Ammonium carbamate

Repeated dose toxicity: oral

Administrative data

Endpoint:

short-term repeated dose toxicity: oral

Type of information:

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

Adequacy of study:

weight of evidence

Reliability:

2 (reliable with restrictions)

Rationale for reliability incl. deficiencies:

other: Comparable to guideline standards with acceptable restrictions: - only two doses, incomprehensive reporting

Cross-reference

Reason / purpose for cross-reference:

reference to same study

Data source

Reference

Reference Type:

publication

Title:

Toxicity and carcinogenicity of acidogenic or alkalogenic diets in rats; effects of feeding NH4Cl, KHCO3 or KCl

Author:

Lina, B.A.R. and Kuijpers, M.H.M.

Year:

2004

Bibliographic source:

Food and Chemical Toxicology 42, 135 - 153

Materials and methods

Test guideline

Qualifier:

equivalent or similar to guideline

Guideline:

OECD Guideline 407 (Repeated Dose 28-Day Oral Toxicity Study in Rodents)

Deviations:

yes

Remarks:

(only two doses tested, no satellite recovery group, no neurobahavioural tests, no opthalmological test)

GLP compliance:

not specified

Limit test:

no

Test material

Test material information

Constituent 1

Reference substance name:

Ammonium chloride

EC Number:

235-186-4

EC Name:

Ammonium chloride

Cas Number:

12125-02-9

Molecular formula:

ClH4N

IUPAC Name:

ammonium chloride

Details on test material:

- Name of test material (as cited in study report): Ammonium chloride (NH4Cl)
- Analytical purity: >99.5%
- Source: Merck, Darmstadt, Germany
- Catalogue No.:1145

Test animals

Species:

rat

Strain:

Wistar

Sex:

male/female

Details on test animals or test system and environmental conditions:

TEST ANIMALS
- Source: TNO Central Institute for the Breeding of Laboratory Animals, Zeist, the Netherlands
- Age at study initiation: 5 weeks of age
- Weight at study initiation: no data
- Fasting period before study: no data
- Housing: groups of five of the same sex and the same treatment group, in suspended stainless steel cages with wire-mesh floor and front
- Diet: ad libitum; the Institute’s cereal based rodent diet (Rutten and De Groot, 1992, Food and Chemical Toxicology 30, 601–610).
- Water: ad libitum; tap water
- Acclimation period: no data

ENVIRONMENTAL CONDITIONS
- Temperature (°C): 22+/- 2°C
- Humidity (%): 35–70%
- Air changes (per hr): at least 10/h
- Photoperiod (hrs dark / hrs light): 12/12

Administration / exposure

Route of administration:

oral: feed

Vehicle:

unchanged (no vehicle)

Analytical verification of doses or concentrations:

not specified

Duration of treatment / exposure:

4 weeks

Frequency of treatment:

daily

Doses / concentrations

Doses / concentrations 1

Remarks:

Doses / Concentrations:
2 and 4% NH4Cl
Basis:
nominal in diet

Doses / concentrations 2

Remarks:

Doses / Concentrations:
2214.5 and 4228.5 mg/kg bw/d calculated from food consumption
Basis:
actual ingested

No. of animals per sex per dose:

10

Control animals:

yes, plain diet

Details on study design:

- Dose selection rationale: no data

Examinations

Observations and examinations performed and frequency:

DETAILED CLINICAL OBSERVATIONS: Yes
- Clinical signs and survival were checked daily

BODY WEIGHT: Yes
- Body weights of the individual rats were determined weekly

FOOD CONSUMPTION AND COMPOUND INTAKE:
- Food intake of the five rats in each cage was determined weekly over a one week period

WATER CONSUMPTION A: Yes
- Time schedule for examinations: recorded daily per cage in week 1

HAEMATOLOGY: Yes
- Time schedule for collection of blood: the haematological examinations were conducted in week 4
- How many animals: all
- Parameters checked: haemoglobin concentration, packed cell volume, red blood cells, mean corpuscular volume, mean corpuscular haemoglobin and mean corpuscular haemoglobin concentration, thrombocytes, total white blood cells prothrombin time and differential white blood cells.

BLOOD GAS ANALYSIS
- pH, pCo2, pO2. The amount of bicarbonate and base excess were then calculated

CLINICAL CHEMISTRY: Yes
- Time schedule for collection of blood: from abdominal aorta at necropsy
- Animals fasted: no data
- How many animals: 10/sex/dose
- Parameters checked: alkaline phosphatase, aspartate aminotransferase, alanine aminotransferase, gamma glutamyl transferase, lactate dehydrogenase, total protein, albumin, total bilirubin, urea, creatinine and calcium, inorganic phosphate, chloride, and sodium and potassium.

URINALYSIS: Yes
- Urinary acid indices were determined in weeks 1 and 4 in urine samples obtained from rats placed in metabolism cages with no food and water for 2– 3 h at the start of the light period. Urinary pH was measured in individual samples.
- The concentrating ability of the kidneys was examined by collecting urine from 10 rats/sex/group, during the last 16 h of a 24-h water deprivation period in week 4. During the 16-h collection period no feed was available. The samples were analysed for volume and density. In addition, examinations for protein, glucose, occult blood, ketones, bilirubin and urobilinogen appearance and microscopy of the sediment were carried out in the individual samples collected in week 4 (results of these examinations were not reported in the publication).

Sacrifice and pathology:

SACRIFICE
At the end of the 4-week, all rats were killed under light ether anaesthesia.

GROSS PATHOLOGY
- Femur composition was assessed at autopsy at the end of the 4 week in 10 rats/sex/group.
- Any abnormalities were recorded and the adrenals, brain, pituitary, kidneys, liver, spleen, testes, ovaries, thyroid, thymus and heart were weighed. Paired organs were weighed together.

HISTOPATHOLOGY
- Doses examined: all animals of each sex in the control group and the NH4Cl high-dose groups
- Organs examinedin control and high dose group: adrenals, kidneys, liver, lungs (inflated with the fixative), mesenteric lymph nodes, spleen, testes, urinary bladder (inflated with the fixative), and all gross lesions.
- Organs examined in the low dose group: Adrenals, heart, kidneys, stomach, mammary glands, thyroid, testes, urinary bladder and uterus.

Statistics:

Numerical data were evaluated for statistical significance using the following statistical tests: one-way analysis of (co)variance [AN(C)OVA] followed by Dunnett’s tests, or least significance difference (LSD) tests. Rates were evaluated by Mann–Whitney’s U test. Histopathological data were evaluated by two-sided Fisher exact probability test. A P value of less than 0.05 was considered to indicate statistical significance.

Results and discussion

Results of examinations

Details on results:

CLINICAL SIGNS AND MORTALITY
- There was the usual random incidence of ageing symptoms that occur in this strain of rats when maintained over a period of time, but there were no treatment-related abnormalities in condition or behaviour after 4 weeks
- Mortality rate was not affected by treatment of NH4Cl. None of the rats died during after 4 weeks

BODY WEIGHT AND WEIGHT GAIN
- 4% NH4Cl resulted in marked and significant (p < 0.01) growth depression at the end of the 4 weeks (male 25.5% and female 18.6% below control). 2.0 % NH4Cl also markedly decreased (p < 0.05) growth in females. Although statistical analysis indicated a statistical difference to control animals, this difference is less than 10%, videlicet 7.5% below control value

FOOD INTAKE:
- Food intake was decreased in the 4% NH4Cl group. Otherwise, there were no consistent differences in food intake among the groups (data not shown).

WATER INTAKE
- The feeding of 4% NH4Cl after 4 weeks resulted in increases in water intake up to 40–60%. With 2.0% NH4Cl, water intake was ca. 20% increased in males at all stages but not in females (data not shown)

BLOOD GAS ANALYSIS
After 4 weeks of feeding of NH4Cl, a dose related decrease in base excess (Male 4.2 and -2.2 vs. 7.2 mmol/l in control in the low dose and high dose, respectively; Female 1.5 and -0.2 vs. 7.6 mmol/l in controls in the low and high dose, respectively) associated with lower blood pH and bicarbonate concentration.

URINARY EXAMINATIONS
- measured in the firts week of feeding, 2.0 % NH4Cl caused a significant (p < 0.01) decrease in urinary pH in both sexes (5.6 vs 7.6 in controls for males and 5.7 vs 7.7 in controls in females). 4% NH4Cl reduced the urinary pH to a similar extent (data not shown).
- There was a dose dependent increase in the net acid excretion (data not shown).

HEMATOLOGY and CLINICAL CHEMISTRY
- There were no consistent or treatment-related effects on red blood cell variables, clotting potential or total and differential white blood cell counts in any of the groups.
- Plasma chloride levels were significantly increased after treatment with 2.0% NH4Cl after 4 weeks (4.0% over controls) in males only. With 4% NH4Cl the increases were in both sexes ranging from 8.7-11.8 % after 4 weeks.
- Alkaline phosphatase activity was increased with 4% NH4Cl after 4 weeks (22.6-50.0% over controls) in both sexes and in males fed 2.0% NH4Cl for 4 weeks (27.7% over controls). With 4% NH4Cl, decreases were noted in creatinine concentration and aspartate amino transferase activity, and increases in bilirubin and inorganic phosphate concentrations (no data shown in publication).

ORGAN WEIGHTS
- There were no consistent or treatment-related changes in the weights of the liver, spleen, ovaries, pituitary, thyroid, thymus or heart. The relative weight of the kidneys (relative to body weight) was increased in rats of both sexes fed 2.0 NH4Cl (11-13.6% above controls) or 4% NH4Cl (16-19%) above controls. The increase in kidney weights were due to hypertrophy which was not associated with any adverse histopathological findings. The relative weight of the adrenals was increased in males fed 4% NH4Cl (24.7% above controls). This is in line with zona glomerulosa hypertrophy seen at this dose.
- The analysis of the femur at the end of the studies did not reveal any effects of NH4Cl on femur weight, calcium content or on total bone substance measured as fat free solid.

GROSSPATHOLOGY
- Macroscopic examination at necropsy after 4 weeks did not reveal significant differences among the treatment groups and the controls.

HISTOPATHOLOGY
- Most histopathological changes observed in the various studies were about equally distributed among the treatment groups and the controls and represent normal background pathology for rats of this strain and age. A number of treatment-related non-neoplastic changes were, however observed. Dose-related increases in the incidence of zona glomerulosa hypertrophy (adrenal) occurred in both sexes. Statistical significance was not seen

Effect levels

Dose descriptor:

NOAEL

Remarks:

(28 days; systemic effects)

Effect level:

ca. 2 214.5 mg/kg bw/day (actual dose received)

Sex:

male/female

Basis for effect level:

other: Based on significant weight loss (18-25% below control value) in both sexes of animals treated with 4228.5 mg/kg bw

Target system / organ toxicity

Critical effects observed:

not specified

Any other information on results incl. tables

It may be concluded that most of the observed changes can be regarded as physiological adaptations to the feeding of acid forming salt, and that the rats showed a remarkable adaptive capacity of adaptation