Iron trinitrate

Repeated dose toxicity: oral

Administrative data

Endpoint:

sub-chronic toxicity: oral

Type of information:

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

Adequacy of study:

key study

Reliability:

2 (reliable with restrictions)

Rationale for reliability incl. deficiencies:

other: The study was conducted according to an appropriate OECD test guideline, with acceptable restrictions. No information on the GLP status of the study.

Data source

Reference

Reference Type:

publication

Title:

Lack of Carcinogenicity of Ferric Chloride in F344 Rats.

Author:

Sato M, Furukawa F, Toyoda K, Mitsumori K, Nishikawa A, Takahashi M

Year:

1992

Bibliographic source:

Food and Chemical Toxicology 30(10): 837-842

Materials and methods

Test guideline

Qualifier:

equivalent or similar to guideline

Guideline:

OECD Guideline 408 (Repeated Dose 90-Day Oral Toxicity Study in Rodents)

Deviations:

yes

Remarks:

No information on GLP, limited details on environmental conditions, limited details on examinations conducted, and no urinalysis.

Principles of method if other than guideline:

Range-finding study performed for a 2-year carcinogenicity study

GLP compliance:

not specified

Limit test:

no

Test material

Test material information

Constituent 1

Reference substance name:

ferric chloride hexahydrate

IUPAC Name:

ferric chloride hexahydrate

Test material form:

other: solid

Details on test material:

- Name of test material (as cited in study report): Ferric chloride hexahydrate
- Substance type: Iron salt
- Physical state: Solid
- Analytical purity: 98.5%
- Impurities (identity and concentrations): No data
- Composition of test material, percentage of components: No data
- Purity test date: No data
- Lot/batch No.: No data
- Expiration date of the lot/batch: No data
- Stability under test conditions: It was confirmed that a 2% solution of ferric chloride in distilled water was stable for one week at room temperature.
- Storage condition of test material: No data

Test animals

Species:

rat

Strain:

Fischer 344

Sex:

male/female

Details on test animals or test system and environmental conditions:

TEST ANIMALS
- Source: Charles River Japan Inc
- Age at study initiation: Six weeks
- Weight at study initiation: No data
- Fasting period before study: No data
- Housing: No data
- Diet (e.g. ad libitum): Ad libitum
- Water (e.g. ad libitum): Ad libitum
- Acclimation period: One week


ENVIRONMENTAL CONDITIONS
- Temperature (°C): No data
- Humidity (%): No data
- Air changes (per hr): No data
- Photoperiod (hrs dark / hrs light): No data


IN-LIFE DATES: No data

Administration / exposure

Route of administration:

oral: drinking water

Vehicle:

water

Details on oral exposure:

PREPARATION OF DOSING SOLUTIONS: Ferric chloride was dissolved in distilled water at concentrations of 0 (control), 0.12, 0.25, 0.5, 1.0 or 2.0 % (w/v).

Analytical verification of doses or concentrations:

not specified

Duration of treatment / exposure:

13 weeks

Frequency of treatment:

Continuous

Doses / concentrations

Doses / concentrations 1

Remarks:

Doses / Concentrations:
0.12, 0.25, 0.5, 1.0 and 2.0% w/v
Basis:
nominal in water

Doses / concentrations 2

Remarks:

Doses / Concentrations:
80, 154, 277, 550 and 1231 mg/kg bw/day in male rats, 88, 176, 314, 571 and 1034 mg/kg bw/day in female rats
Basis:
other: approximate dose received

No. of animals per sex per dose:

Ten

Control animals:

other: Drinking water only

Details on study design:

- Dose selection rationale: This is a dose-range finding study to determine doses for a two-year carcinogenicity study.
- Rationale for animal assignment (if not random): No data
- Rationale for selecting satellite groups: No satellite group
- Post-exposure recovery period in satellite groups: None
- Section schedule rationale (if not random): No data

Positive control:

No

Examinations

Observations and examinations performed and frequency:

CAGE SIDE OBSERVATIONS: Yes
- Time schedule: Daily

DETAILED CLINICAL OBSERVATIONS: No data

BODY WEIGHT: Yes
- Time schedule for examinations: Weekly

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: No
- 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): Yes


OPHTHALMOSCOPIC EXAMINATION: No


HAEMATOLOGY: Yes
- Time schedule for collection of blood: At the end of the exposure period
- Anaesthetic used for blood collection: No data
- Animals fasted: No data
- How many animals: All survivors
- Parameters examined: no data


CLINICAL CHEMISTRY: Yes
- Time schedule for collection of blood: At the end of the exposure period
- Animals fasted: No data
- How many animals: All survivors
- Parameters examined: no data

URINALYSIS: No

NEUROBEHAVIOURAL EXAMINATION: No

Sacrifice and pathology:

GROSS PATHOLOGY: Yes, for all survivors. No further details.
HISTOPATHOLOGY: Yes, all major organs and tissues. No further details.

Other examinations:

no further information

Statistics:

Body weight and the daily water intake data were analysed statistically using Student's t-test.

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):

effects observed, treatment-related

Ophthalmological findings:

not examined

Haematological findings:

effects observed, treatment-related

Clinical biochemistry findings:

effects observed, treatment-related

Urinalysis findings:

not examined

Behaviour (functional findings):

not examined

Organ weight findings including organ / body weight ratios:

not examined

Gross pathological findings:

no effects observed

Histopathological findings: non-neoplastic:

effects observed, treatment-related

Histopathological findings: neoplastic:

no effects observed

Details on results:

CLINICAL SIGNS AND MORTALITY: No deaths or clinical signs of toxicity were observed.


BODY WEIGHT AND WEIGHT GAIN: In the 1.0 and 2.0% groups there was a reduced body weight gain of at least 10% compared with the controls at the termination of treatment.


WATER CONSUMPTION AND COMPOUND INTAKE (if drinking water study): There was a significant suppression in the intake of drinking water observed in groups given concentrations of 0.5% or more (no further detail).


HAEMATOLOGY: Males of the treated groups had significantly increased red blood cell counts (no further details).


CLINICAL CHEMISTRY: Higher levels of serum iron were observed in males (control: 102±5 g/dl; 0.12%: 107±11 g/dl;
0.25%: 109±8  g/dl; 0.5%: 129±22  g/dl; 1.0%: 139±10  g/dl; 2.0%: 156 g/dl).


GROSS PATHOLOGY: No findings reported.


HISTOPATHOLOGY: NON-NEOPLASTIC: In examinations of sections stained with haematoxylin and eosin, brown pigment deposition was observed only in the keratin layers of the oesophageal mucosa in the groups given concentrations of 0.25% or higher, and in the laminae propriae of the large intestine in the 2.0% group. In sections stained with Berlin blue, increased numbers of positive pigments were also observed in the hepatocytes and Kupffer cells of the liver, the cartilage of the trachea and bronchus, the keratin mucosal layers of the tongue, the forestomach, the mucous layers of the small intestines, the white pulp of the spleen, the tubular epithelium of the kidney and the adipose tissues of the groups given 0.25% and higher. The intensity of the staining was marked in the intestine and liver.


HISTOPATHOLOGY: NEOPLASTIC: not investigated in this range-finding study.

Effect levels

1

Dose descriptor:

NOAEL

Effect level:

0.5 other: %

Based on:

test mat.

Sex:

male/female

2

Dose descriptor:

NOAEL

Effect level:

5 000 ppm

Based on:

test mat.

Sex:

male/female

3

Dose descriptor:

NOAEL

Effect level:

>= 277 - <= 314 mg/kg bw/day (actual dose received)

Based on:

test mat.

Sex:

male/female

Basis for effect level:

other: Equivalent to 277 and 314 mg/kg bw/day in males and females, respectively.

4

Dose descriptor:

NOAEL

Effect level:

>= 57 mg/kg bw/day (actual dose received)

Based on:

element

Sex:

male/female

Basis for effect level:

other: NOAEL for Fe (III), based on iron content in the test material

Target system / organ toxicity

Critical effects observed:

not specified

Any other information on results incl. tables

Doses: The dose of ferric chloride in mg/kg bw/day was calculated from the drinking water intake and the final weight of male and female rats

Dose of ferric chloride hexahydrate		Final weight (kg)		Final daily water intake (ml/d)		Dose (mg/kg bw/day)
%	mg/l	Males	Females	Males	Females	Males	Females
0	0	0.35	0.195	23.5	15	0	0
0.12	1.2	0.345	0.19	23	14	80	88
0.25	2.5	0.34	0.185	21	13	154	176
0.5	5.0	0.325	0.175	18	11	277	314
1.0	10.0	0.3	0.175	16.5	10	550	571
2.0	20.0	0.195	0.145	12	7.5	1231	1034

Applicant's summary and conclusion

Conclusions:

ferric chloride hexahydrate was administered to rats in their drinking water for 13 weeks, the NOAEL was 0.5% (equivalent to 277 and 314 mg/kg bw/day in males and females, respectively).
The NOAEL was 0.5% (equivalent to 277 and 314 mg/kg bw/day in males and females, respectively) based on the reduced body weight gain.
The equivalent NOAEL doses expressed as iron trinitrate would be 246 mg/kg bw/day for males and 281 mg/kg bw/day for females.

Executive summary:

Ferric chloride hexahydrate was administered to Fischer 344 rats (10/sex/dose) in their drinking water for 13 weeks, at concentrations of 0.12, 0.25, 0.5, 1.0 and 2.0% (equivalent to approximately 80, 154, 277, 550 and 1231 mg/kg bw/day in male rats, 88, 176, 314, 571 and 1034 mg/kg bw/day in female rats). All deaths and clinical signs of toxicity were recorded. Body weights were measured weekly. At the end of the dosing period, all survivors were killed for haematological, clinical chemistry and pathological exminations. All major organs and tissues were examined microscopically.

There were no deaths or clinical signs of toxicity. There was a significant reduction in body weight gains at the two highest doses at the end of the treatment period. Treated males had increased levels of serum iron and higher red blood cell counts compared with controls. In microscopic examinations of sections stained with haematoxylin and eosin, brown pigment deposition was observed only in the keratin layers of the oesophageal mucosa in the groups given concentrations of 0.25% or higher, and in the laminae propriae of the large intestine in the 2.0% group. In sections stained with Berlin blue, increased numbers of positive pigments (an indication of iron overload) were also observed in the hepatocytes and Kupffer cells of the liver, the cartilage of the trachea and bronchus, the keratin mucosal layers of the tongue, the forestomach, the mucous layers of the small intestines, the white pulp of the spleen, the tubular epithelium of the kidney and the adipose tissues of the groups given 0.25% and higher. The intensity of the staining was marked in the intestine and liver. The NOAEL was 0.5% (equivalent to 277 and 314 mg/kg bw/day in males and females, respectively) based on the reduced body weight gain.