Calcium carbonate

Endpoint summary

Key value for chemical safety assessment

Effects on fertility

Description of key information

A number of supporting studies are reported in the literature where calcium has been administered to pregnant women. No teratogenic or reproductive effects were noted. However, the excessive consumption of antacid tablets during pregnancy can lead to neonatal hypercalcaemia. These effects were fully reversed following birth and hospital treatment.

Link to relevant study records

Reference

Endpoint:

screening for reproductive / developmental toxicity

Type of information:

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

Adequacy of study:

key study

Justification for type of information:

1. HYPOTHESIS FOR THE ANALOGUE APPROACH
Bulk calcium carbonate and uncoated nano calcium carbonate are chemically identical. The only difference between them is the particle size, which may result in minor differences in physico-chemical properties. However, in biological systems and in the environment, both bulk and uncoated nano calcium carbonate are expected to have identical (eco)toxicological and environmental fate properties.
2. SOURCE AND TARGET CHEMICAL(S) (INCLUDING INFORMATION ON PURITY AND IMPURITIES)
Uncoated nano calcium carbonate (Source): Refer to IUCLID section 1.2 for information on purity and impurities.
Bulk calcium carbonate (Target): Refer to IUCLID section 1.2 for information on purity and impurities.
The majority of new studies have been performed using uncoated nano calcium carbonate as the test material, hence it has been designated as the ‘Source’ in the read-across justification. However, for some endpoints data on bulk calcium carbonate are available and used to read-across to uncoated nano calcium carbonate so for these endpoints, marked with a * in the read-across justification document, bulk calcium carbonate is the ‘Source’.
3. ANALOGUE APPROACH JUSTIFICATION
Please refer to the justification attached in Section 13 of the dataset
4. DATA MATRIX
Please refer to the justification attached in Section 13 of the dataset

Reason / purpose for cross-reference:

read-across: supporting information

Reason / purpose for cross-reference:

read-across source

Key result

Dose descriptor:

NOEL

Effect level:

1 000 mg/kg bw/day (actual dose received)

Based on:

test mat.

Sex:

male/female

Basis for effect level:

other: No toxicologically significant effects observed at any dose level.

Sexual maturation:

not examined

Key result

Dose descriptor:

NOAEL

Generation:

F1

Effect level:

>= 1 000 mg/kg bw/day (actual dose received)

Based on:

test mat.

Sex:

male/female

Basis for effect level:

other: No statistically significant treatment-related effects at the high dose

Reproductive effects observed:

not specified

Effect on fertility: via oral route

Endpoint conclusion:

no adverse effect observed

Study duration:

subacute

Species:

rat

Effect on fertility: via inhalation route

Endpoint conclusion:

no study available

Effect on fertility: via dermal route

Endpoint conclusion:

no study available

Additional information

A repeat dose oral toxicity study combined with a reproduction/ developmental toxicity screening test was performed in the rat in accordance with OECD TG 422 (Dunster, 2010) and GLP. Uncoated nano calcium carbonate was administered by gavage to three groups, each of ten male and ten female Wistar rats, for up to forty-eight consecutive days (including a two week maturation phase, pairing, gestation and early lactation for females), at dose levels of 0, 100, 300 and 1000 mg/kg bw/day. Pairing of animals within each dose group was undertaken on a one male: one female basis within each treatment group on Day 15 of the study, with females subsequently being allowed to litter and rear their offspring to Day 5 of lactation. During the lactation phase, daily clinical observations were performed on all surviving offspring, together with litter size and offspring weights and assessment of surface righting reflex. There were no treatment related effects observed on mating, fertility or gestation length at any dose level. The offspring litter size, viability, growth and development were all comparable to controls and no adverse effects were noted. Since no treatment-related effects were observed for reproduction, a NOEL for reproductive toxicity was considered to be 1000 mg/kg bw/day. The results of this study are read across to bulk calcium carbonate.

Effects on developmental toxicity

Description of key information

Bulk calcium carbonate showed no signs of developmental toxicity in a prenatal developmental screening toxicity test.

Link to relevant study records

Reference

Reference 1

Endpoint:

developmental toxicity

Type of information:

experimental study

Adequacy of study:

key study

Reliability:

2 (reliable with restrictions)

Rationale for reliability incl. deficiencies:

comparable to guideline study with acceptable restrictions

Qualifier:

equivalent or similar to guideline

Guideline:

OECD Guideline 414 (Prenatal Developmental Toxicity Study)

Deviations:

yes

Remarks:

Minor deviations from the guideline which do not affect the outcome of the study.

Principles of method if other than guideline:

This study was designed to evaluate the developmental effects of moderate dietary calcium increases in rats fed nutritionally adequate diets. Female Charles River CD/VAF Plus rats were given 0.50 (control), 0.75, 1.00 or 1.25% dietary calcium as calcium carbonate in AIN-76A diets for 6 weeks before mating, during mating and for 20 days of gestation. On gestation day 20, the animals were killed and caesarean sections were performed.

GLP compliance:

not specified

Limit test:

no

Species:

rat

Strain:

other: CD/VAF Plus Charles River

Details on test animals or test system and environmental conditions:

TEST ANIMALS
- Source: Charles River Laboratories, Inc. (Wilmington, MA, USA)
- Age at study initiation: Females - 52 days old; Males - 44 days old
- Weight at study initiation: Females - approximately 217 ± 1 g
- Fasting period before study: None
- Housing: Stainless steel cages
- Diet (e.g. ad libitum): ad libitum
- Water (e.g. ad libitum): ad libitum
- Acclimation period: 6 days


ENVIRONMENTAL CONDITIONS
- Temperature (°C): 18 - 27 °C
- Humidity (%): 25 - 72 %
- Air changes (per hr): no data
- Photoperiod (hrs dark / hrs light): 12/12

Route of administration:

oral: feed

Vehicle:

unchanged (no vehicle)

Details on exposure:

DIET PREPARATION
- Rate of preparation of diet (frequency): Once
- Mixing appropriate amounts with (Type of food): AIN-76A Diet
- Storage temperature of food: -1 - -6 °C

Analytical verification of doses or concentrations:

yes

Details on analytical verification of doses or concentrations:

Concentration of calcium analysed using the methods of the Association of Official Analytical Chemists

Details on mating procedure:

- If cohoused:
- M/F ratio per cage: 1:2
- Length of cohabitation: No data - at least overnight for one night
- Verification of same strain and source of both sexes: yes
- Proof of pregnancy: sperm in vaginal smear referred to as day 0 of pregnancy

Duration of treatment / exposure:

Females: 6 weeks prior to mating, through mating to day 20 of gestation

Frequency of treatment:

Rats allowed access to feed ad libitum

Duration of test:

Approximately 62 days

No. of animals per sex per dose:

Females: Pre-mating - 69/dose; Pregnant rats - 45-48/dose
Males: Received test material only during the mating period

Control animals:

other: The group receiving 0.5% calcium were used as the control group

Maternal examinations:

CAGE SIDE OBSERVATIONS: No data

DETAILED CLINICAL OBSERVATIONS: Yes
- Time schedule: No data

BODY WEIGHT: Yes
- Time schedule for examinations: Every 3 days for the 6 weeks before mating and during gestation, but not during mating (females only)

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: No
- Compound intake calculated as time-weighted averages from the consumption and body weight gain data: No
- Food consumption for each animal determined and mean daily diet consumption calculated as g food/animal/day: Yes (females only)

POST-MORTEM EXAMINATIONS: Yes
- Sacrifice on gestation day # 20
- Organs examined: Major organs (no further details reported)

Ovaries and uterine content:

The ovaries and uterine content was examined after termination: Yes
Examinations included:
- Gravid uterus weight: No data
- Number of corpora lutea: Yes
- Number of implantations: Yes
- Number of early resorptions: Yes
- Number of late resorptions: Yes

Fetal examinations:

- External examinations: Yes: all per litter
- Soft tissue examinations: Yes: half per litter
- Skeletal examinations: Yes: half per litter
- Head examinations: No data

Statistics:

The incidence of clinical parameters was analysed by comparing treatment groups with the control group by using Fisher's exact test.
Data on feed consumption were analysed using analysis of variance (ANOVA) and a protected least significant difference (LSD) test (two-tailed).
Weight gains were analysed using analysis of covariance (ANOCOVA) after adjusting for the initial body weight, and an LSD test (two-tailed).
Trend analysis was performed on feed consumption.
The number of corpora lutea, the average number of implantations, the average number of viable foetuses for each litter and the average number of male or female foetuses for each litter were analysed by ANOVA and a LSD test (one-tailed).
Data on implantation efficiency, early deaths, late deaths, and total resorptions (early and late deaths) were analysed by the Freeman-Tukey arcsine transformation for binomial proportions. The transformed data for each litter were then analysed by ANOVA and a LSD test (one-tailed), comparing the control group with experimental groups.
Fisher's exact test was used to analyse data on litters with runts and litters totally resorbed, comparing the controls group with treatment groups.
Foetal body weights and crown-rump lengths were analysed by a nested ANOVA and a LSD (one-tailed).
Data on the specific litter incidence of sternebral, skeletal and visceral variations were analysed using Fisher's exact test.
Proportions of litters with foetuses showing external variations were analysed using Fisher's exact test.
The average number of foetuses with variations per litter was analysed using the Freeman-Tukey arcsine transformation and then an ANOVA and a protected LSD test (one-tailed).
Litters that had foetuses with one or more sternebral, skeletal and visceral variations were analysed with Fisher's exact test.
Trend analysis was performed on the litter data for external variations using the Cox exact one-tailed test for unadjusted positive trend.

Historical control data:

No historical control data were provided

Details on maternal toxic effects:

Maternal toxic effects:no effects

Details on maternal toxic effects:
Clinical findings included alopecia, exudate around the eyes, exudate around the nose, bent teeth, lesion, lump in the left flank or leg and mammary lump. The number of animals with alopecia was significantly increased in animals fed 1.00 % calcium (8.7 vs 0 % in the control) during the 6 weeks before mating and in the groups fed 0.75 and 1.00 % calcium (10 and 16.3 % respectively vs 0 % in the control) during gestation.
One non-pregnant animal was observed to have diarrhoea on day 4 after assignment to the 1.25 % calcium diet.
One pregnant female fed 1.00 % dietary calcium developed bleeding from the vagina on days 15 and 16 of gestation. The incidence of diarrhoea and bleeding from the vagina was not considered to be related to the dietary calcium.
Three females died during the study, but the deaths were not attributable to excess calcium in the diet.
Values for mean daily feed consumption per female given excess dietary calcium for 6 weeks before mating and during gestation are shown in Table 1. The 0.75, 1.00 and 1.25 % groups ate slightly more than the controls during most of the study, but not all the increases were statistically significant.
Mean values for body weight gain for animals given excess dietary calcium for 6 weeks before mating and gestation are given in Table 2. Although some differences in weight gain between control and treated groups were significant, there was no consistent pattern of increase or decease in the body weight gain.
Maternal reproductive parameters are listed in Table 3. There were no adverse reproductive effects associated with ingestion of excess dietary calcium. Significant increases were observed in implantation efficiency at 1.25 % dietary calcium and in the average number of viable female foetuses at 0.75 and 1.25 % dietary calcium. These increases were not dose related and were not considered to be caused by dietary calcium.

Key result

Dose descriptor:

NOAEC

Effect level:

> 1.25 other: % w/w calcium

Basis for effect level:

other: maternal toxicity

Details on embryotoxic / teratogenic effects:

Embryotoxic / teratogenic effects:no effects

Details on embryotoxic / teratogenic effects:
There were no adverse effects to the foetuses caused by feeding excess dietary calcium to the dams (Table 3). Foetal body weights and crown-rump lengths of male and female animals given excess dietary calcium were similar to those of the control groups. Feeding excess dietary calcium had no effect on the number of litters with male or female runts. The incidences of specific external variations are shown in Table 4. In the control group, one foetus did not possess a mouth (astomia) and a lower jaw (agnathia). At the 1.25 % dietary calcium level, one foetus had multiple anomalies: agnathia, cleft palate, protruding tongue, beak-like snout, malformed head, open eye on the right side, oedema of the trunk, omphalocele, exposed heart and lungs (diaphragmatic cleft), club foot on the left front paw, scoliosis, imperforate anus and kinked tail. The occurrence of this foetus with multiple anomalies was not thought to be related to feeding the rats excess dietary calcium before and during pregnancy. The data in table 4 were analysed by trend analysis and Fisher's exact test. The Cox exact trend test showed no significant positive trends in the litter incidence of individual or combined variations. When analysed by Fisher's exact test, there were no significant differences between treated and control groups in the litter incidences of these same parameters. Under the conditions of the study, there were no significant increases in the litter incidence of specific or combined external variations. The numbers of foetuses with sternebral ossification deficiencies and the litter incidences are summarised in Table 5. A comparison of the proportion of litters with foetuses having sternebral variations showed no significant differences among the groups given excess calcium and the control group. The increase in the number of litters with foetuses with bipartite sternebrae in the 1.25 % group was not significant (p=0.0854). The incidences of sternebral variations are presented in Table 6. The variations counted are based on the classification of the types and not on the individual sternebrae affected. The average number of foetuses per litter with one or more sternebral variations increased in the 0.75, 1.00 and 1.25 % groups, but the differences were not significant. There was no significant increase in the average number of foetuses per litter with at least two or three sternebral variations in the groups fed excess dietary calcium. Statistical analysis of the proportion of litters that had foetuses with variations (at least one, two or three) showed no significant increase between the excess calcium and the control groups. The incidences of specific skeletal variations, excluding sternebrae, were not dose related and there were no significant differences among any of the values (Table 7). The average number of sacral and caudal vertebrae was not affected by dietary calcium (data not shown). The average number of foetuses per litter with at least one skeletal variation was lower in the 0.75, 1.00 and 1.25 % dietary calcium groups than in the control (Table 8). However, the ANOVA of the Freeman-Tukey arcsine-transformed data showed no differences in the average number of foetuses with variations (p>0.10). The proportion of litters that had foetuses with variations (at least one, two or three) showed no dose-related or significant differences between any of the values obtained in this study. There was no dose related effect on the incidence of any other type of soft-tissue variation (Table 9). The proportion of litters with hydrocephalus was greater in the control group than in the treatment groups, but the differences were not significant. The incidence of visceral problems by foetus and by litter is shown in Table 10. The average number of foetuses with at least one, two or three soft-tissue variations was similar in all groups. Although the average number of foetuses with at least one variation was lower in the 1.00 and 1.25 % groups, ANOVA did not indicate a significant decrease compared with controls. Although there was a dose-dependent decrease in the proportion of litters with foetuses with at least one variation. Fisher's exact test (two-tailed) did not show any significant difference. Thus the proportion of litters with foetuses with at least one two or three soft-tissue variations was similar in all groups.

Key result

Dose descriptor:

NOAEC

Effect level:

1.25 other: % w/w calcium

Based on:

test mat.

Sex:

male/female

Basis for effect level:

other: developmental toxicity

Abnormalities:

not specified

Developmental effects observed:

not specified

Table 1: Mean daily feed consumption (g/animal/day) of rats given excess dietary calcium.

Concentration of Calcium in the Diet (%)
Days	0.5	0.75	1.00	1.25
	Non-pregnant rats 6 weeks before mating
	(69)	(69)	(69)	(69)
0-6	15.2 ± 0.4	15.7 ± 0.3	15.8 ± 0.3	16.4 ± 0.4
6-12	17.4 ± 0.3	17.7 ± 0.3	17.8 ± 0.3	18.2 ± 0.3
12-18	17.0 ± 0.3	17.6 ± 0.3	18.1 ± 0.3^b	18.4 ± 0.3^c
18-24	17.9 ± 0.3	18.1 ± 0.3	18.8 ± 0.3^a	18.9 ± 0.3^b
24-30	17.6 ± 0.3	17.9 ± 0.3	18.8 ± 0.3^b	18.6 ± 0.3^a
30-36	17.8 ± 0.3	17.7 ± 0.3	18.6 ± 0.3^d	18.4 ± 0.2
36-42	17.8 ± 0.3	17.8 ± 0.3*	18.6 ± 0.3^a	18.5 ± 0.3^d
0-42	17.3 ± 0.2	17.5 ± 0.2*	18.1 ± 0.2^a	18.2 ± 0.2^b
	Pregnant rats from days 0-20
	(45)	(47)	(44)	(48)
0-6	19.6 ± 0.4	20.4 ± 0.4	20.8 ± 0.4^a	21.0 ± 0.3^b
6-12	22.8 ±0.4	23.3 ± 0.3	23.9 ± 0.4^d	24.2 ± 0.4^a
12-18	23.5 ± 0.4	24.3 ± 0.3	24.1 ± 0.5	24.2 ± 0.3
18-20	23.7 ± 0.5	24.0 ± 0.4	23.7 ± 0.6	23.0 ± 0.5
0-20	22.2 ± 0.4	22.8 ± 0.3	23.0 ± 0.4	23.1 ± 0.3

* Because one animal was autopsied in extremis on day 40 in the 0.75% group the number of rats was reduced to 68 for the data for days 36 -42 and 0 -42.

Number of animals are indicated in parantheses. Values are means ± SEM. superscripts indicate differences between control and test values ( ^aP ≤0.05; ^bP≤0.01; ^cP ≤0.001; ^d 0.05 P 0.01 )

Table 2: Mean body weight gains and initial body weights of rats given excess dietary calcium

Concentration of Calcium in the Diet (%)
Days	0.5	0.75	1.00	1.25
	Body weight  gain (g) of non-pregnant rats 6 weeks before mating
	(69)	(69)	(69)	(69)
0-6	8.1 ± 1.4	9.9 ± 1.2	9.1 ± 1.1	7.7 ± 1.4
6-12	22.1 ± 1.1	21.1 ± 0.9	21.7 ± 1.1	20.1 ± 1.1
12-18	12.1 ± 0.8	14.7 ± 0.9^a	16.1 ±1.0^a	16.3 ± 0.7^a
18-24	16.3 ± 0.8	14.0 ± 0.7^a	15.7 ± 0.9	14.1 ± 0.7^a
24-30	9.0 ± 0.6	9.8 ± 0.8	11.9 ± 0.9^a	10.6 ± 0.7
30-36	11.0 ±0.9	9.2 ± 0.8	10.1 ± 1.0	9.6 ± 0.8
36-42	5.3 ± 0.7	5.8 ± 0.9*	4.9 ± 0.8	6.4 ± 0.6
0-42	83.9 ± 2.9	85.1 ± 3.1*	89.5 ± 3.0	84.7 ± 2.6
	Initial body weight (g) of non pregnant rats
0	217.7 ± 1.0	217.6 ± 1.0	217.4 ± 1.0	217.7 ± 1.0
	Body weight gain (g) of pregnant rats
	(45)	(47)	(44)	(48)
0-6	29.8 ±1.2	29.7 ± 1.3	29.8 ± 1.1	29.4 ± 1.0
6-12	30.4 ± 1.0	32.0 ± 0.8	30.2 ± 0.7	31.9 ± 0.9
12-18	45.2 ± 1.8	51.1 ± 1.4^a	48.0 ± 2.6	48.9 ± 1.5
18-20	25.8 ± 1.1	27.9 ± 0.9	26.2 ± 1.1	25.1 ± 1.0
0-20	131.2 ± 3.1	140.7 ± 2.6	134.2 ± 3.9	135.3 ± 3.0
	Initial body weight (g) of pregnant rats
0	297.6 ± 3.6	300.8 ± 3.7	306.5 ± 4.8	300.0 ± 3.8

* Because one animal was autopsied in extremis on day 40 in the 0.75% group the number of rats was reduced to 68 for days 36-42 and 0 -42

Values marked with superscripts indicate significant differences between control and test values ^aP ≤0.05; ^bP≤0.01; ^cP ≤0.001; ^d 0.05 P 0.10

Table 3: Maternal and reproductive outcome at autopsy and foetal data from rats given excess dietary calcium

Concentration of Calcium in the Diet (%)
Parameter	0.5	0.75	1.00	1.25
No. of pregnant females	45	47	44	48
No. of corpora lutea/female	16.27 ± 0.4	16.83 ± 0.40	16.52 ± 0.27	15.93 ± 0.32
Implantation frequency  (% female)*$	82.01 ± 3.73	90.29 ± 1.89	86.51 ± 3.46	93.41 ± 2.28^b
No of implants/female*	13.58 ± 0.68	15.26 ± 0.43	14.25 ±  0.60	15.15 ± 0.50
No of viable foetuses/female*	11.82 ± 0.66	13.53 ± 0.46	12.82 ± 0.65	13.54 ± 0.58
No of viable foetuses/litter*	6.29 ± 0.43	6.49 ± 0.33	6.95 ± 0.49	6.83 ± 0.37
Viable female foetuses/litter*	5.53 ± 0.39	7.04 ± 0.38^b	5.86 ± 0.37	6.71 ± 0.43^a
Resorptions/female (mean %)*‡	12.50 ± 1.98	11.15 ± 1.73	12.92 ± 3.19	13.81 ± 3.09
No of early deaths/litter$	1.71	1.70	1.41	1.56
No. of late deaths/litter$	0.04	0.02	0.02	0.04
No of early and late deaths/litter$‡	1.76	1.72	1.43	1.60
No of litters totally resorbed	0	0	1	2
Body weight (g)
Males	3.84	3.90	3.83	3.79
Females	3.61	3.69	3.66	3.58
Crown rump length (cm)
Males	4.0	4.0	4.0	4.0
Females	3.9	3.9	3.9	3.9
No of runts
Males	5	5	3	3
Females	4 (4)	4 (1)	1 (1)	3 (2)

* Means ± SEM

$ Statistical analysis was performed on these data after application of the Freman-Tukey arcsine transformation

‡ The number of resorptions includes early deaths and late deaths per litter

§ The number of litters is indicated in parentheses

Values marked with superscripts indicate significant differences between control and test values ^aP ≤0.05; ^bP≤0.01; ^cP ≤0.001; ^d 0.05 P 0.10

Table 4: Incidence of specific external variations in foetuses from rats given excess dietary calcium

Concentration of Calcium in the Diet (%)
Parameter	0.5	0.75	1.00	1.25
No. foetuses (litters examined)	532 (45)	636 (47)	564 (43)	650 (46)
No of foetuses (litters with variations)	2 (2)	2 (2)	1 (1)	8 (6)
Heamorrhages, external	1 (1)	-	1 (1)	2 (2)
Cleft palate	-	1 91)	-	2 (2)
Agnethia	1 (1)	-	-	1 (1)
Astomia	1 (1)	-	-	-
Thread like tail	-	1 (1)	-	1 (1)
Kinked tail	-	-	-	2 (2)
Oedema	-	-	-	3 (2)
Malformed head	-	-	-	1 (1)
Beaked snout	-	-	-	1 (1)
Protruding tongue	-	-	-	1 (1)
Open eye	-	-	-	1 (1)
Diaphragmatic cleft*	-	-	-	1 (1)
Omphalocele	-	-	-	1 (1)
Club foot	-	-	-	1 (1)
Seollosis	-	-	-	1 (1)
Imperforate anus	-	-	-	2 (2)

The number of litters is indicated parentheses

* This parameter includes exposed heart and lungs

Table 5: Incidence of specific external variations in foetuses from rats given excess dietary calcium

Concentration of Calcium in the Diet (%)
Parameter	0.5	0.75	1.00	1.25
No. foetuses (litters examined)	251 (44)	305 (47)	273 (43)	313 (46)
No of foetuses with specific stenebral variations
Incomplete ossification	74 (31)	84 (35)	88 (36)	95 (35)
Bipartite	4 (4)	3 (3)	4 (3)	10 (10)^d
Unossified	50 (27)	49 (27)	40 (18)	54 (31)
Malagnified	8 (7)	2 (2)	10 (9)	11 (11)
Fused	1 (1)	0 (0)	0 (0)	1 (1)

The number of litters is indicated parentheses

Superscript indicates differences between control and test values (^d0.05  P 0.10)

Table 6: Incidence of sternebral variations in foetuses of rats given excess dietary calcium

Concentration of Calcium in the Diet (%)
Parameter	0.5	0.75	1.00	1.25
Sternebral variations*
Total no.	137	138	142	171
Mean no./litter	3.11	2.94	3.30	3.72
Foetuses with one or more sternbral variations
Total no.	105	118	112	138
Mean no./litter$	2.39	2.51	2.60	3.00
Foetuses affected (%)	41.83	38.69	41.03	44.09
Litters with foetuses with one or more variations
Total no.	37	42	38	39
Litters affected (%)	84.09	89.36	88.37	84.78
No of foetuses examined	251	305	273	313
No of litters examined	44	47	43	46

* Statistical analysis was not performed on this parameter

$ Statistical analysis was performed on these data after application of the Freeman-Tukey arcsine transformation

Conclusions:

Both the non-pregnant and pregnant rats in the 0.75, 1.00 and 1.25% groups ate slightly more than did the control group during most of the intervals measured, but not all the increases were statistically significant. There was no consistent pattern of increase or decrease in weight gain. No dose-related changes were found in maternal clinical findings, the average number of implantations, resorptions and viable foetuses, or foetal length or weight. Under the conditions of the study, there were no statistically significant increases as compared with the control group in the litter incidence regarding specific external, visceral or skeletal variations of the foetuses. Dietary calcium was neither foetotoxic nor teratogenic at the concentrations used.

Therefore, as no adverse effects were noted at this dose level (1.25% Ca in diet), the NOAEL for teratogenic and maternal toxic effects in rats is in excess of 1963 - 2188 mg/kg bw/day of calcium carbonate.

Reference 2

Endpoint:

developmental toxicity

Type of information:

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

Adequacy of study:

supporting study

Justification for type of information:

1. HYPOTHESIS FOR THE ANALOGUE APPROACH
Bulk calcium carbonate and uncoated nano calcium carbonate are chemically identical. The only difference between them is the particle size, which may result in minor differences in physico-chemical properties. However, in biological systems and in the environment, both bulk and uncoated nano calcium carbonate are expected to have identical (eco)toxicological and environmental fate properties.
2. SOURCE AND TARGET CHEMICAL(S) (INCLUDING INFORMATION ON PURITY AND IMPURITIES)
Uncoated nano calcium carbonate (Source): Refer to IUCLID section 1.2 for information on purity and impurities.
Bulk calcium carbonate (Target): Refer to IUCLID section 1.2 for information on purity and impurities.
The majority of new studies have been performed using uncoated nano calcium carbonate as the test material, hence it has been designated as the ‘Source’ in the read-across justification. However, for some endpoints data on bulk calcium carbonate are available and used to read-across to uncoated nano calcium carbonate so for these endpoints, marked with a * in the read-across justification document, bulk calcium carbonate is the ‘Source’.
3. ANALOGUE APPROACH JUSTIFICATION
Please refer to the justification attached in Section 13 of the dataset
4. DATA MATRIX
Please refer to the justification attached in Section 13 of the dataset

Reason / purpose for cross-reference:

read-across source

Reason / purpose for cross-reference:

read-across: supporting information

Vehicle:

water

Dose descriptor:

NOEL

Effect level:

1 000 mg/kg bw/day (actual dose received)

Based on:

test mat.

Basis for effect level:

other: developmental toxicity

Abnormalities:

not specified

Developmental effects observed:

not specified

Effect on developmental toxicity: via oral route

Endpoint conclusion:

no adverse effect observed

Study duration:

subchronic

Species:

rat

Effect on developmental toxicity: via inhalation route

Endpoint conclusion:

no study available

Effect on developmental toxicity: via dermal route

Endpoint conclusion:

no study available

Additional information

A prenatal developmental toxicity study was performed to evaluate the developmental effects of moderate dietary calcium increases in rats fed nutritionally adequate diets (Shackelford et al, 1993). The test method used was not reported; however, it is considered to be similar to OECD TG 414. Female Charles River CD/VAF Plus rats were given 0.50 (control), 0.75, 1.00 or 1.25% dietary calcium as calcium carbonate in AIN-76A diets for 6 weeks before mating, during mating and for 20 days of gestation. On gestation day 20, the animals were killed and caesarean sections were performed.

Both the non-pregnant and pregnant rats in the 0.75, 1.00 and 1.25% groups ate slightly more than did the control group during most of the intervals measured, but not all the increases were statistically significant. There was no consistent pattern of increase or decrease in weight gain. No dose-related changes were found in maternal clinical findings, the average number of implantations, resorptions and viable foetuses, or foetal length or weight. Under the conditions of the study, there were no statistically significant increases as compared with the control group in the litter incidence regarding specific external, visceral or skeletal variations of the foetuses. Dietary calcium was neither foetotoxic nor teratogenic at the concentrations used.

Since no adverse effects were noted at the highest dose level tested (1.25% Ca in diet), the NOAEL for teratogenic and maternal toxic effects in rats is in excess of 1.25% Ca, equivalent to approximately 1963 - 2188 mg/kg bw/day of calcium carbonate.

Toxicity to reproduction: other studies

Additional information

A study is available in the literature which investigates how dietary calcium and lead interact to modify maternal blood pressure, erythropoiesis and foetal neonatal growth in rats during pregnancy and lactation (Bogden et al, 1995). Female rats were fed dietary calcium carbonate at concentrations of 0.1, 0.5 and 2.5 g/100g. After one week the animals were mated and after pregnancy was confirmed the animals were fed drinking water containing either 0 or 250 mg/L of lead. This treatment was continued for the duration of pregnancy and for one week of lactation. Three control groups were fed the same diets without lead exposure.

The dietary calcium concentrations that were used did not interfere with the ability of the rats studied to maintain pregnancy and deliver normal pups.

 

A human study investigating the long term effect of calcium supplementation during pregnancy on the blood pressure of the offspring is reported in the literature (Belizan et al, 1997). The trial examined the effectiveness of 2 g of elemental calcium supplementation per day (four tablets of calcium carbonate 500 mg) for the prevention of hypertensive disorders of pregnancy. Supplementation was started at 20 weeks' gestation and continued until delivery.

The reports of women with anaemia, premature rupture of membranes, diabetes mellitus, third trimester haemorrhage, and receiving medical treatment, and the numbers of hospital admissions, inductions of labour, and caesarean sections were closely similar in the calcium and placebo groups.

At birth the weight, length, gestational age, and the rate of preterm delivery were closely similar between the two groups. A higher proportion of the newborn infants in the calcium group had an Apgar score <7 at the first minute (20/248 vs 10/251), and more newborns were admitted to the neonatal intensive care unit (38/248 vs 29/251) (not significant). However, these results were not statistically significant.

Therefore calcium supplementation at a dose of 2 g/day during pregnancy did not cause any adverse effects in either the mother or child.

 

Clinical cases are available in the literature which report the excessive ingestion of antacid tablets during pregnancy, potentially leading to the development of hypercalcaemia (milk-alkali syndrome). The amount of calcium carbonate ingested was significantly greater than that which would be consumed in the diet and from following the recommended guidelines for antacid consumption (one patient consumed 7,500 – 10,500 mg calcium carbonate/day from midway through the first trimester to the onset of labour). Following hospital treatment both the mother and child were reported to be fully recovered with no lasting side effects or teratogenic effects.

Justification for classification or non-classification

Under the conditions of the OECD TG 422 study, uncoated nano calcium carbonate administered to male and female rats up to a dose level of 1000 mg/kg bw/day for a period of up to 48 days is not toxic to reproduction and has no effect on fertility or development. No treatment-related effects were observed for reproduction; hence, a NOEL for reproductive toxicity was considered to be 1000 mg/kg bw/day. The results of this study are read across to bulk calcium carbonate. The prenatal developmental toxicity study also demonstrated that calcium carbonate was neither foetotoxic nor teratogenic at the concentrations used. Since no adverse effects were noted at the highest dose level tested (1.25% Ca in diet), the NOAEL for teratogenic and maternal toxic effects in rats is in excess of 1.25% Ca, equivalent to approximately 1963 - 2188 mg/kg bw/day of calcium carbonate.

Based on these results, bulk calcium carbonate does not warrant classification for reproductive/developmental toxicity under CLP.