Bis(D-gluconato-O1,O2)zinc

Repeated dose toxicity: oral

Administrative data

Endpoint:

sub-chronic toxicity: oral

Type of information:

experimental study

Adequacy of study:

key study

Study period:

1981

Reliability:

2 (reliable with restrictions)

Rationale for reliability incl. deficiencies:

comparable to guideline study with acceptable restrictions

Data source

Reference

Reference Type:

publication

Title:

Unnamed

Year:

1981

Report date:

1981

Materials and methods

Test guideline

Qualifier:

equivalent or similar to guideline

Guideline:

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

GLP compliance:

not specified

Limit test:

no

Test material

Test material information

1

Reference substance name:

Zinc sulphate

EC Number:

231-793-3

EC Name:

Zinc sulphate

Cas Number:

7733-02-0

Molecular formula:

H2O4S.Zn

IUPAC Name:

Zinc sulphate

Test material form:

solid: particulate/powder

Specific details on test material used for the study:

- Name of test material: Zinc sulphate heptahydrate
- Substance type: Pure active substance
- Physical state: Crystal
- Analytical purity: 99.9 %

Test animals

Species:

mouse

Strain:

ICR

Sex:

male/female

Details on test animals or test system and environmental conditions:

TEST ANIMALS
- Source: Shizuoka Agriculture Cooperative Association for Laboratory Animals
- Age at study initiation: Four weeks
- Weight at study initiation: 25-30 g (male); 20-25 g (female)
- Fasting period before study: No data
- Housing: Four animals housed in each stainless cage with a wire-meshed bottom
- Diet: Pulverized chows F (Oriental Yeast Co.), ad libitum, mixed with test material
- Water: Ad libitum
- Acclimation period: One wk


ENVIRONMENTAL CONDITIONS
- Temperature: 24±1 °C
- Humidity: 55±5 %
- Air changes (per hr): No data
- Photoperiod: 10 h dark/14 h light cycle

Administration / exposure

Route of administration:

oral: feed

Vehicle:

other: Mixed with basic feed

Details on oral exposure:

DIET PREPARATION:
- Mixing appropriate amounts with (Type of food): Pulverized chows M (produced by Oriental Yeast Co.

Analytical verification of doses or concentrations:

no

Duration of treatment / exposure:

13 weeks

Frequency of treatment:

daily

Doses / concentrations

Doses / concentrations 1

Dose / conc.:

300 ppm

Doses / concentrations 2

Dose / conc.:

3 000 ppm

Doses / concentrations 3

Dose / conc.:

30 000 ppm

No. of animals per sex per dose:

12

Control animals:

yes, plain diet

Details on study design:

- Dose selection rationale: not specified
- Rationale for animal assignment: random
The animals were divided into four groups, each of which included 12 animals of each sex and fed on diets containing Zinc sulphate at four different concentration levels 0, 300, 3,000 and 30,000 ppm, for 13 weeks.

Positive control:

No

Examinations

Observations and examinations performed and frequency:

DETAILED CLINICAL OBSERVATIONS: Yes
- Time schedule: Daily

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: Yes
- Compound intake calculated as time-weighted averages from the consumption and body weight gain data: Yes

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: Yes

WATER CONSUMPTION: Yes
- Time schedule for examinations: Twice a week

HAEMATOLOGY: Yes
- Time schedule for collection of blood: Not specified
- Anaesthetic used for blood collection: Yes (under light ether anaesthesia)
- How many animals: 96
- Parameters checked: Erythrocyte count, hemoglobin, leukocyte count, differential count of leukocyte

CLINICAL CHEMISTRY: Yes
- Time schedule for collection of blood: Not specified
- How many animals: 96
- Parameters checked: Total plasma protein, alkaline phosphatase, glucose, urea nitrogen, SGOT, SGPT, cholesterol and calcium.

Sacrifice and pathology:

GROSS PATHOLOGY: Yes
After necropsy at the termination of the study the following organs were weighed: Brain, pituitary, thyroid, heart, thymus, liver, kidney, spleen, adrenals, gonads (testes or ovaries), and muscles (triceps surae).

HISTOPATHOLOGY: Yes (see table)
- For histopathology following organs and tissues were collected: Brain, pituitary, thyroid, heart, thymus, liver, kidney, spleen, adrenals, gonads (testes or ovaries), and muscles (triceps surae), submaxillary glands, lungs, mesenteric lymph nodes, pancreas, stomach, small and large intestine, accessory genital organs, bone and bone marrow (sternum and femur), and lesions of gross abnormalities
- 3 or 4 µm paraffin sections from the specimens were stained with hematoxylineosin, periodic acid Schiff's reaction and azan for microscopic observations.

Other examinations:

None

Statistics:

Student's t-test was used to estimate the statistical differences between controls and treated groups.

Results and discussion

Results of examinations

Clinical signs:

effects observed, treatment-related

Description (incidence and severity):

- At 30,000 ppm: Depressed spontaneous motility before death or sacrifice.
- At ≤ 3,000 ppm: No treatment related toxic signs

Mortality:

mortality observed, non-treatment-related

Description (incidence):

- At 30,000 ppm: Four males and one female in this group were found dead or killed in extremis during the study. Histological findings of these animals revealed impairment of the urinary tract and regressive changes in the exocrine gland of the pancreas. Mortality was 33.3% in males and 8.3% in females.

Body weight and weight changes:

effects observed, treatment-related

Description (incidence and severity):

- At 30,000 ppm: A more prominently retarded growth resulting in smaller body size than those of other groups.
- At 300 ppm: A significant but very slight depression of weight gain was seen in females for a week after commencement, followed by a rapid recovery to the control level.

Food consumption and compound intake (if feeding study):

effects observed, treatment-related

Description (incidence and severity):

- At 30,000 ppm: The food intake of male and female mice was depressed during the first week of the study in comparison to that of the controls but showed a tendency to recover afterwards. Average food intake of these animals then remained at only a slightly lower level than that of the control group.

Food efficiency:

effects observed, treatment-related

Description (incidence and severity):

Markedly lower than those of the control group during the first week of the study, corresponding to the decrease in food intake.
-At 30,000 ppm: The overall average food efficiency was much lower than the control group.
- At 3,000 ppm: No data

Water consumption and compound intake (if drinking water study):

effects observed, treatment-related

Description (incidence and severity):

-At 30,000 ppm: Decreased in both sexes during the first week. Though males soon recovered, females showed persistent lower intake during the study.

Ophthalmological findings:

not examined

Haematological findings:

effects observed, treatment-related

Description (incidence and severity):

Male and female mice in the 30,000 ppm group showed moderately lower values in hematocrit and hemoglobin concentration than those of the control group; the leukocyte count in males also decreased moderately. Morphological changes of erythrocyte anisocytosis, polychromatophilia and poikilocytosis, were seen in six males and four females which were reported by necropsy or microscopic observation to have fore-stomach ulcers. Though some significant fluctuations were seen in hematocrit, no dose dependent abnormalities were found in hemoglobin concentration and erythrocyte count in males or females at less than 3,000 ppm group.

Clinical biochemistry findings:

effects observed, treatment-related

Description (incidence and severity):

-At 30,000 ppm: A slight to moderate decrease in total protein, glucose and cholesterol and a moderate to marked increase in alkaline phosphatase and urea nitrogen. Additional significant changes occurring in these animals were depression of SGPT level and increase in calcium level in females, and an increase of SGOT level in males
- At 3,000 ppm: Some fluctuations with significant differences from controls were seen in several parameters but these were all within the acceptable historical limits of mice of this strain and age.

Urinalysis findings:

not examined

Behaviour (functional findings):

not examined

Immunological findings:

not examined

Organ weight findings including organ / body weight ratios:

effects observed, treatment-related

Description (incidence and severity):

-At 30,000 ppm: Coincidental increase in absolute and relative weight was found in the thyroids of males and the kidneys of females.
- At ≤ 3,000 ppm: Statistically not significant values when compared to control.

Gross pathological findings:

effects observed, treatment-related

Description (incidence and severity):

-At 30,000 ppm: Marked emaciation, ischemic discoloration of the kidney and thyroid, atrophy of the pancreas, edematous thickening of the upper small intestine and slight splenomegaly were recorded in addition to several cases of fore stomach ulcer.

Neuropathological findings:

not examined

Histopathological findings: non-neoplastic:

effects observed, treatment-related

Description (incidence and severity):

-At 30,000 ppm: There were lesions attributable to the treatment in the pancreas, upper intestine, stomach, spleen and kidney. The pancreatic acinar cells had swollen nuclei with an increased number of clarified nucleoli and whirl-like profiles in their cytoplasm which were more basophilically stained than the controls. Single cell necrosis of the acinar cells was also a common feature in these animals. Moreover, a decrease in the number of acinus and ductule like metaplasia of acinar cells was demonstrated. There was mucosal catarrh in the upper intestine with proliferation of epithelial cells and edema at lamina propria, slight to moderate ulcerative lesions in the boundary of the fore-stomach and proliferation of erythropoietic immature cells in the splenic red pulp of these animals. Regressive changes of the renal cortex were observed in the females.
- At ≤ 3,000 ppm: Statistically not significant values when compared to control

Histopathological findings: neoplastic:

no effects observed

Effect levels

1

Dose descriptor:

NOEL

Effect level:

3 000 ppm

Based on:

test mat. (total fraction)

Sex:

male/female

Remarks on result:

other: approximately equivalent to 458 mg/kg/day on male, 479 mg/kg/day on female.

2

Dose descriptor:

LOEL

Effect level:

30 000 ppm

Based on:

test mat. (total fraction)

Sex:

male/female

Basis for effect level:

body weight and weight gain

clinical biochemistry

clinical signs

food consumption and compound intake

food efficiency

gross pathology

haematology

histopathology: non-neoplastic

organ weights and organ / body weight ratios

water consumption and compound intake

Target system / organ toxicity

1

Critical effects observed:

yes

Lowest effective dose / conc.:

30 000 ppm

System:

immune system

Organ:

leucocyte development

Treatment related:

yes

2

Critical effects observed:

yes

Lowest effective dose / conc.:

30 000 ppm

System:

gastrointestinal tract

Organ:

intestine

kidney

pancreas

spleen

stomach

Treatment related:

yes

3

Critical effects observed:

yes

Lowest effective dose / conc.:

30 000 ppm

System:

haematopoietic

Organ:

erythrocyte development

Treatment related:

yes

Applicant's summary and conclusion

Conclusions:

Under the test conditions, the NOEL of the test material in mice was determined to be 3,000 ppm (approximately equivalent to 458 mg/kg/day in male mice or 479 mg/kg/day in female mice).

Executive summary:

A study was conducted to evaluate the sub chronic (13 wk) toxicity of the test material in ICR mice. The study followed was equivalent or similar to OECD Guideline 408 (Repeated Dose 90-Day Oral Toxicity in Rodents).

Mice of both sexes were fed a diet containing test substance at 0, 300, 3000 and 30000 ppm for 13 wk. The clinical signs of the animals, body weight, food, chemical and water intake, food efficiency, hematological, biochemical examination, necropsy and organ weight and histopathological examination were performed.

Animals in the 30,000 ppm group showed retarded growth along with low food intake, abnormal values in a few hematological parameters, decreased water intake and significant deviations in biochemical parameters. Histopathological lesions included catarrh at the upper intestine, ulcers at the boundary of fore- and glandular stomach, proliferation of erythropoietic immature cells in the splenic red pulp as well as pancreatic lesions.

Under the test conditions, NOEL of the test material in mice was determined to be 3,000 ppm (approximately equivalent to 458 mg/kg/day in male mice or 479 mg/kg/day in female mice.