Registration Dossier

Administrative data

Endpoint:
developmental toxicity
Type of information:
experimental study
Adequacy of study:
key study
Reliability:
2 (reliable with restrictions)
Rationale for reliability incl. deficiencies:
study well documented, meets generally accepted scientific principles, acceptable for assessment

Data source

Referenceopen allclose all

Reference Type:
publication
Title:
Teratogenesis Studies with EDTA and its Salts in Rats
Author:
Schardein, J.L. et al
Year:
1981
Bibliographic source:
Toxicology and Applied Pharmacology 61, 423-428 (1981)
Reference Type:
secondary source
Title:
Teratogenesis Studies with EDTA and its Salts in Rats
Author:
Schardein, J.L. et al
Year:
1981
Bibliographic source:
Toxicol. Appl. Pharmacol., 61, 423-428, (1981) Cited in: Bibra Bulletin 21, No. 1, pp 12-13, (1982)

Materials and methods

Test guideline
Qualifier:
equivalent or similar to guideline
Guideline:
OECD Guideline 414 (Prenatal Developmental Toxicity Study)
Version / remarks:
study was conducted before guideline was adopted
Deviations:
yes
Remarks:
treatment from gestation day 7 to 14
Principles of method if other than guideline:
Edetic acid (EDTA) and four of its salts, disodium, trisodium, calcium di-sodium, and tetrasodium EDTA, were studied for teratogenic potential in rats. Equimolar doses based on 1000 mg/kg were given by gastric intubation on days 7 to 14 of gestation. On day 21 of gestation the dams of each group were sacrificed and litter data for each dam collected.
GLP compliance:
no
Limit test:
yes

Test material

Constituent 1
Chemical structure
Reference substance name:
Trisodium hydrogen ethylenediaminetetraacetate
EC Number:
205-758-8
EC Name:
Trisodium hydrogen ethylenediaminetetraacetate
Cas Number:
150-38-9
Molecular formula:
C10H16N2O8.3Na
IUPAC Name:
trisodium hydrogen 2,2',2'',2'''-(ethane-1,2-diyldinitrilo)tetraacetate
Test material form:
solid
Details on test material:
Name in the publication: trisodium edetate

Test animals

Species:
rat
Strain:
other: CD albino
Details on test animals or test system and environmental conditions:
TEST ANIMALS
- Source: Charles River Breeding Laboratories
- Weight at study initiation: mean weight 241 g
- Diet: Purina Lab Chow ad libitum
- Water: tap water ad libitum


Administration / exposure

Route of administration:
oral: gavage
Vehicle:
other: 0.2 M phosphate buffer
Details on exposure:
- pH of dosing solution: 7.9

Analytical verification of doses or concentrations:
no
Details on mating procedure:
- Impregnation procedure: cohoused
- If cohoused:
- M/F ratio per cage: 1:1
- Proof of pregnancy: vaginal plug, sperm in vaginal smear referred to as day 0 of pregnancy
Duration of treatment / exposure:
8 days (day 7 to day 14 of gestation)
Frequency of treatment:
daily, the total daily dose was equally divided into two daily doses
Duration of test:
21 days
Doses / concentrations
Dose / conc.:
1 245 mg/kg bw/day (actual dose received)
Remarks:
divided into two equal daily doses
No. of animals per sex per dose:
20
Control animals:
yes, concurrent no treatment
yes, concurrent vehicle
Details on study design:
- Dose selection rationale: Dose levels were chosen from preliminary studies with EDTA in which there was some evidence of both maternal and fetotoxicity at 1000 mg/kg bw/day.

Examinations

Maternal examinations:
CAGE SIDE OBSERVATIONS: Yes

BODY WEIGHT: Yes
- Time schedule for examinations: once a week

FOOD CONSUMPTION: Yes

POST-MORTEM EXAMINATIONS: Yes
- Sacrifice on gestation day 21



Ovaries and uterine content:
The ovaries and uterine content was examined after termination: Yes
Examinations included:
- Number of implantations: Yes
- Number of early resorptions: Yes
- Number of late resorptions: Yes
Fetal examinations:
- External examinations: each fetus
- Gross inspection, slicing and visceral abnormalties: 1/3 of the litter
- Skeletal examinations: 2/3 of the litter
Statistics:
Means and standard errors were calculated for litter size, followed by analysis of variance. Pre- and postimplantation losses, embryonic viability, and fetal survival were evaluated by analysis of covariance. Fetal weights were also evaluated by analysis of covariance following calculation of mean weight/litter by sex, the values representing means and standard errors of mean litter weights. Significant variance by either analysis of variance or covariance was further evaluated by Dunnett's t test to locate the source of variance. Sex distribution was analyzed by partitioned a chi-squeared test.

Results and discussion

Results: maternal animals

General toxicity (maternal animals)

Clinical signs:
effects observed, treatment-related
Description (incidence and severity):
Significant drug-related reactions included diarrhea and reduced activity. Diarrhea generally occurred daily following treatment and ultimately disappeared on the last day of dosing or the day after. The incidence of diarrhea was 35%. Transient episodes of reduced activity were seen on the first day of dosing in one dam.
Dermal irritation (if dermal study):
not examined
Mortality:
mortality observed, non-treatment-related
Description (incidence):
Two dams had to be sacrificed as a result of a dosing error (intubarion into the lungs).
Body weight and weight changes:
effects observed, treatment-related
Description (incidence and severity):
Treatment with the test item caused reduced body weight gain. Recovery was evident once treatment was terminated (see Table 1).


Food consumption and compound intake (if feeding study):
effects observed, treatment-related
Description (incidence and severity):
During treatment (gestation days 7-14) food intake was slihghtly decreased in the test item groups compared to the vehicle and untreated controls. During the post-treatment period (gestation days 14-21) food intake for the treated dams was similar to or slightly greater than that of dams in both control groups (see Table 1).
Food efficiency:
not examined
Water consumption and compound intake (if drinking water study):
not examined
Ophthalmological findings:
not examined
Haematological findings:
not examined
Clinical biochemistry findings:
not examined
Urinalysis findings:
not examined
Behaviour (functional findings):
not examined
Immunological findings:
not examined
Organ weight findings including organ / body weight ratios:
not examined
Gross pathological findings:
not specified
Neuropathological findings:
not examined
Histopathological findings: non-neoplastic:
not examined
Histopathological findings: neoplastic:
not examined
Other effects:
not examined

Maternal developmental toxicity

Number of abortions:
no effects observed
Description (incidence and severity):
No abortions were noted.
Pre- and post-implantation loss:
no effects observed
Total litter losses by resorption:
no effects observed
Early or late resorptions:
no effects observed
Dead fetuses:
no effects observed
Changes in pregnancy duration:
no effects observed
Changes in number of pregnant:
no effects observed

Effect levels (maternal animals)

open allclose all
Key result
Dose descriptor:
LOAEL
Remarks:
general maternal toxicity
Effect level:
1 245 mg/kg bw/day (actual dose received)
Based on:
test mat.
Basis for effect level:
other: diarrhea, slightly reduced food intake, slightly reduced body weight gain
Key result
Dose descriptor:
NOAEL
Remarks:
developmental toxicity
Effect level:
>= 1 245 mg/kg bw/day (actual dose received)
Based on:
test mat.
Basis for effect level:
other: no adverse developmental effects noted

Results (fetuses)

Fetal body weight changes:
no effects observed
Description (incidence and severity):
Fetal body weights were similar to those of the two control groups (see Table 2).
Reduction in number of live offspring:
no effects observed
Description (incidence and severity):
Mean litter size and number of live fetuses per dam were similar to those in the two control groups (see Table 2).
Changes in sex ratio:
no effects observed
Description (incidence and severity):
The sex ratio was close to 50% in all groups (see Table 2).
Changes in litter size and weights:
not examined
Changes in postnatal survival:
not examined
External malformations:
effects observed, non-treatment-related
Description (incidence and severity):
A total of 1084 fetuses from EDTA and EDTA salt-treated dams were examined. These were compared to 237 fetuses from 19 dams treated with the vehicle and 278 fetuses from 20 untreated dams.
One untreated control fetus had multiple abnormalities including eye defect, ectrodactyly, and a curly tail.
Skeletal malformations:
effects observed, non-treatment-related
Description (incidence and severity):
Only summary data from all EDTA and EDTA salt--treated groups were reported. Of the 752 fetuses treated with EDTA or its salts and were skeletally examined 24 fetuses has skeletal abnormalities. These included 20 with bifid vertebra, 1 with agenesis of ribs, 2 with inhibition of osteogenesis of the shull or ribs and 1 with malformed ribs. There was no definitive pattern regarding treatment with a particular compound and the occurrence of anomalies.

None of the fetuses in the vehicle control group had abnormalities while 8 untreated control fetuses exhibited some major defect.
One untreated control fetus was stunted and had multiple abnormalities including ectrodactyly, and a curly tail. Five additional control fetuses had bifid vertebrae while 2 had malformed vertebrae or sternebrae.
Visceral malformations:
no effects observed
Description (incidence and severity):
On untreated control fetus had multiple abnormalities including eye defect. Histological examination of the eyes revealed a cataract in one eye and a dysmorphic lens and retina in the other.
Other effects:
not examined

Effect levels (fetuses)

Key result
Dose descriptor:
NOAEL
Effect level:
>= 1 245 mg/kg bw/day (actual dose received)
Basis for effect level:
other: no adverse treatment-related effects noted

Fetal abnormalities

Abnormalities:
no effects observed

Overall developmental toxicity

Developmental effects observed:
no

Any other information on results incl. tables

Table 1: Food consumption and weight gain in rats treated with EDTA and EDTA salts on days 7 through 14 of gestation

Days Control (untreated) Vehicle Control 967 mg/kg bw/day EDTA 1243 mg/kg bw/day disodium EDTA 1245 mg/kg bw/day trisodium EDTA 1340 mg/kg bw/day calcium disodium EDTA 1374 mg/kg bw/day tetrasodium EDTA
Mean food consumption (g/day)
0-7 20.7 21.4 19.9 20.4 19.9 20.4 19.0
7-14 22.3 22.5 18.4 17.5 19.1 20.9 18.5
14-21 24.7 25.3 26.7 27.2 25.7 26.5 25.6
Mean body weight gain (g)
0-7 31.9 35.1 37.6 35.6 33.2 37.2 26.9
7-14 28.5 26.1 16.5 13.7 20.4 25.1 18.3
14-21 102.7 93.3 104.4 100.2 98.4 108.1 94.2

Table 2: Reproductive data in dams receiving EDTA and EDTA salts on days 7 through 14 of gestation

Fetuses
Sex Body weight (mean g ± SE) Number
Treatment No of dams Litter size (mean ± SE) Post implantation loss (%) M F M F Dead Live Resorbed
Control (untreated) 20 14.0 ± 0.4 4 52 48 5.3 ± 0.1 5.6 ± 0.1 1 278 12
Vehicle Control 19 12.5 ± 0.1 3 50 50 5.3 ± 0.1 5.7 ± 0.1 0 237 7
967 mg/kg bw/day EDTA 17 12.7 ± 0.6 3 49 51 5.5 ± 0.1 5.7 ± 0.1 0 216 6
1243 mg/kg bw/day Na2 EDTA 19 12.6 ± 0.4 1 56 44 5.4 ± 0.1 5.6 ± 0.1 0 202 3
1245 mg/kg bw/day Na3 EDTA 18* 12.4 ± 1.0 8 48 52 5.4 ± 0.2 5.7 ± 0.1 0 210 11
1340 mg/kg bw/day CaNa2 EDTA 17 13.5 ± 0.4 2 52 48 5.3 ± 0.1 5.6 ± 0.1 0 230 4
1374 mg/kg bw/day Na4 EDTA 19 11.9 ± 0.7 4 47 52 5.2 ± 0.1 5.5 ± 0.1 0 226 10

* 2 dams had to be killed due to dosing errors

Applicant's summary and conclusion

Conclusions:
No test item-related effects on embryo-fetal development could be detected in the study. Under the conditions of the test the NOAEL for maternal and embryo-featl developmental toxicity in rats is >= 1245 mg/kg bw/day.