Caffeine

Developmental toxicity / teratogenicity

Administrative data

Endpoint:

developmental toxicity

Type of information:

experimental study

Adequacy of study:

key study

Study period:

no data

Reliability:

2 (reliable with restrictions)

Rationale for reliability incl. deficiencies:

other: Acceptable, well documented publication which meets basic scientific principles.

Data source

Reference

Reference Type:

publication

Title:

A study of the teratogenic potential of caffeine ingested in drinking water.

Author:

Collins TFX et al.

Year:

1983

Bibliographic source:

Fd Chem Toxic 21 (6): 763-777.

Materials and methods

Test guideline

Qualifier:

equivalent or similar to guideline

Guideline:

OECD Guideline 414 (Prenatal Developmental Toxicity Study)

Principles of method if other than guideline:

The study was carried out according to Collins TFX et al. (1981). A study of the teratogenic potential of caffeine given by oral intubation to rats Regul Toxic Pharmac 1: 355.

GLP compliance:

not specified

Limit test:

no

Test material

Test material information

Constituent 1

Reference substance name:

Caffeine

EC Number:

200-362-1

EC Name:

Caffeine

Cas Number:

58-08-2

Molecular formula:

C8H10N4O2

IUPAC Name:

1,3,7-trimethyl-3,7-dihydro-1H-purine-2,6-dione

Details on test material:

- Name of test material (as cited in study report): Caffeine
- Analytical purity: >= 98.5%
- Stability under test conditions: confirmed
No further data.

Test animals

Species:

rat

Strain:

Osborne-Mendel

Details on test animals or test system and environmental conditions:

TEST ANIMALS
1) female Osborne-Mendel (FDA strain) rats
- Source: FDA rat-breeding colony
- Age at study initiation: 99-140 days
- Weight at study initiation: 219-271 g on day 0 of the experiment (236.8 ± 0 .6 g, mean ± SEM)
- Fasting period before study: no
- Housing: individually in hanging cages with wire bottoms
- Diet (ad libitum): Purina Rat Chow
- Water (ad libitum): dosing/control solutions
- Acclimation period: no data

2) sexually mature male Osborne-Mendel (FDA strain) rats
- Age at study initiation: ca. 3 - 7 months

ENVIRONMENTAL CONDITIONS
- Temperature (°C): 21-24 ± 1 °C
- Humidity (%): 40-50 ± 10%
- Photoperiod (hrs dark / hrs light): 12/12

IN-LIFE DATES: no data

Administration / exposure

Route of administration:

oral: drinking water

Vehicle:

water

Details on exposure:

PREPARATION OF DOSING SOLUTIONS: no data

VEHICLE: distilled water

Analytical verification of doses or concentrations:

not specified

Details on mating procedure:

- Impregnation procedure: cohoused
- If cohoused:
- M/F ratio per cage: 1/1
- Length of cohabitation: overnight
- Verification of same strain and source of both sexes: yes
- Proof of pregnancy: finding of sperm referred to as day 0 of pregnancy

Duration of treatment / exposure:

days 0 through 20 of gestation

Frequency of treatment:

continuously in the drinking water

Duration of test:

until day 20 of gestation

Doses / concentrations

Doses / concentrations 1

Remarks:

Doses / Concentrations:
ca. 10.1, 27.4, 50.7, 86.6, 115.8, 160.9, 204.5 mg/kg bw/d
Basis:
actual ingested

Doses / concentrations 2

Remarks:

Doses / Concentrations:
70, 180, 360, 700, 1000, 1500, 2000 ppm in the drinking water
Basis:
nominal in water

No. of animals per sex per dose:

61 pregnant rats per group

Control animals:

yes, concurrent no treatment

Examinations

Maternal examinations:

CAGE SIDE OBSERVATIONS: No data

DETAILED CLINICAL OBSERVATIONS: No data

BODY WEIGHT: No data

FOOD CONSUMPTION: Yes
Food consumption was measured on days 7, 1 4 and 20

WATER CONSUMPTION AND COMPOUND INTAKE : Yes
The caffeine solution or distilled water was poured into glass bottles fitted with stainless-steel ball-point non-drip tubes, and the fluid was provided ad lib. from day 0 until the animals were killed on day 20. Water intake was determined by weight difference of the water bottle.

POST-MORTEM EXAMINATIONS: Yes
- Sacrifice on gestation day # 20
- Organs examined: uterus
- Females were examined for gross abnormalities prior to sacrifice

Ovaries and uterine content:

The ovaries and uterine content was examined after termination: Yes
Examinations included:
- Gravid uterus weight: No data
- Number of corpora lutea: Yes
- Number of implantations: Yes
- Number of early resorptions: Yes
- Number of late resorptions: Yes

Fetal examinations:

- External examinations: Yes: all per litter
Each live fetus was weighed, sexed and examined for gross external malformations under magnification, and the crown-rump length was measured. A runt was considered to be any fetus that weighed less than 70% of the average weight of the concurrent male or female control.
- Soft tissue examinations: Yes: half per litter
- Skeletal examinations: Yes: half per litter
- Head examinations: No data

Statistics:

Data were analysed by appropriate methods including
- Fisher's Exact test,
- Analysis of variance,
- Least significant difference (LSD) test (two-tail),
- Freeman-Tukey arc-sine transformation,
- Cox Exact Trend test.
Significance levels: less than or equal to 0.05, 0.01, 0.001 or 0.0001 as appropriate.

Results and discussion

Results: maternal animals

Maternal developmental toxicity

Details on maternal toxic effects:

Details on maternal toxic effects:
No overt behavioural differences were noted in any of the treated animals, and no dose-related gross effects were seen. Three females died during the experiment, one each from the groups given 360, 1500 or 2000 ppm caffeine. Autopsy findings of the three females that died were unremarkable.
At the four highest dose levels there was a dose-related decrease in fluid consumption, and at the three highest levels consumption was significantly lower than in the controls.
Food consumption was significantly lower than control in all dose groups.
Dose levels of 70, 180 and 360 ppm had no apparent effect on maternal weight gain during any period or on the total weight gained during gestation. Dose levels of 700, 1000, 1500 and 2000 ppm caused significantly low total maternal weight gains from day 0 to day 20.

Effect levels (maternal animals)

1

Dose descriptor:

NOAEL

Effect level:

ca. 50.7 mg/kg bw/day (actual dose received)

Basis for effect level:

other: maternal toxicity

2

Dose descriptor:

NOAEL

Effect level:

ca. 10.1 mg/kg bw/day (actual dose received)

Basis for effect level:

other: developmental toxicity

Results (fetuses)

Details on embryotoxic / teratogenic effects:

Details on embryotoxic / teratogenic effects:
Dosages of 1500 and 2000 ppm were associated with decreased implantation efficiency, increased resorptions and decreased mean numbers of viable fetuses. Numbers of runts were significantly increased after dosages of 1000 - 2000 ppm.
Fetal body weight and length were decreased and edematous fetuses were increased at dosages of 700 - 200 ppm. The test substance did not produce any dose-related gross anomalies. Only two animals with missing or hypoplastic nails were produced, both in the 1500 ppm. Sternebral ossification deficiencies were increased at all dose levels except 70 ppm. Skeletal ossification deficiencies were increased in a dose-related manner at the four highest dose levels.

Effect levels (fetuses)

Dose descriptor:

NOAEL

Effect level:

> 204.5 mg/kg bw/day (actual dose received)

Basis for effect level:

other: teratogenicity

Fetal abnormalities

Abnormalities:

not specified

Overall developmental toxicity

Developmental effects observed:

not specified

Applicant's summary and conclusion

Executive summary:

Caffeine dissolved in drinking-water was available ad libitum to Osborne-Mendel rats at dose levels of 0, 70, 180, 360, 700, 1000, 1500 or 2000 ppm during days 0-20 of gestation. The corresponding daily caffeine intakes were 0, 10.1, 27.4, 50.7, 86.6, 115.8, 160.9 and 204.5 mg/kg bw. Dosages of 160.9 and 204.5 mg/kg bw/d were associated with decreased implantation efficiency, increased resorptions and decreased mean numbers of viable fetuses. Numbers of runts were significantly increased after dosages of 115.8 - 204.5 mg/kg bw/d. Fetal body weight and length were decreased and edematous fetuses were increased at dosages of 86.6 - 204.5 mg/kg bw/d. Contrary to results seen after gavage studies, caffeine available ad libitum in drinking-water produced no dose-related gross anomalies. Only two animals with missing or hypoplastic nails were produced, both in the 160.9-mg/kg bw/d group. Sternebral ossification deficiencies were increased at all dose levels except 10.1 mg/kg bw/d. Skeletal ossification deficiencies were increased in a dose-related manner at the four highest dose levels. Caffeine given by water bottle produced ossification deficiencies similar to those seen after intubation, but at higher dosages.

NOAELs were 50.7, 10.1, and >204.5 mg/kg bw/d for maternal toxicity, developmental toxicity, and teratogenicity, respectively.