Magnesium

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:

supporting study

Reliability:

2 (reliable with restrictions)

Rationale for reliability incl. deficiencies:

other: Study is in principle comparable to the OECD guidelines of testing for chemicals. Materials, methods and results are well described and documented. No information on analytic available.

Data source

Reference

Reference Type:

publication

Title:

A 90-day repeated dose oral toxicity study of magnesium chloride in F344 rats

Author:

Takizawa, T. et al.

Year:

2000

Bibliographic source:

Bull. Natl. Inst. Health Sci. 118: 63-70.

Materials and methods

Test guideline

Test guideline 1

Qualifier:

equivalent or similar to guideline

Guideline:

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

Deviations:

no

Test guideline 2

Qualifier:

equivalent or similar to guideline

Guideline:

EU Method B.26 (Sub-Chronic Oral Toxicity Test: Repeated Dose 90-Day Oral Toxicity Study in Rodents)

Deviations:

no

Principles of method if other than guideline:

In order to examine the toxicity of magnesium chloride hexahydrate, four groups of 10 male and 10 female rats received the compound by dietary supplementation at 2.5, 0.5, 0.1 or 0 % for 90 days.

GLP compliance:

not specified

Limit test:

no

Test material

Test material information

Constituent 1

Reference substance name:

Magnesium chloride

EC Number:

232-094-6

EC Name:

Magnesium chloride

Cas Number:

7786-30-3

IUPAC Name:

magnesium dichloride

Constituent 2

Reference substance name:

7791-18-6

Cas Number:

7791-18-6

IUPAC Name:

7791-18-6

Constituent 3

Reference substance name:

magnesium chloride hexahydrate

IUPAC Name:

magnesium chloride hexahydrate

Test material form:

other: solid

Details on test material:

- Name of test material (as cited in study report): magnesium chloride hexahydrate
- Substance type: technical product
- Physical state: solid

No further information given.

Test animals

Species:

rat

Strain:

Fischer 344

Sex:

male/female

Details on test animals or test system and environmental conditions:

TEST ANIMALS: F344/DuCrj rats (SPF)
- Source: Nippon Charles River
- Age at study initiation: 5 week old (at purchase); 6 week old (after acclimatisation)
- Housing: in groups of 3 or 4 individuals in plastic cages with wood chip bedding (Soft Chip; Sankyo Lab Service Corp.) in a breeding room with a barrier system; the cages were exchanged properly
- Diet (ad libitum): base diet CRF-1 from Oriental Yeast Co., Ltd
- Water (ad libitum): tap water
- Acclimation period: 8 days

ENVIRONMENTAL CONDITIONS
- Temperature (°C): 23 ± 2 °C
- Humidity (%): 55 ± 5%
- Air changes (per hr): 18 times/hr
- Photoperiod (hrs dark / hrs light): 12-hr light and darkness cycling (artificial lightning 07:00 ~ 19:00)

No further information given.

Administration / exposure

Route of administration:

oral: feed

Vehicle:

unchanged (no vehicle)

Details on oral exposure:

DIET PREPARATION
- A solid compound diet was used that was obtained by pelletizing a mixture of magnesium chloride hexahydrate provided by Ako Kasei Co., Ltd. and base diet CRF-1 from Oriental Yeast Co., Ltd.

- The stability of this compound diet was proven in a storage test comprising 3 months closed storing under cooling at a dark place followed by 2 weeks open storing under diffuse light at ambient temperature.
- The diet was stored in a cooled feed storage until use and administered to the animals within the period for which diet stability was guaranteed.

No further information given.

Analytical verification of doses or concentrations:

not specified

Details on analytical verification of doses or concentrations:

no data

Duration of treatment / exposure:

90 days

Frequency of treatment:

in diet ad libitum (continuous)

Doses / concentrations

Doses / concentrations 1

Remarks:

Doses / Concentrations:
2.5% test substance
Basis:
nominal in diet

Doses / concentrations 2

Remarks:

Doses / Concentrations:
0.5% test substance
Basis:
nominal in diet

Doses / concentrations 3

Remarks:

Doses / Concentrations:
0.1% test substance
Basis:
nominal in diet

No. of animals per sex per dose:

10 rats/sex/group (4 groups)

Control animals:

yes, plain diet

Details on study design:

- Dose selection rationale:
The test substance concentration in the compound diet was determined from the results of a 2-week pretest conducted before the present study. In this pretest with a test substance concentration of 5% (i.e. the concentration which is usually considered as upper limit for the maximum test substance concentration in food additive tests conducted with compound diets) and 2.5% (the half of aforementioned concentration), no decrease in diet consumption was observed in both the 5% and 2.5% group. However, diarrhea, increase of water consumption and a tendency of inhibited body weight increase occurred in both groups. Furthermore, autopsy revealed softening of caecum and colon content. From these results, the maximum concentration applied in the present study was set at the concentration (i.e. 2.5%) at which the administration of the test substance is likely to show effects, and for the concentration applied in the further administration groups this concentration was divided by a common ratio of 5 to achiev 0.5 and 0.1% dietary levels.

- Rationale for animal assignment (if not random):
The animals were allocated to four groups of 10 rats per sex in such a way that the body weight (based on the newest body weight at the day before the begin of the experiment) of each animal was close to the average body weight of the respective group.

No further information given.

Positive control:

no data

Examinations

Observations and examinations performed and frequency:

CAGE SIDE OBSERVATIONS: Yes
- Time schedule: every day
- Cage side observations: general condition of the animals (including mortality)

DETAILED CLINICAL OBSERVATIONS: No data

BODY WEIGHT: Yes
- Time schedule for examinations: weekly

FOOD CONSUMPTION AND COMPOUND INTAKE (if feeding study): Yes
- Food consumption for each animal determined and mean daily diet consumption calculated as g food/kg body weight/day: weekly
- Compound intake calculated as time-weighted averages from the consumption and body weight gain data: No data

FOOD EFFICIENCY: No data

WATER CONSUMPTION AND COMPOUND INTAKE (if drinking water study): Yes
- Time schedule for examinations: weekly

OPHTHALMOSCOPIC EXAMINATION: No

HAEMATOLOGY: Yes, blood was collected from the dorsal aorta
- Time schedule for collection of blood: on completion of the administration period
- Anaesthetic used for blood collection: with ether
- Animals fasted: overnight
- How many animals: all surviving animals
- Parameters checked: read blood cell count (RBC), hemoglobin (Hb), hematocrit value (Ht), mean corpuscular volume (MCV), mean corpuscular hemoglobin (MCH), mean corpuscular hemoglobin concentration (MCHC), white blood cell count (WBC) and platelet count (Plt) were measured using a multi-automatic blood cell counter (M-2000, Toa Electronic Co.)
- In addition, differential counts using an automatic blood cell analyzer (MICROX HEG-120A, Tateishi Electric Co.) were conducted: band neutrophilocytes (Band), segmented neutrophilocytes (Seg), eosinocytes (Eosino), basocytes (Baso), lymphocytes (Lympho), monocytes (Mono) and nucleated erythrocytes (Ebl)

CLINICAL CHEMISTRY: Yes, blood was collected from the dorsal aorta
- Time schedule for collection of blood: on completion of the administration period
- Anaesthetic used for blood collection: with ether
- Animals fasted: overnight
- How many animals: all surviving animals
- Parameters checked: total protein (TP), albumin (Alb), albumin/globulin (A/G), total cholesterol (T-Cho), γ-glutamyl transpeptidase (γGTP), cholinesterase (ChE), alanine transaminase (AlT), aspartic acid transaminase (AsT), lactic acid dehydrogenase (LDH), alkaline phosphatase (ALP), blood urea nitrogen (BUN), creatinine (Cre), Calcium (Ca), inorganic phosphorus (IP), sodium (Na), potassium (K) and chloride (Cl) were measured by SRL Co. (Tokyo)

URINALYSIS: No

NEUROBEHAVIOURAL EXAMINATION: No

No further information given.

Sacrifice and pathology:

After blood-taking the animals were killed by exsanguination and subjected to autopsy.

GROSS PATHOLOGY: Yes
- Brains, thymus glands, lungs, spleens, livers, adrenal glands, suprarenal glands, kidneys, and testis were weighed.

HISTOPATHOLOGY: Yes
- Animals of the control group and the maximum dose group were examined.
- The organs were fixed with 10% neutral-buffered formalin, or as for testis (from 5 animals of the control group and 5 animals from the maximum dose group) in Bouin's fixature, were cut into thin slices according to conventional methods and stained with hematoxylin-eosin stain.
- Organs examined: lymph nodes (neck section and mesenteries), thymus glands, bones and bone marrow (sternal bones and thighbones), thymus glands, tracheas, lungs, hearts, thyroids and parathyroids, tongues, gullets, proventriculuses, glandular stomachs, duodenums, small intestines (jejunum and ileum), colons (cecum, intestinum colon, rectum), livers, pancreases, urinary bladders, kidneys, suprarenal glands, vesicular glands, prostate gland, epididymises, ovaries and oviducts, uteruses, vaginas, brains, hypophysises, sciatic nerves, skeletal muscles, spinal cords, eyeballs and adnexal organs.

No further information given.

Other examinations:

no data

Statistics:

As for the body weight measured over the whole administration period, and the hematological and histopathological findings, an intergroup comparison by a one-way analysis of variance was conducted, and those items which showed significance at a significance level of 5% were compared to the control group by a two-sided test according to Dunnett with significance levels of 5% and 2%.

Results and discussion

Results of examinations

Clinical signs:

effects observed, treatment-related

Mortality:

mortality observed, treatment-related

Body weight and weight changes:

effects observed, treatment-related

Food consumption and compound intake (if feeding study):

no effects observed

Food efficiency:

not specified

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

no effects observed

Ophthalmological findings:

not examined

Haematological findings:

no effects observed

Clinical biochemistry findings:

no effects observed

Urinalysis findings:

not examined

Behaviour (functional findings):

not examined

Organ weight findings including organ / body weight ratios:

no effects observed

Gross pathological findings:

not specified

Histopathological findings: non-neoplastic:

no effects observed

Histopathological findings: neoplastic:

not specified

Details on results:

CLINICAL SIGNS AND MORTALITY
- No treatment related death was observed during the study.
- Transient soft stool was observed in males and females of the 2.5 % group (from week 2 of administration).

BODY WEIGHT AND WEIGHT GAIN
- Slight reduction in body weight gain was noted in the 2.5 % group (males) from week 5 of administration onwards, and the difference to the control group became significant in the last week of administration.

FOOD CONSUMPTION AND COMPOUND INTAKE (if feeding study)
- No effects on food consumption were observed.
- Compound intake was 62, 308 or 1600 mg/kg bw/d in males and 59, 299 or 1531 mg/kg in females of the 0.1, 0.5 or 2.5% groups, respectively.

WATER CONSUMPTION AND COMPOUND INTAKE (if drinking water study)
- Water consumption was increased in males and females of the 2.5 % group onwards.
- This was considered as a change of rather low toxicologic relevance.

HAEMATOLOGY
- No toxically relevant changes.
- Some effects with respect to erythrocyte-related values (decreased RBC, Hb and Ht in males; incresaed Hb and MCH in females) observed in the present study were considered of low toxicological significance.

CLINICAL CHEMISTRY
- No toxically relevant changes.
- Some effects (decreased total cholesterol, cholinesterase, lactic dehydrogenase and alkaline phosphatase, and an increase of inorganic phosphorus), mainly observed in males, were mild and not accompanied by histopathological changes, and thus considered as toxicologically unproblematic.

ORGAN WEIGHTS
- No treatment-ralted changes were observed.
- Single incidences of decreased organ weights were correlated to body weight changes and not considered of toxicological significance.

HISTOPATHOLOGY: NON-NEOPLASTIC
- No differences were observed between the control and treated groups.

No further information given.

Effect levels

1

Dose descriptor:

NOAEL

Effect level:

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

Based on:

test mat.

Sex:

male

Basis for effect level:

other: Transient soft stool was observed in the 2.5% group and, in addition, a suppression of body weight gain was found in males.

2

Dose descriptor:

NOAEL

Effect level:

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

Based on:

test mat.

Sex:

female

Basis for effect level:

other: Transient soft stool was observed in females of the 2.5% group.

Target system / organ toxicity

Critical effects observed:

not specified

Any other information on results incl. tables

A translation of the japanese study is attached on this robust study summary. Table with the dose levels, food consumption and the intakes of MgCl2 are presented there.

Applicant's summary and conclusion

Conclusions:

Magnesium chloride administered to rats as an admixture to solid food over a period of 90 days caused transient soft stool in male and female animals of the 2.5% group and suppression of body weight gain in males. Thus, the dose of 0.5% (i.e. 308 mg/kg bw/d for males and 299 mg/kg bw/d for females) was considered as the no-observed-adverse-effect-level (NOAEL).