Reaction mass of Magnesium peroxide and magnesium carbonate and magnesium hydroxide and magnesium oxide

Developmental toxicity / teratogenicity

Administrative data

Endpoint:

developmental toxicity

Type of information:

experimental study

Adequacy of study:

key study

Study period:

09-03-2010 to 14-05-2010

Reliability:

1 (reliable without restriction)

Rationale for reliability incl. deficiencies:

other: GLP study conduced according to guidelines.

Cross-reference

Reason / purpose for cross-reference:

reference to same study

Data source

Reference

Reference Type:

study report

Title:

Unnamed

Year:

2010

Materials and methods

Test guideline

Test guideline 1

Qualifier:

according to guideline

Guideline:

other: OECD Guideline 422 (Combined Repeated Dose Toxicity Study with the Reproduction / Developmental Toxicity Screening Test)

Deviations:

no

Test guideline 2

Qualifier:

according to guideline

Guideline:

other: EPA EPA OPPTS 870.3650 (Combined Repeated Dose Toxicity Study with the Reproduction/Developmental Toxicity Screening Test, July 2000)

Deviations:

no

Test guideline 3

Qualifier:

according to guideline

Guideline:

other: OECD Guideline 421 (Reproduction / Developmental Toxicity Screening Test)

Test guideline 4

Qualifier:

according to guideline

Guideline:

other: EPA OPPTS 870.3550( Reproduction/Developmental Toxicity Screening Test July 200)

Deviations:

no

GLP compliance:

yes

Test material

Test material information

Constituent 1

Reference substance name:

Magnesium hydroxide

EC Number:

215-170-3

EC Name:

Magnesium hydroxide

Cas Number:

1309-42-8

Molecular formula:

H2MgO2

IUPAC Name:

magnesium dihydroxide

Details on test material:

-Molecular formula: Mg (OH) 2
-Molecular weight: 58.32
-Stability in water: Stable
-Description: White powder
-Test substance storage: At room temperature in the dark
- Stability under storage conditions: Stable
- Hygroscopic: yes store in a closed vessel
-pH: ±10 (10 %H2O)
-Solubility in water: 0.00014 mol/L

Test animals

Species:

rat

Strain:

Wistar

Details on test animals or test system and environmental conditions:

-Source: Charles River laboratories France, L’Arbresle Cedex, France.
-Age at study initiation: Approximately 11 weeks
-Housing:
Lactation: Pups were kept with the dam until termination in Macrolon cages (MIII type, height 18cm)
General: Sterilised sawdust as bedding material and paper as cage enrichment. During activity monitoring animals were housed individually in Macrolon cages with sterilised sawdust as bedding material. No cage-enrichment was provided during activity monitoring.
Diet: Free access to pelleted rodent diet.
Water: Free access to tap water
-Acclimation period: At least 5 days prior to the start of treatment

Environmental Conditions:
Animals were housed in a controlled environment.
Temperature (°C): 21±3°C (actual range 19.7- 21.7°C)
-Humidity (%): A relative humidity of 40-70% (actual range: 34-73%)
- Air changes per hour: 15 air changes per hour
-Photoperiod (Hrs dark/hrs light): 12 hours artificial light and 12 hours darkness per day.

Administration / exposure

Route of administration:

oral: gavage

Vehicle:

water

Analytical verification of doses or concentrations:

yes

Details on analytical verification of doses or concentrations:

Accuracy and homogeneity were determined for formulations prepared for use during treatment. Duplicate samples( approx 500 mg), which were taken from the formulations using a pipette, were accurately weighed into volumetric flasks of 50ml. For determination of accuracy, samples were taken at 50% height or at 90%, 50% and 10% height. The latter set of samples was also used for the determination of the homogeneity of the samples. The volumetric flasks were filled up to the mark with 4% aqueous HNO3. The solutions were further diluted with 4% aqueous HNO3 to obtain concentrations within the calibration range.

Details on mating procedure:

- M/F ratio per cage: Following a minimum of 14 days of exposure for the males and females, one female was cohabitated with one male of the same treatment group, avoiding sibling mating
- Length of cohabitation: Once mating had occurred, the males and females were separated
- Proof of pregnancy: Detection of mating was confirmed by evidence of sperm in the vaginal lavage, by staging of the estrous cycle and/or by the appearance of an intravaginal copulatory plug. This day was designated Day 0 post-coitum. Once mating had occurred, the males and females were separated.
- After successful mating each pregnant female was caged: Females were individually housed in Macrolon cages (MIII type, height 18cm). the females were allowed to litter normally. Day 1 of lactation was defined as the day when a litter was found complete (i.e. membranes,placentas cleaned up, nest built up and/or feeding of pups started). Females that were littering were left undisturbed.

Duration of treatment / exposure:

Pups were not treated directly, but were potentially exposed to the test substance in utero and through lactational transfer.
Males were exposed for 29 days, i.e. 2 weeks prior to mating, during mating, and up to termination. Females were exposed for 41-45 days, i.e. during 2 weeks prior to mating, during mating, during post-coitum, and during at least 4 days of lactation. Females 41, 46, 48, 49 (group 1, table 1), 53, 59 (group 2, table 1), 61, 62, 68 (group3, table 1) and 76 (group 4, table 1) were not dosed during littering.

Frequency of treatment:

Once daily for 7 days per week, approximately the same time each day with a maximum of 6 hours difference between the earliest and latest dose. Animals were dosed up to the day prior to scheduled necropsy.

No. of animals per sex per dose:

Four groups of ten male and ten female Wistar (Hans) rats were exposed by oral gavage to the test substance.

Control animals:

yes, concurrent no treatment

Details on study design:

-Dose selection rationale: Based on the results of a 10-day dose range finding study.
-Rationale for animal selection: This species and strain of rat has been recognized as appropriate for general and reproduction toxicity studies.

Examinations

Maternal examinations:

Detailed clinical Observations;
Time schedule: Daily detailed clinical observations were made in all animals immediately after dosing. Once, prior to the start of treatment and at weekly intervals this was also performed outside the home cage in a standard arena. Arena observations were not performed when the animals were mating, or housed individually.

Functional Observations:
The following tests were performed on the selected 5 animals/sex/group:
-hearing ability, pupillary reflex, static righting reflex and grip strength.
-motor activity test.
During the motor activity test, males were caged individually and females were caged with their pups. The selected males were tested during week 4 of treatment and the selected females were tested during lactation. In order to avoid hypothermia of pups, dams were removed from the pups for not more than 30-40 minutes.

Ovaries and uterine content:

The ovaries and uterine content was examined after termination: Yes
Examinations included:
- Number of corpora lutea: Yes (Table 1)
- Number of implantations: Yes (Table 1)
All animals were subjected to macroscopic examination, with special attention being paid to reproductive organs. Descriptions of all macroscopic abnormalities were recorded. The numbers of former implantation sites and corpora lutea were recorded for all paired females.

Fetal examinations:

-External examinations: Yes:
Each litter was examined to determine the following if possible:
Mortality/ Viability: The numbers of live and dead pups on Day 1 of lactation and daily thereafter were determined. If possible, defects or cause of death were evaluated.
Clinical signs: At least once daily, detailed clinical observations were made in all animals.
Body weights: Live pups were weighed on Days 1 and 4 of lactation.
Sex: Sex was determined for all pups on Days 1 and 4 of lactation.

Statistics:

For each group the following calculations were performed:
Mating (%) - (Number of females mated/Number of females paired) x 100
Fertility Index (%) – (Number of pregnant females/Number of females paired) x 100
Conception index (%) – (Number of pregnant females/Number of females mated) x 100
Gestation index (%) – (Number of females bearing live pups/ Number of pregnant females) x 100
Duration of gestation – Number of days between confirmation of mating and the beginning of parturition.
Percentage live males at first litter check – (Number of live male pups at first litter check/ Number of live pups at First Litter Check) x 100
Percentage live females at First Litter Check – (Number of live female pups at First Litter Check/ Number of live pups at First Litter Check) x 100
Percentage of postnatal loss Days 0-4 of lactation – (Number of dead pups on Day 4 of lactation/ Number of live pups at First Litter Check) x 100
Viability index (%) – Number of live pups on Day 4 of lactation/ Number of pups born alive) x 100

Results and discussion

Results: maternal animals

Effect levels (maternal animals)

Dose descriptor:

NOAEL

Effect level:

>= 1 000 mg/kg bw/day

Based on:

test mat.

Basis for effect level:

other: developmental toxicity

Results (fetuses)

Effect levels (fetuses)

Dose descriptor:

NOAEL

Effect level:

> 1 000 mg/kg bw/day

Based on:

test mat.

Basis for effect level:

other: developmental toxicity

Fetal abnormalities

Abnormalities:

not specified

Overall developmental toxicity

Developmental effects observed:

not specified

Treatment related:

not specified

Any other information on results incl. tables

No toxicologically relevant effects on gestation index and duration, parturition, maternal care and early postnatal pup development (mortality, clinical signs, body weight and macropsy) were observed.

Gestation:

The gestation index was 100% for all groups and the duration of gestation was similar between control and treated groups.

Parturition/ Maternal care: No signs of difficult or prolonged parturition were noted among the pregnant females. Examination of cage debris of pregnant females revealed no signs of abortion or premature birth. No deficiencies in maternal care were observed.

Early postnatal pup development: The number of dead and living pups at first litter check, postnatal loss, viability index and sex ratio were unaffected by treatment, and clinical signs, body weight and external macroscopy did not reveal treatment-related findings.

Mortality: Two pups of the control group, three pups at 110 mg/kg, and one pup at 330 mg/kg were found dead or missing during lactation. The missing pups were most likely cannibalized. No pups were found dead or missing at 1000 mg/kg. No toxicological relevance was attributed to these dead/missing pups since the mortality incidence did not show a dose related trend and remained within the range considered normal for puppies of this age.

Clinical signs: Incidental clinical symptoms of pups consisted of blue spot on the back and scabbing of the snout or back. The nature and incidence of these clinical signs remained within the range considered normal for pups of this age, and were therefore considered to be of no toxicological relevance.

Body weights: body weights of pups were considered to have been unaffected by treatment.

 

Table 1:

Corpora Lutea and Implantation sites:

Females F0-Generation

 

		Group 1 Control	Group 2 110 mg/kg	Group 3 330 mg/kg	Group 4 1000 mg/kg
Corpora lutea	Mean	14.0	15.1	14.7	14.5
	St.Dev	2.2	2.7	4.4	2.7
	N	10	10	10	10
Implantation sites	Mean	12.6	13.1	12.6	13.2
	St.Dev	1.2	1.6	1.2	1.4
	N	10	10	10	10

Table 2:

Developmental data:

F0- generation- Lactation

 

	Group 1 Control	Group 2 110 mg/kg	Group 3 330 mg/kg	Group 4 1000 mg/kg
Litters
Total	10	10	10	10
Duration of gestation
Mean (+)	21.3	21.3	21.3	21.2
St. Dev	0.5	0.5	0.5	0.4
N	10	10	10	10
Dead pups at first litter check
Litters affected (#)	0	1	1	0
Total	0	1	1	0
Mean	0.0	0.1	0.1	0.0
St.Dev	0.0	0.3	0.3	0.0
N	10	10	10	10
Living pups at first litter check
% of Males/Females	45/55	51/49	55/45	54/46
Total	116	121	118	123
Mean	11.6	12.1	11.8	12.3
St.Dev	1.1	1.4	1.5	1.8
N	10	10	10	10
Postnatal loss
% of Living Pups	1.7	1.7	0.0	0.0
Litters affected	2	2	0	0
Total	2	2	0	0
Mean	0.2	0.2	0.0	0.0
St.Dev	0.4	0.4	0.0	0.0
N	10	10	10	10
Viability Index	98.3	98.3	100.0	100.0

 

Applicant's summary and conclusion

Conclusions:

No reproduction/developmental toxicity were observed at any dose level. Based on these results, a parental, reproduction and developmental No Observed Adverse Effect Level (NOAEL) of at least 1000 mg/kg/day was determined.

Executive summary:

Overall, no toxicologically relevant changes were noted in any of the parental parameters investigated in this study.