Registration Dossier

Toxicological information

Repeated dose toxicity: oral

Currently viewing:

Administrative data

Endpoint:
chronic toxicity: oral
Type of information:
experimental study
Adequacy of study:
weight of evidence
Study period:
1983
Reliability:
2 (reliable with restrictions)
Rationale for reliability incl. deficiencies:
comparable to guideline study with acceptable restrictions

Data source

Reference
Reference Type:
publication
Title:
Unnamed
Year:
1983

Materials and methods

Test guideline
Qualifier:
equivalent or similar to guideline
Guideline:
OECD Guideline 452 (Chronic Toxicity Studies)
Deviations:
yes
Remarks:
only two doses
GLP compliance:
not specified
Limit test:
yes

Test material

Constituent 1
Chemical structure
Reference substance name:
Cytidine 5'-(trihydrogen diphosphate), mono[2-(trimethylammonio)ethyl] ester, hydroxide, inner salt, monosodium salt
EC Number:
251-689-1
EC Name:
Cytidine 5'-(trihydrogen diphosphate), mono[2-(trimethylammonio)ethyl] ester, hydroxide, inner salt, monosodium salt
Cas Number:
33818-15-4
Molecular formula:
C14H26N4O11P2.Na
IUPAC Name:
Sodium[(2R,3S,4R,5R)-5-(4-amino-2-oxo-1,2-dihydropyrimidin-1-yl)-3,4-dihydroxyoxolan-2-yl]methyl({[2-(trimethylazaniumyl)ethyl]phosphanato}oxy)phophonate
Test material form:
solid

Test animals

Species:
dog
Strain:
Beagle
Sex:
male/female
Details on test animals or test system and environmental conditions:
TEST ANIMALS
- Source: from own quarters
- Age at study initiation: 6 months of age
- Diet: Animals were fed once a day with "Pan-lab-dog" standard diet.
- Water: ad libitum

Administration / exposure

Route of administration:
oral: gavage
Details on route of administration:
esophageal cannulation
Vehicle:
other: agar
Details on oral exposure:
- VEHICLE
- Concentration in vehicle: 25% test item in 0.25% agar suspension
- Amount of vehicle (if gavage): 6 ml/kg
Analytical verification of doses or concentrations:
not specified
Duration of treatment / exposure:
6 months
Frequency of treatment:
Daily
Doses / concentrations
Dose / conc.:
1 500 mg/kg bw/day (nominal)
Remarks:
of CDP-choline
No. of animals per sex per dose:
3/sex in the test item group
1/sex in the control group
Control animals:
yes, concurrent vehicle
Positive control:
No

Examinations

Observations and examinations performed and frequency:
BODY WEIGHT: Yes
- Time schedule for examinations: weekly

HAEMATOLOGY: Yes
- Time schedule for collection of blood: At the begining and at the end of the test.
- Anaesthetic used for blood collection: Yes
- Animals fasted: Not specified.
- Parameters checked: hematocrit, hemoglobin, RBC, WBC, leukocyte formula.

CLINICAL CHEMISTRY: Yes
- Time schedule for collection of blood: At the begining and at the end of the test.
- Animals fasted: Not specified.
- Parameters checked: glucose, GOT, GPT, uric nitrogen (BUN), chloride, protein, globulin, lipids, cholesterol, bilirubin

URINALYSIS: Yes
- Time schedule for collection of urine: At the begining and at the end of the test.
- Metabolism cages used for collection of urine: not specified
- Animals fasted: Not specified.
- Parameters checked: pH, urinary density and qualitative determination
Sacrifice and pathology:
GROSS PATHOLOGY: Yes
All animals included in the study were submitted to a complete necropsy. A careful gross examination of organs was made and their weight recorded.
The following organs were examined and prepared for their histologic study:
Organs: liver (lower left and central lobules), kidney (both), heart, spleen, lung, ovary, testicle, mesenteric lymphatic ganglia (in some animals).

HISTOPATHOLOGY: Yes
The previously mentioned organs were examined. They were fixed in 10% neutral formol, then in paraffin and cut into 6 micrometers pieces. Dying of every organ was made with hematoxylin-eosin abd Perls' in the spleen for hemosiderin identification.
Statistics:
The number on animals used prevented from carrying out a stadistical study and comparison between both groups.The results shown are individually considered and restricted to the appraisal whether respective values fall within normal ranges.

Results and discussion

Results of examinations

Clinical signs:
no effects observed
Mortality:
no mortality observed
Body weight and weight changes:
no effects observed
Food consumption and compound intake (if feeding study):
not examined
Food efficiency:
not examined
Water consumption and compound intake (if drinking water study):
not examined
Ophthalmological findings:
not examined
Haematological findings:
no effects observed
Clinical biochemistry findings:
no effects observed
Urinalysis findings:
no effects observed
Behaviour (functional findings):
not examined
Immunological findings:
not examined
Organ weight findings including organ / body weight ratios:
no effects observed
Gross pathological findings:
no effects observed
Neuropathological findings:
not examined
Histopathological findings: non-neoplastic:
effects observed, non-treatment-related
Description (incidence and severity):
Liver: No signs of toxicity. The alterations found were characteristic of liver parenchyma and not related to the test item.They appear with similar frequency and intensity in control animals. Among the most frequent onces can be mentioned small granulomas ands infiltrations of inflamatory cells, necrosis of isolated liver cells and small areas of necrosis, and other alterations such as cumulations of bilirubin, multinucleate eosinophilic bodies and rather irregular distribution of liver glycogen from a group of animals to another.

Kidney: No nephrotoxic signs were found in any of the animals. It was observed numerous cortical granulomas of probable parasitic origin in one control animal and one animal from the test item group, problable provoked by Toxcara canis. Some cortical areas of intersticial infoltration were also appreciated.

Heart: One female from CDP-choline group showed a noticeable myocardial necrosis area. None of the other animals showed any increase of necrosis, hence the non-toxic character of the mentioned lesion os assumed.

Lung: The existence of an intersticial pneumonic process in various development degrees was observedd in all animals under study; one of them had various granulomatous foci, with remains of hemosiderin from small old hemorrhages provoked by this process.

Testicle: Some multinucleate giant cells were found sporadically in the tubular openings, without bearing any relationship with the test item.
Histopathological findings: neoplastic:
no effects observed
Other effects:
not examined

Effect levels

Key result
Dose descriptor:
NOAEL
Effect level:
1 500 mg/kg bw/day (nominal)
Based on:
test mat.
Sex:
male/female
Basis for effect level:
body weight and weight gain
clinical biochemistry
gross pathology
histopathology: non-neoplastic
mortality
organ weights and organ / body weight ratios
urinalysis

Target system / organ toxicity

Key result
Critical effects observed:
no

Any other information on results incl. tables

Table 1: Weight data (g):

Weeks

Control (male)

CDP-choline (male)

 

Control (female)

CDP-choline (female)

 

4

9.5

12.2

9.8

12.9

6

9.5

12.3

9.3

12.3

8

9.6

13.3

9.5

12.7

10

9.6

14.0

9.6

13.0

12

9.8

14.3

9.8

12.2

14

9.8

13.8

9.4

12.9

12

9.7

14.1

9.6

12.9

18

9.0

13.9

9.5

12.2

20

9.0

15.1

10.4

13.5

22

9.2

14.9

10.4

13.7

24

8.9

15.2

10.6

14.6

Weights for CDP-choline group (male and female) correspond to three animals' average

Table 2: Hematology parameters, 6 months

Parameters

Control (male)

Control (female)

CDP-choline

(male 1)

CDP-choline

(male 2)

CDP-choline

(male 3)

CDP-choline

(female 1)

CDP-choline

(female 2)

CDP-choline

(female 3)

Hematocrit

43

42

42

47

39

51

40

44

Hemoglobin

15.7

15.5

12.8

12.1

11.6

12.6

9.8

10.3

RBC

8430000

8040000

8070000

7800000

7480000

7230000

8960000

8550000

WBC

6000

6500

6700

11950

5100

10400

4200

6700

Lymphocite

 

41

44

45

30

38

42

48

42

Neutrophyl

58

55

54

70

61

57

52

58

Basophyl

0

0

0

0

0

0

0

0

Eosinophyl

1

1

0

0

0

1

0

0

Monocyte

0

0

1

0

1

0

0

0

Table 3: Chemistry parameters, 6 months.

Parameters

Control (male)

Control (female)

CDP-choline

(male 1)

CDP-choline

(male 2)

CDP-choline

(male 3)

CDP-choline

(female 1)

CDP-choline

(female 2)

CDP-choline

(female 3)

Glucose

70

66

66

78

75

79

102

79

GOT

63

35

43

55

30

43

76

41

GPT

81

95

68

31

35

35

37

27

BUN

16.5

12.5

19.5

18.3

32

19

18

23.5

Chloride

355

461.5

532.5

461.5

426

461.5

532.5

426

Lipids

574.1

629.6

751.9

796.3

592.6

685.2

703.7

777.8

Cholesterol

155.5

168.2

177.3

170.9

135.5

181.8

157.3

191.8

Bilirubin

0.4

0.3

0.1

0.1

0.3

0.2

0.1

0.2

Proteins

7.5

6.0

0.1

6.7

6.0

6.5

7

6.9

Globulins

6.3

5.6

5.5

4

4.6

5.1

6

4.6

Table 4: Urinalysis, 6 months.

Parameters

Control (male)

Control (female)

CDP-choline

(male 1)

CDP-choline

(male 2)

CDP-choline

(male 3)

CDP-choline

(female 1)

CDP-choline

(female 2)

CDP-choline

(female 3)

Urobilinogen

0.1

0.1

0.1

 

0.1

0.1

0.1

0.1

Blood

+ weak

+ weak

+ weak

+ weak

+ weak

+ +

NF

NF

Bilirubin

NF

NF

NF

NF

NF

NF

NF

NF

Ketones

NF

NF

NF

NF

NF

NF

NF

NF

Glucose

NF

NF

NF

NF

NF

NF

NF

NF

Proteins

NF

NF

NF

NF

NF

NF

NF

NF

pH

7

6

6

6

6

6

6

6

Density

1.06

1.02

1.00

1.03

1.00

1.02

1.00

1.00

Aspect

normal

normal

normal

normal

turbid

normal

normal

normal

NF = not found

Table 5: Weight of organs (g) and weight rate (%) after necropsy

Animal/ Organ

Liver

Spleen

Heart

R. kidney

L. kidney

Lung

R testicle

R. ovary

L. testicle

L. ovary

Control (female)

8.9 kg

38.1

4.3%

64.6

0.7%

69.3

0.8 %

21.1

0.2 %

20.7

0.2 %

81.7

0.9 %

0.3

0.003 %

5.4

0.1 %

Control (male)

10.6 kg

474.4

45.5%

69.9

0.7%

71.1

0.7 %

30.8

0.3 %

25.3

0.2

85.5

0.8 %

1.84

0.2 %

-

-

CDP-choline (male 1)

17.3 kg

496.5

4%

93

0.5%

93.3

0.5 %

42.1

0.2 %

44.3

0.3 %

165.2

1 %

12

0.1 %

12.5

0.1 %

CDP-choline (male 2)

16.7 kg

646.2

3.9%

92.5

0.6%

132.1

0.8%

42.2

0.3%

41.6

0.2%

137.9

0.8%

11.2

0.1%

10.6

0.1%

CDP-choline (male 3)

11.7 kg

427.0

3.6%

46.8

0.4%

94.5

0.8%

26.9

0.2%

27.7

0.2%

92.2

0.8%

10.4

0.1%

7.9

0.1%

CDP-choline (female 1)

15.6 kg

544.8

3.5%

84.8

0.5%

94.2

0.6%

31.0

0.2%

31.0

0.2%

22.2

0.1%

0.3

0.002%

1.2

0.01%

CDP-choline (female 2)

12.3 kg

655.0

5.3%

140.6

1.1%

79.0

0.6%

31.2

0.3%

31.3

0.3%

110.8

0.9%

1.4

0.01%

1.1

0.01%

CDP-choline (female 3)

15.8 kg

785.0

5%

107.0

0.7%

106.0

0.7%

32.2

0.2%

33.0

0.2%

125.9

0.8%

0.7

0.004%

1.6

0.01%

Applicant's summary and conclusion

Conclusions:
As no toxic effects were observed related to the test item, it can be concluded that citicoline has a NOAEL of 1500 mg/kg bw/day in dogs, after a 6 months oral administration.
Executive summary:

A 6-months repeated dose toxicity study was performed on Beagle dogs following a method similar to OECD 452. Citicoline was orally administered to 5 male and 5 female dogs, at doses of 1500 mg/kg/day and one control animal per sex which only received the vehicle. All animals were thoroughly observed daily for the onset of any toxic signs. Body weight changes were evaluated weekly, and haematology, clinical chemistry and urine parameters were also evaluated at the beginning and at the end of the test. After finalization of treatment, a gross pathology and histopathology analysis were conducted. No mortality or other effects were observed, except for some histopathological findings not related to the test item administration. The study did not reveal any clinical signs of toxicity. Under these conditions, Citicoline was found to have a NOAEL of 1500 mg/kg bw/day in dogs.