organ:

stomach

Treatment related:

yes

Dose response relationship:

yes

Relevant for humans:

presumably yes

Conclusions:

Under the conditions described the systemic no-observed-adverse-effect level (NOAEL) was 5 mg/kg body weight for males and females.
With respect to the findings resulting from the local irritative effects on the forestomach mucosa a NOAEL could not be established in females. The changes in
the forestomach in a single female at 1 mg/kg is considered as a borderline case. In males the NOAEL for local toxicity was 1 mg/kg.

Executive summary:

In a subacute toxicity study according to OECD test guideline 407 4,6-dichlor-5-fluorpyrimidine (100 % a.i.)] was administered to 5 Wistar rats/sex/dose in demineralized water containing 2 % v/v Cremophor EL at dose levels of 0, 1, 5, 25 mg/kg bw/day.

At the highest dose of 25 mg/kg bw clinical signs. 3 males and 2 females showed increased salivation. At 25 mg/kg mean body weights of males were (on most days statistically) significantly lower compared to the control group. The differences reached up to 22% and were 7% at the end of the study. The differences in body weights between the 25 mg/kg female group and the control group were not statistically significant but the body weight development was retarded also in this group. The difference to control reached up to 9% and were 8% at the end of the study. Compared to the respective control group relative weights of livers at 25 mg/kg were 14% higher in males and 16% in females. In 25 mg/kg females relative weights of hearts were significantly increased. At histopathology, squamous cell hyperplasia with hyperkeratosis in the forestomach was observed with incidences of 0/0/5/4 in males and 0/1/5/5 in females. The severity score increased with the dose. The difference to control was 21%. At 25 mg/kg, hyperplasia and hyperkeratosis were accompanied by ulceration of the forestomach epithelium and chronic inflammation with giant cells of the underlying submucosa. These lesions were scored as grade 2 or 3 (slight to moderate).

Occasionally foreign bodies namely hair fragments were detectable within the inflammatory lesions. The number of adipocytes was reduced in the femoral (males and females) and sternal (only females) bone marrow at 25 mg/kg. Concomitantly, granulopoiesis appeared to be increased in both femoral and sternal bone marrow in rats of either sex dosed at 25 mg/kg. Based on these results the systemic NOAEL is 5 mg/kg bw for both males and females. With respect to the findings resulting from the local irritative effects on the forestomach mucosa a NOAEL could not be established in females. The changes in the forestomach in a single female at 1 mg/kg is considered as a borderline case. In males the NOAEL for local toxicity was 1 mg/kg.

 

This subacute toxicity study in the rat is acceptable and satisfies the guideline requirement for a subacute oral study in rats.

Cross-reference 2

Reason / purpose for cross-reference:

reference to same study

Reference

Endpoint:

skin sensitisation: in vivo (non-LLNA)

Type of information:

experimental study

Adequacy of study:

key study

Study period:

2001-07-29 to 2003-07-14

Reliability:

1 (reliable without restriction)

Rationale for reliability incl. deficiencies:

guideline study

Qualifier:

according to guideline

Guideline:

OECD Guideline 406 (Skin Sensitisation)

Version / remarks:

17 July 1992

Qualifier:

according to guideline

Guideline:

EPA OPPTS 870.2600 (Skin Sensitisation)

Version / remarks:

August 1998

GLP compliance:

yes

Type of study:

guinea pig maximisation test

Justification for non-LLNA method:

A valid GPMT conducted according to guideline is available, which is reliable without restrictions and adequate for classification and labelling purposes. Potency estimation is not mandatory when existing guideline and GLP conforming data are available, which were conducted before the new annex of the REACH Regulation entered into force. For this reason and for reasons of animal welfare no additional LLNA was conducted.

Species:

guinea pig

Strain:

Hartley

Remarks:

Crl: HA for the main study and HsdPoc:DH for the rasnge finding study

Sex:

female

Details on test animals and environmental conditions:

TEST ANIMALS
- Source: Charles River Laboratory Animal Breeders in 88353 Kißlegg and Harlan Winkelmann GmbH Laboratory Animal Breeders in 33176 Borchen
- Females (if applicable) nulliparous and non-pregnant: yes
- Microbiological status of animals, when known:
- Age at study initiation: 4 weeks
- Weight at study initiation: 272-377 g
- Housing: the animals were conventionally kept in type IV Makrolon® cages, in groups of five during the adaptation period and in groups of two or three per cage throughout the study period. Low-dust wood shavings supplied by Ssniff Spezialdiaten GmbH, Soest, were used as bedding.
- Diet (e.g. ad libitum): ad libitum, "PROVIMI KLIBA 3420 - Maintenance Diet for Guinea Pigs" supplied by PROVOMI KLIBA AG
- Water (e.g. ad libitum): ad libitum, tap water
- Acclimation period: 5 days
- Indication of any skin lesions: not reported

ENVIRONMENTAL CONDITIONS
- Temperature (°C): 22 +/- 3
- Humidity (%): 40- 60
- Air changes (per hr):10
- Photoperiod (hrs dark / hrs light):12/12

No. of animals per dose:

20 animals for the main study and treatment groups and 10 animals for the control and the range-finding experiments

Details on study design:

RANGE FINDING TESTS:

MAIN STUDY
A. INDUCTION EXPOSURE
- No. of exposures: two, epicutaneous and intradermal
- Exposure period: 48 h for epicutaneous
- Test groups:
Epicutaneous:
Test item group: 0.5 ml 1% HEC 5725-DCF-PYRIMIDINE
intradermal:
Injection site: medial/bilateral 0.5% HEC 5725-DCF-PYRIMIDINE formulated in polyethylene glycol 400
Injection site: caudal/bilateral 0.5% HEC 5725-DCF-PYRIMIDINE formulated at equal parts in polyethylene
glycol 400 and complete Freund's adjuvant
- Control group:
Epicutaneous: 0.5 ml polyethylene glycol 400
Intradermal: Injection site: cranial/bilateral complete Freund's adjuvant (DIFCO LAB.) diluted 1:1 with sterile physicological saline solution
- Site: Dorsal
- Frequency of applications: Epicutaneous once one week prior to intradermal injection.
Intradermal once 21 days before challenge
- Duration: Epicutaneous 48 h
- Concentrations: Intradermal 0.5%; Epicutaneous: 1%

B. CHALLENGE EXPOSURE
- No. of exposures: One
- Day(s) of challenge: 22nd
- Exposure period: 24 h
- Test groups: 1% test item in polyethylene glycol
- Control group: 1% test item in polyethylene glycol
- Site: the 1% test item
formulation was placed on the right flank (caudal) of the animals of the test item group and the control group and held securely in place on the skin with a ORABAND® self-adhesive tape for 24 hours.
- Concentrations: 1%
- Evaluation (hr after challenge): 21 h

Positive control results:

alpha-Hexylzimtaldehyd formulated in sterile physicological saline solution

Key result

Reading:

2nd reading

Hours after challenge:

48

Group:

positive control

Dose level:

5% induction and 12% challenge

Remarks on result:

other: total number of animals was not reported: 100% dermal reaction to positive control

Key result

Reading:

2nd reading

Hours after challenge:

48

Group:

negative control

Dose level:

1%

No. with + reactions:

0

Total no. in group:

10

Key result

Reading:

1st reading

Hours after challenge:

24

Group:

negative control

Dose level:

1%

No. with + reactions:

0

Total no. in group:

10

Key result

Reading:

2nd reading

Hours after challenge:

48

Group:

test chemical

Dose level:

1%

No. with + reactions:

18

Total no. in group:

20

Clinical observations:

Appearance and behaviour of the test item group were not different from the control group.

Key result

Reading:

1st reading

Hours after challenge:

24

Group:

test chemical

Dose level:

1%

No. with + reactions:

20

Total no. in group:

20

Clinical observations:

Appearance and behaviour of the test item group were not different from the control group.

Interpretation of results:

Category 1A (indication of significant skin sensitising potential) based on GHS criteria

Conclusions:

After the intradermal induction the animals in the test item group showed strong effects up to encrustation at the injection sites of the first induction.
The challenge with the 1% test item formulation led to skin effects (grade 1-3) in 20
of 20 animals (100%) in the test item group and no skin effects were seen in the
control group animals. In summary, by comparing the results in the treatment group and in the control group under the conditions of the maximization test and with respect to the evaluation criteria the test item therefore exhibits a skin sensitization potential.

Executive summary:

In a dermal sensitization study according to OECD test guideline 406 (1992) with 4,6-dichlor-5-fluorpyrimidine (100 % a.i).] in polyethylene glycol, young adult Hartley guinea pigs(20/treatment group) were tested using the method of Magnusson and Kligman. After the intradermal induction the animals in the test item group showed strong effects up to encrustation at the injection sites of the first induction. The challenge with the 1% test item formulation led to skin effects (grade 1-3) in 20 of 20 animals (100%) in the test item group and no skin effects were seen in the control group animals.

Based on the results 4,6-dichlor-5-fluorpyrimidine is classified according to Regulation (EU) No. 1272/2008 (CLP) and the Globally Harmonized System for Classification and Labelling of Chemicals (GHS) as skin sensitizer Category 1A (May cause an allergic skin reaction).

Cross-reference 3

Reason / purpose for cross-reference:

reference to same study

Reference

Endpoint:

skin irritation: in vivo

Type of information:

experimental study

Adequacy of study:

key study

Study period:

2001-08-28 to 2001-10-08

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:

17 July 1992

GLP compliance:

yes

Species:

rabbit

Strain:

Himalayan

Details on test animals or test system and environmental conditions:

TEST ANIMALS
- Source: LPT Laboratory of Pharmacology and Toxicology KG D-24601 Lohndorf/Post Wankendorf
- Age at study initiation: approx 6-11 months
- Weight at study initiation: 2.3-2.9 kg

Type of coverage:

semiocclusive

Preparation of test site:

shaved

Remarks:

dermal application to the shaved intact dorsal skin

Vehicle:

unchanged (no vehicle)

Controls:

yes, concurrent no treatment

Amount / concentration applied:

0.5 mL/patch

Duration of treatment / exposure:

4 h

Observation period:

14 days

Number of animals:

3

Details on study design:

TEST SITE
- Area of exposure: approx 6 cm²
- Type of wrap if used: The test substance was applied to the test site and then covered with a gauze patch. The patch was held in contact with the skin by means of a semi-occlusive dressing for the duration of the exposure period.

OBSERVATION TIME POINTS
(indicate if minutes, hours or days) 60 min, 24, 48, 72 h and 4 to 14 days

SCORING SYSTEM: according to test guideline

Irritation parameter:

edema score

Basis:

animal #2

Time point:

24/48/72 h

Score:

4

Max. score:

4

Reversibility:

not fully reversible within: 14 days, area of application dark discoloured, swelling on left abdomen increasing to: induration of the skin and skin in area of application discoloured and indurated, skin around area of application red and warm

Irritation parameter:

erythema score

Basis:

animal #2

Time point:

24/48/72 h

Score:

4

Max. score:

4

Reversibility:

not fully reversible within: 14 days, area of application dark discoloured, swelling on left abdomen increasing to: induration of the skin and skin in area of application discoloured and indurated, skin around area of application red and warm

Irritation parameter:

edema score

Basis:

animal: #1 - #3

Time point:

other: 60 min

Score:

2

Max. score:

4

Reversibility:

other: animals 1 and 3 died therafter

Irritation parameter:

erythema score

Basis:

animal: #1 - #3

Time point:

other: 60 min

Score:

1

Max. score:

4

Reversibility:

other: Animal 1 and 3 died thereafter

Interpretation of results:

Category 1 (corrosive) based on GHS criteria

Conclusions:

Under the present test conditions all three animals showed an erythema (grade 1) 1 hour after patch removal.
All three animals showed an oedema (grade 2) 1 hour after patch removal, animal no. 2 (grade 1) in addition at the examination time-points 24 and 48 hours after patch removal.
Additionally, dark discolouration of skin of the area of application and a swelling on the left abdomen were noted in animal no. two 24 hours to 14 days after patch removal. Induration of skin was noted in animal no. two 72 hours to 7 days after patch removal. Furthermore, from 8 to 14 days after patch removal discolouration and induration of the area of application was noted while the skin around this area was red and warm.
Animal nos. 1 and 3 were found dead at the examination time point 24 hours after patch removal.

Executive summary:

In a primary dermal irritation study according to OECD guideline 404 (1992), young adult Himalayan rabbits (3/treatment) were dermally exposed to 0.5 mL/patch of 4,6-dichlor-5-fluorpyrimidine (100 % a.i.) for 4 hours to 6 cm² of shaved dorsal skin. Animals then were observed for 14 days. Irritation was scored as required by the test guideline.

Under the present test conditions all three animals showed an erythema (grade 1) 1 hour after patch removal.

All three animals showed an oedema (grade 2) 1 hour after patch removal, animal no. 2 (grade 1) in addition at the examination time-points 24 and 48 hours after patch removal.

Additionally, dark discolouration of skin of the area of application and a swelling on the left abdomen were noted in animal no. two 24 hours to 14 days after patch removal. Induration of skin was noted in animal no. two 72 hours to 7 days after patch removal. Furthermore, from 8 to 14 days after patch removal discolouration and induration of the area of application was noted while the skin around this area was red and warm.

Animal nos. 1 and 3 were found dead at the examination time point 24 hours after patch removal. In this study, 4,6-dichlor-5-fluorpyrimidine is extremely irritating to the skin based on the presented results. Based on these results the test item is classified according to Regulation (EU) No. 1272/2008 (CLP) and the Globally Harmonized System for Classification and Labelling of Chemicals (GHS) as corrosive Category 1 (Causes severe skin burns and eye damage).

Cross-reference 4

Reason / purpose for cross-reference:

reference to same study

Reference

Endpoint:

acute toxicity: oral

Type of information:

experimental study

Adequacy of study:

key study

Study period:

2001-08-09 to 2002-08-26

Reliability:

1 (reliable without restriction)

Rationale for reliability incl. deficiencies:

guideline study

Qualifier:

according to guideline

Guideline:

OECD Guideline 423 (Acute Oral toxicity - Acute Toxic Class Method)

Version / remarks:

22 March 1996

GLP compliance:

yes

Test type:

acute toxic class method

Limit test:

no

Species:

rat

Strain:

Wistar

Remarks:

HsdCpb:WU

Sex:

male/female

Details on test animals or test system and environmental conditions:

TEST ANIMALS
- Source: Harlan Winkelmann GmbH, Borchen, District of Paderborn
- Females (if applicable) nulliparous and non-pregnant: yes
- Age at study initiation: males approx. 10 weeks and females approx. 9 weeks
- Weight at study initiation: males: 242-266 g; females: 172-188 g
- Fasting period before study: not reported
- Housing: grouped, conventionally in polycarbonate cages; the bedding consisted of low-dust wood granules type BK 8/15 (supplier: Ssniff, Spezialdiäten GmbH, Soest/Westphalia).
- Diet (e.g. ad libitum): ad libitum, "NAFAG Ò No. 9441 W 10" (manufacturer: Eberle Nafag AG, Gossau)
- Water (e.g. ad libitum): ad libitum, tap water
- Acclimation period: 5 days

ENVIRONMENTAL CONDITIONS
- Temperature (°C): 22° +/- 2°C
- Humidity (%): 55 +/- 5
- Air changes (per hr): 10
- Photoperiod (hrs dark / hrs light): 12/12

Route of administration:

oral: gavage

Vehicle:

other: The test substance was formulated in demineralized water with the aid of 2 % Cremophor EL before administration.

Details on oral exposure:

VEHICLE
- Concentration in vehicle: 200, 20 and 2.5 mg/mL
- Amount of vehicle (if gavage): 10 mL/kg bw

MAXIMUM DOSE VOLUME APPLIED: 10 mL/kg bw


CLASS METHOD (if applicable) acute toxic class
- Rationale for the selection of the starting dose: as required by the test guideline

Doses:

2000, 200 and 25 mg/kg bw

No. of animals per sex per dose:

3

Control animals:

no

Details on study design:

- Duration of observation period following administration: 14 days
- Frequency of observations and weighing: The body weights of the rats are recorded on day 1 before administration and then weekly.
- Necropsy of survivors performed: yes
- Other examinations performed: clinical signs, body weight

Statistics:

Classification is based on cut-off values as indicated in the OECD guideline.

Key result

Sex:

male/female

Dose descriptor:

LD50

Effect level:

> 30 - < 50 mg/kg bw

Based on:

test mat.

Mortality:

All animals of the 2000 mg/kg bw and the 200 mg/kg bw group died within 5 to 50 min and 20 min to 2 h, respectively.

Clinical signs:

irregular respiration

observations of tremors

salivation

other:

Body weight:

other body weight observations

Remarks:

The body weight and the body weight development of males and females were not affected by the treatment.

Gross pathology:

In animals that died during the observation period the following changes were detected:
Dark-red discoloration of liver
Slightly collapsed lung
Pale discoloration of kidneys
Pale discoloration of spleen
No gross pathologic changes were observed in animals sacrificed at the end of the study period.

Interpretation of results:

Category 2 based on GHS criteria

Conclusions:

In the present study conducted according to OECD test guideline 423 (1996) three animals were administered a single dose of either 2000, 200 or 25 mg/kg bw of the test substance and observed for further 14 days. All animals of the 2000 mg/kg bw and the 200 mg/kg bw groups died at least within 2h. None of the animals of the 25 mg/kg bw group died, thus the LD50 value was determined at > 30 < 50 mg/kg bw.

Executive summary:

In an acute oral toxicity study according to OECD guideline 423, adopted 22 March 1996, 6 female, fasted, 9 weeks old Wistar strain rats and 3 male, fasted, 10 weeks old Wistar strain rats were given a single oral dose of 4,6-dichlor-5-fluorpyrimidinbe in demineralized water containing 2% Cremophor by gavage at a dose of 2000, 200 (females) and 25 mg/kg bw (males and females) and observed for 14 days.

6/6 animals of the 2000 and 200 mg/kg bw groups died 50 min and 2h after dosing, respectively. Clinical signs shown by the animals found dead included decreased, uncoordinated gait, labored breathing, increased salivation, narrowed palpebral fissures, and temporary tremor,. lateral position, temporary convulsions, reddened skin, increased motility, and tachypnea.

A dose of 25 mg/kg body weight was tolerated by male and female rats without mortalities and clinical signs.

The body weight and the body weight development of males and females were not affected by the treatment.

In animals that died during the observation period the following changes were detected:

Dark-red discoloration of liver

Slightly collapsed lung

Pale discoloration of kidneys

Pale discoloration of spleen

No gross pathologic changes were observed in animals sacrificed at the end of the study period.

Oral LD50 (rat, combined) > 30< 50 mg/kg bw

Cross-reference 5

Reason / purpose for cross-reference:

reference to same study

Reference

Endpoint:

acute toxicity: dermal

Type of information:

experimental study

Adequacy of study:

key study

Study period:

2005-04-05 to 2005-08-01

Reliability:

1 (reliable without restriction)

Rationale for reliability incl. deficiencies:

guideline study

Qualifier:

according to guideline

Guideline:

OECD Guideline 402 (Acute Dermal Toxicity)

Version / remarks:

23 February 1987

GLP compliance:

yes

Test type:

standard acute method

Limit test:

no

Species:

rat

Strain:

Wistar

Sex:

male/female

Details on test animals or test system and environmental conditions:

TEST ANIMALS
- Source: Harlan/Winkelmann GmbH, 33178 Borchen, Germany
- Females (if applicable) nulliparous and non-pregnant: yes
- Age at study initiation: 9-13 weeks
- Weight at study initiation: males: 203-255 g; females: 207-232 g
- Fasting period before study:
- Housing: The animals were caged individually in polycarbonate cages on low dust wood granulate bedding (J. Rettenmaier & Sohne, 73494 Rosenberg, Germany).
- Diet (e.g. ad libitum): The animals received the standard diet "Provimi Kliba 3883.0.15 Maus/Ratte Haltung, Kaiseraugst Switzerland", ad libitum
- Water (e.g. ad libitum): tap water, ad libitum
- Acclimation period: 5 days

ENVIRONMENTAL CONDITIONS
- Temperature (°C): 22 +/- 2
- Humidity (%): 55 +/- 5
- Air changes (per hr): 10
- Photoperiod (hrs dark / hrs light): 12/12

Type of coverage:

semiocclusive

Vehicle:

unchanged (no vehicle)

Details on dermal exposure:

TEST SITE
- Area of exposure: between 1.0 and 20.0 cm²
- % coverage: approx. 10%
- Type of wrap if used: For each dose and animal the required amount of the pure liquid test substance was weighed and applied as uniformly and thinly as possible to the test area, covered with a gauze-layer (6.0 cm x 5.0 cm = 30.0 cm²) of a ,,Cutiplast® steril" coated with air-tight ,,Leukoflex®". The gauze strip was placed on the rat's back and secured in place using ,,Peha®-Haft" cohesive stretch tape (8 cm x 23 cm) and additionally covered with a "Lomir biomedical Inc rat jacket", which was connected with a
safety pin to the stretch tape to ensure that the animals could not ingest the test
substance.

REMOVAL OF TEST SUBSTANCE
- Washing (if done): After 24 h the dressings were removed and the area was rinsed with tepid water using soap and gently patting the area dry.
- Time after start of exposure: 24 h

TEST MATERIAL
- Amount(s) applied (volume or weight with unit): between 20.5 and 75.0 mg/cm²

Duration of exposure:

24 h

Doses:

160, 400, 1000, 4000 mg/kg bw

No. of animals per sex per dose:

5

Control animals:

not specified

Details on study design:

- Duration of observation period following administration: 21 days
- Frequency of observations and weighing: Clinical signs and mortality rates were determined several times on the day of application and subsequently at least once daily for an observation period of at least 21 days. The weight gain of the animals was checked weekly until the end of the study.
- Necropsy of survivors performed: yes
- Other examinations performed: clinical signs, body weight

Statistics:

The LD50 value was calculated with the aid of a software program according to Spearman, Karber (D.J. Finney; Statistical method in biological assay, 2nd Edition, Griffin, London, 524-530; 1971). The algorithm was taken from L. Sachs (Angewandte Statistik, 6th Edition 1984, pp. 178 ff.). Where calculation of the LD50 using the software program was not possible or meaningful, an assessment was made based on the applied dose and dose-response curve, respectively.

Key result

Sex:

male

Dose descriptor:

LD50

Effect level:

365 mg/kg bw

Based on:

test mat.

Key result

Sex:

female

Dose descriptor:

LD50

Effect level:

304 mg/kg bw

Based on:

test mat.

Mortality:

All animals of the 1000 and 4000 mg/kg bw treatment group died within 2 days. In the 400 mg/kg bw dose group 3 males and 4 females died after 2 to 3 days. None of the animals of dose group 160 mg/kg bw died.

Clinical signs:

other:

Body weight:

other body weight observations

Remarks:

There were no toxicological effects on body weight or body weight development in the surviving males and females.

Gross pathology:

Gross Pathology Findings
4000 mg/kg bw.:
stomach: fluid, gas-filled.
1000 mg/kg bw.:
stomach: consistency-changes light colored, partly reddening.
400 mg/kg bw.:
stomach: partly reddening, hemorrhagic watery fluid; lung: pale discolorations;
skin: black to blue discolorations of the treatment area, scars on the treatment area.
160 mg/kg bw.:
skin: scars on the treatment area.

Interpretation of results:

Category 3 based on GHS criteria

Conclusions:

Based on the present investigations, HEC 5725-DCF-Pyrimidine is to be regarded to have the following LD50 values:
LD50 rat, male : 365 mg/kg body weight (235-567 mg/kg)
rat, female : 304 mg/kg body weight (212-435 mg/kg)
So it is regarded as low toxic after dermal application.

Executive summary:

In an acute dermal toxicity study according to OECD test guideline 402 (1987), groups of young adult males and female Wistar rats (5/sex) were dermally exposed to 4,6-dichlor-5-fluorpyrimidine (100 % a.i) for 24 hours to 10 % of body surface area at doses of 160, 400, 1000 and 4000 mg/kg bw.  Animals then were observed for 21 days.

Dermal LD50 Males =365 mg/kg bw

                        Females = 304 mg/kg bw

4,6-dichlor-5-fluorpyrimidine is of low Toxicity based on the LD50 value for female Wistar rats. Animals of both high dose groups (1000 and 4000 mg/kg bw) died within 2 days. In all dose groups clinical signs related to treatment were observed corresponding to the findings during necropsy.

 Based on the results the substance needs to be classified according to Regulation (EU) 1272/2008 (CLP) and the Globally Harmonized System for Calssification and Labelling of Chemicals (GHS) as acute toxic via the dermal route Castegory 3 “Toxic in contact to skin”.

 

Data source

Reference

Reference Type:

other company data

Title:

Unnamed

Year:

2006

Report date:

2006

Materials and methods

Objective of study:

absorption

Test guideline

Qualifier:

no guideline followed

Principles of method if other than guideline:

Toxicokinetic statement for HEC 5725-DCF-Pyrimidine based on available toxicology data

GLP compliance:

no

Test material

Test material information

1

Reference substance name:

-

EC Number:

446-560-3

EC Name:

-

Cas Number:

213265-83-9

Molecular formula:

C4HCl2FN2

IUPAC Name:

4,6-dichloro-5-fluoropyrimidine

Results and discussion

Toxicokinetic / pharmacokinetic studies

Details on absorption:

Based on the provided data, the absorption of HEC 5725-DCF-Pyrimidine after oral administration is likely to be good after both oral and dermal administration. Dermal administration of 500 µL to the skin of three rabbits led to the death of two of the three animals within 24 hours, and in rats the dermal LD50 is approximately 300 mg/kg bw.
Repeated oral gavage administration led to decreased body weight at at 25 mg/kg bw/day in conjunction with local signs of irritation in the forestomach at 1, 5, and 25 mg/kg bw/day, although the majority of findings observed in the 28-day oral study may have been due simply to the local irritation observed in the stomach rather than to systemic toxicity of the test substance.

Applicant's summary and conclusion

Conclusions:

The substance is considered to be absorbed equally oral and dermal based on the available data.