Iron trinitrate

Developmental toxicity / teratogenicity

Administrative data

Endpoint:

developmental toxicity

Type of information:

migrated information: read-across from supporting substance (structural analogue or surrogate)

Adequacy of study:

key study

Study period:

No data

Reliability:

1 (reliable without restriction)

Rationale for reliability incl. deficiencies:

other: The study was well documented and meets generally accepted scientific principles, and conducted in compliance with GLP.

Data source

Reference

Reference Type:

study report

Title:

Unnamed

Year:

2002

Materials and methods

Test guideline

Qualifier:

according to guideline

Guideline:

other: OECD 422

Deviations:

no

Principles of method if other than guideline:

Refer to summary presented in section 7.8.1

GLP compliance:

yes

Limit test:

no

Test material

Test material information

Constituent 1

Reference substance name:

7782-63-0

EC Number:

616-510-7

Cas Number:

7782-63-0

IUPAC Name:

7782-63-0

Constituent 2

Reference substance name:

Iron (II) sulfate heptahydrate

IUPAC Name:

Iron (II) sulfate heptahydrate

Test material form:

solid: crystalline

Details on test material:

- Name of test material (as cited in study report): Iron (II) sulfate heptahydrate
- Physical state: blue-green crystalline powder
- Analytical purity: 91.1% (Lot No. 010628) and 90.6% (Lot No. 010903)
- Impurities (identity and concentrations): Mg 0.3%, Mn 0.19%, Zn 75 ppm (Lot No. 010628) and Mg 0.28%, Mn 0.16%, Zn 84 ppm (Lot No. 010903)
- Storage condition of test material: room temperature under argon
- Stability under test conditions: The stability of test concentrations was confirmed for 6 hours in the dark at room temperature.

Test animals

Species:

rat

Strain:

Sprague-Dawley

Details on test animals or test system and environmental conditions:

TEST ANIMALS
- Source: Charles River Laboratories Japan, Inc., Yokohama, Japan
- Age at study initiation: 10 weeks
- Weight at study initiation: male: 341 -383 g; female: 222 -255 g
- Housing: stainless steel cage
- Diet (ad libitum): CRF-1 from Oriental Yeast Co., Ltd.
- Water (ad libitum): tap water
- Acclimation period: 12 days

ENVIRONMENTAL CONDITIONS
- Temperature (°C): 20 - 26
- Humidity (%): 40 - 70
- Air changes (per hr): 12
- Photoperiod (hrs dark / hrs light): 12 / 12

Administration / exposure

Route of administration:

oral: gavage

Vehicle:

water

Details on exposure:

PREPARATION OF DOSING SOLUTIONS: Test substance was prepared with water for injection purposes. Gavage solutions were prepared freshly each time and used within 6 hours.

Analytical verification of doses or concentrations:

yes

Details on analytical verification of doses or concentrations:

The concentrations were measured in samples used for males at the first treatment and at the end of administration.

Details on mating procedure:

- M/F ratio per cage: 1/1
- Length of cohabitation: up to 14 days
- Proof of pregnancy: vaginal plug / sperm in vaginal smear referred to as day 0 of pregnancy

Duration of treatment / exposure:

Males: Total of 49 days beginning 14 days before mating.
Females: Total of 42-47 days beginning 14 days before mating.

Frequency of treatment:

Once daily

Duration of test:

49 days

Doses / concentrations

Remarks:

Doses / Concentrations:
30, 100, 300, 1000 mg/kg bw/day
Basis:
actual ingested

No. of animals per sex per dose:

12

Control animals:

yes, concurrent vehicle

Details on study design:

- Dose selection rationale: Doses selected for the main studies were based on gross pathology findings observed in the 14-days preliminary studies (Study No. 100520P).

Examinations

Maternal examinations:

Observation performed and frequency: General condition was observed twice a day (before and after treatment). 

Male body weights were  measured twice a week. Female body weights were measured twice a week  during the pre-mating and mating period and on days 0, 7, 14 and 21 of gestation and days 0 and 4 of lactation. 

Food consumption was determined twice a week during the pre- and post-mating period in males, and twice a week during the pre-mating period and on days 2, 9, 16 and 21 of gestation and day 4 of lactation in females. 

For 6 males and females per group, urinalysis was carried out before final administration. 

Blood samples were collected from the abdominal aorta on next day of the last  administration and hematological  and blood chemical examinations were carried out. 

Ovaries and uterine content:

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

Fetal examinations:

- External examinations: Yes:
- Soft tissue examinations: No data
- Skeletal examinations: No data
- Head examinations: No data

Statistics:

Bartlett´s test, Dunnett´s test

Indices:

copulation index, fertility index, gestation index, delivery index, implantation index, live birth index and viability index

Historical control data:

No information

Results and discussion

Results: maternal animals

Maternal developmental toxicity

Details on maternal toxic effects:

Maternal toxic effects:yes

Details on maternal toxic effects:
CLINICAL SIGNS AND MORTALITY (PARENTAL ANIMALS)
males: 1000 mg/kg bw/day: 1 died on day 27 and salivation; 300 mg/kg bw/day: salivation
females: 1000 mg/kg bw/day: 1 died on day 19 and salivation; 300 mg/kg bw/day: salivation

BODY WEIGHT
males: 1000 mg/kg bw/day: reduced between days 11 and 49;
females: 1000 mg/kg bw/day: reduced during pregnancy on day 21 (not significant)

FOOD CONSUMPTION
males/females: 1000 mg/kg bw/day: reduced on day 3

REPRODUCTIVE FUNCTION: ESTROUS CYCLE (PARENTAL ANIMALS)
no effect on estrous cycle


REPRODUCTIVE PERFORMANCE (PARENTAL ANIMALS)
- All 12 pairs of each dose group copulated successfully (copulation index 100%).
- All females except one female became pregnant (fertility index: 100).
-The gestation index was 100% for all dose groups

ORGAN WEIGHTS (PARENTAL ANIMALS)
- males: 1000 mg/kg bw/day: absolute and relative weight of adrenals, relative weight of liver increased; absolute testes weight increased at 300 mg/kg bw/day, but not at 1000 mg/kg bw/day;
absolute weights of pituitary and heart were decreased; relative weight of brain and testes increased: these changes are considered to be due to the significant body weight loss

- females: 1000 mg/kg bw/day: absolute and relative weight of liver increased
relative weight of uterus increased at 1000 mg/kg bw/day, but this was considered to be due to the significant body weight loss.

- Significant higher absolute testis weight observed in males exposure to the 300 mg/kg bw/day, but not in males treated with 1000 mg/kg bw/day. This increase is not considered to be treatment related. The statistically significant higher relative testis weights (1000 mg/kg bw/d) may be due to reduced body weights. No effect on epididymis weights, no abnormalities in testes, epididymides and prostates observed.


GROSS PATHOLOGY (PARENTAL ANIMALS)
- males:
1000 mg/kg bw/day: thymus: atrophy in 2 animals; stomach inflammation and ulcers in glandular stomach (1 animal); bleeding (1 case); inflammatory cell infiltration in submucosal glandular stomach (2 cases); vacuolization of the forestomach epithelium (1 case); liver: yellow-brown pigmentation of periportal hepatocytes (6 cases); pigmentation of periportal Kupffer cells (3 cases): probably due to iron; spleen: extramedullary hematopoiesis (4 cases); yellow-brown pigmentation in red pulp (6 cases); kidney: basophilic changes in tubular epithelium (4 cases); bone marrow: hematopoiesis in the femur (1 case)
300 mg/kg bw/day: spleen: yellow-brown pigmentation in the red pulp (6 cases); extramedullary hematopoiesis (5 cases)

100 mg/kg bw/day: spleen: yellow-brown pigmentation in the red pulp (6 cases); extramedullary hematopoiesis (2 cases); kidney: basophilic changes in tubular epithelium (1 case)

30 mg/kg bw/day: spleen: yellow-brown pigmentation in the red pulp (6 cases); extramedullary hematopoiesis (1 cases); kidney: basophilic changes in tubular epithelium (2 cases)
Control: yellow-brown pigmentation in the red pulp (6 cases); extramedullary hematopoiesis (2 cases)

dead males:
1000 mg/kg bw/day: mineral deposits in heart, lung congestion, pigmentation of periportal hepatocytes
The adrenal glands showed abnormalities (abnormal growth) in the autopsy.

- females:
1000 mg/kg bw/day: liver: yellow-brown pigmentation of periportal hepatocytes (6 cases) probably due to iron; spleen: yellow-brown pigmentation in the red pulp (6 cases); extramedullary hematopoiesis (6 cases)

300 mg/kg bw/day: spleen: yellow-brown pigmentation in the red pulp (6 cases); extramedullary hematopoiesis (6 cases)

100 mg/kg bw/day: spleen: yellow-brown pigmentation in the red pulp (6 cases); extramedullary hematopoiesis (6 cases)

30 mg/kg bw/day: spleen: yellow-brown pigmentation in the red pulp (5 cases); extramedullary hematopoiesis (6 cases)

Control: spleen: yellow-brown pigmentation in the red pulp (6 cases); extramedullary hematopoiesis (6 cases)

dead females:
Lung congestion and edema, mineral deposits in liver
Abnormalities in pituitary (mass), adrenal (enlargement) and thymus (atrophy) were found at autopsy.

HISTOPATHOLOGY (PARENTAL ANIMALS)
males testis:
-mild atrophy of seminiferous tubules: 1/7 animals at 1000 mg/kg bw/d
-moderate atrophy of seminiferous tubules: 1/7 animals at 1000 mg/kg bw/d
-mild hyperplasia of Leydig’s cell: 1/6 animals at 1000 mg/kg bw/d
-mild degeneration of seminiferous tubules: 1/6 animals at control and 1/7 at 1000 mg/kg bw/d
-moderate degeneration of seminiferous tubules: 1/6 animals at control and 1/7 at 1000 mg/kg bw/d
-moderate exfoliated round spermatids of seminiferous tubules: 1/6 animals in the control group and 1/7 at 1000 mg/kg bw/d

males epididymis:
-marked emptiness: 1/7 animals 1/7 at 1000 mg/kg bw/d

males prostate:
-mild lymphoid cell infiltration: 3/6 animals in the control groups
-moderate lymphoid cell infiltration: 3/6 animals at 1000 mg/kg bw/d

The effects observed in histopathology in male reproductive organs occured spontaneous and were not considered compound-related.

females: histopathological examinations of the ovary was not carried out.

Parent animal reproduction:
Reproductive performance displayed no significant changes between treatment groups and controls (number of estrous cases, copulation index, number of days before copulation, fertility index, gestation length, gestation index, delivery conditions, nursing conditions, number of corpora lutea, number of implantation sites, or implantation rate).

Effect levels (maternal animals)

1

Dose descriptor:

NOAEL

Effect level:

100 mg/kg bw/day (actual dose received)

Based on:

test mat.

Basis for effect level:

other: maternal toxicity

2

Dose descriptor:

NOAEL

Effect level:

>= 1 000 mg/kg bw/day (actual dose received)

Based on:

test mat.

Basis for effect level:

other: developmental toxicity

Results (fetuses)

Details on embryotoxic / teratogenic effects:

Embryotoxic / teratogenic effects:no effects

Fetal abnormalities

Abnormalities:

not specified

Overall developmental toxicity

Developmental effects observed:

not specified

Applicant's summary and conclusion

Conclusions:

In a well conducted OECD 422 study conducted to GLP (reliability score 1), the NOAEL for developmental toxicity was at least 1000 mg/kg bw/day iron sulphate heptahydrate (equivalent to 200 mg/kg bw/day iron) in rats. The NOAEL for parental toxicity was 100 mg/kg bw/day (equivalent to 20 mg/kg bw/day iron).

Executive summary:

In a well conducted OECD 422 study conducted to GLP (reliability score 1), the NOAEL for developmental toxicity was at least 1000 mg/kg bw/day iron sulphate heptahydrate (equivalent to 200 mg/kg bw/day iron) in rats. The NOAEL for parental toxicity was 100 mg/kg bw/day (equivalent to 20 mg/kg bw/day iron).