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Key value for chemical safety assessment

Effects on fertility

Effect on fertility: via oral route
Endpoint conclusion:
no adverse effect observed
Dose descriptor:
NOAEL
20 mg/kg bw/day
Species:
rat
Additional information

The reproductive toxicity of zinc compounds has been investigated in one and two generation reproductive toxicity studies in which rats or mice were dosed by gavage or via the diet with soluble zinc compounds (i. e., zinc chloride, zinc sulphate) at exposure levels up to 14 mg Zn/kg bw/day (gavage) or 200 mg Zn/kg bw/day (diet) (Khan et al.,2001, 2003, 2007; Samanta et al, 1986). Further information on potential effects of zinc compounds on male or female reproductive organs could be retrieved from subchronic toxicity studies as conducted by Maita et al.(1981) and Edwards and Buckley (1995).

The most recent one and two generation reproductive toxicity studies conducted by Khan et al.,(2001, 2003, 2007) in rats and mice with zinc chloride provide the most coherent picture on the effects of zinc. All these studies have in common that while effects on fertility such as reduced litter size in F1 and F2 generation have been determined, these were only noticeable at doses which resulted in toxic effects in the dam.

Maita et al. (1981) reported that mice and rats fed with zinc sulphate in dietary concentrations up to 30,000 mg/kg feed did not produce adverse effects on either male or female sex organs after 13 weeks of exposure. This dietary level was equal to ca. 1100 mg or 565 mg Zn/kg bw/day for mice and rats, respectively. Edwards and Buckley (1995) showed that rats exposed to 13 or 60 mg Zn/kg bw/day in the diet over a period of 90 days did not show any detrimental effects on sex organs. In the exposure group of 335 mg Zn/kg bw/day, all males showed hypoplasia in testes and seminiferous tubules in males hypoplastic uterus in females, but these findings are not considered reliable as the animals of this high dose group were generally of poor health conditions and killed for humane reasons prior to study termination.

In addition to those key reproductive toxicity studies(Khan et al.,2001, 2003, 2007; Samanta et al, 1986), some additional studies indicating high oral doses of zinc (i.e., exposures greater that 25 mg day/kg bw/day) to impair fertility as indicated by a decreased number of implantations sites and increased number of resorptions are of note:

A study was carried out to determine the effect of zinc supplementation on the number of implantation sites and resorptions in pregnant rats. The control group consisting of 12 pregnant females was maintained on 10 % vegetable protein diet (containing 30 ppm zinc) from Day 1 through Day 18 of pregnancy. The experimental group consisting of 13 animals was also maintained on the same diet, but received additionally 150 ppm zinc as a 2% zinc sulphate solution administered daily orally. All the animals were sacrificed on Day 18 of pregnancy, and their uteri examined for implantation sites and resorptions. Of a total number of 101 implantation sites in the 12 control animals there were two resorptions, one in each of two animals. In marked contrast, in the 13 zinc supplemented animals, there were 11 resorptions out of 116 implantations. Eight of the animals had at least one resorption each. This difference was statistically significant. The result indicates that oral administration of moderately high levels of zinc (150 ppm) may be associated with harmful effects in the course of pregnancy of rat (Kumar et al., 1976). The low protein diet may have affected the physiology of the animals resulting in an increased sensitivity for zinc. As this hypothesis cannot be further assessed and also considering the limited available study information, this study is only of limited validity for the assessment of effects of zinc exposure on fertility (EU RAR, 2004).

Another study aimed at determining the effect of post-coitum, and pre- and post-coitum dietary zinc supplementation on the conception in the Charles-Foster rat. In the post-coitum study (test 1), two groups of 15 pregnant rats were fed 0 and 4,000 ppm zinc as zinc sulphate in diet (i.e., approximately 200 mg Zn/kg bw/day) from day 1 through day 18 of pregnancy. In the pre- and post-coitum study (test 2), two groups of 15 female rats were treated with same doses for 21 days pre-mating period, maximum 5 days of mating period and 18 days of post-coitum period. All the females were sacrificed on Day 18 of gestation and uterus content and fetuses were examined. In test 1, significant decrease in the incidences of conception and number of implantation sites per mated female was observed in the treatment group with respect to the control group. However, the difference in implantation sites when considered per pregnant female was not significant. In test 2, no significant difference in incidences of conception and implantation sites was observed in the control and treatment groups. In both the tests, there was no treatment-related change in the fetal and placental weights, stillbirths and malformed fetuses were absent and the number of resorption sites was negligible. Based on these results, dietary zinc supplementation at 4,000 ppm did not affect the fetal growth in pregnant rats. This dose, however, altered the normal conception when started after coitus but showed no effect when initiated sufficient time before coitus (Pal et al., 1987).

The available information suggests that high oral doses of zinc (i. e., exposure levels greater than 20 mg Zn/kg bw/day) may adversely affect spermatogenesis and result in impaired fertility indicated by decreased number of implantation sites and increased number of resorptions (US EPA, 2005). However, these effects were only observed in the presence of maternal toxicity as seen in the one or two generation studies conducted by Khan et al.(2001, 2003, 2007) or, in case of the study conducted by Kumar et al. (1976), when other study non-zinc relevant study specificities could have impacted the study outcome.

Overview of experimental key studies on fertility

Test substance

Method

Results

Remarks

Reference

Zinc chloride

One-generation study in rats administered zinc chloride at doses of 0, 3.6, 7.2, 14.4 mg Zn/kg bw/d in water over one generation by gavage. Exposure started 77 days prior to mating

As of 3.6 mg Zn/kg bw/day:

P - Mortality; body weight gain; fertility indext; thymus atrophy

F1 - litter size (non significant); number of surviving pubs (non significant);

As of 7.2 mg Zn/kg bw/day:

P – hemosidosis of spleen; lymphocyte deficiency

F1 - number of surviving pubs; BW gain (PND 21)

2 (reliable with restrictions)

key study

Khan et al., 2001

Zinc chloride

One-generation study in mice administered zinc chloride at doses of 0, 0.75, 1.5 and 3, mg Zn/kg bw/d (male)respectively, 0. 1.5, 3 and 6, mg Zn/kg bw/d (female) in water with 1.5mL HNO3/l over one generation by gavage. Exposure started 49 days prior to mating

0.75 resp. 1.5 mg Zn/kg bw/day:

P- Mortality; body weight gain; abs./rel. Liver/thymus/ spleen weight; fertility indext; number pregnancies

F1- litter size (non significant); number of surviving pubs (non significant);

1.5 resp. 3 mg Zn/kg bw/day:

P- body weight gain; F1– 14day survival index;

3 resp. 6 mg Zn/kg bw/day:

F1– only 1 birth; 9 still births.

2 (reliable with restrictions)

key study

Khan et al., 2003

Zinc chloride

Two-generation study in rats administered zinc chloride at doses of 7.5, 15and 30 mg zinc chloride/kg bw/d (3.6, 7.2 and 14.4 mg Zn/kg bw/day) in water over two successive generations via the oral route. Application procedure not specified but likely oral gavage. Exposure started 77 days prior to mating.

As of 3.6 mg Zn/kg bw/day:

P - Mortality; body weight gain; abs/rel liver/kidney weight; lesions in GI tract, inflammation in prostate

F1 - Mortality; body weight gain; abs/rel brain/prostate/spleen weight;

F2 – no effects

7.2 mg Zn/kg bw/day:

P – abs./rel. brain/seminal vesicle weight;F1 - abs/rel liver/adrenal/seminal vesicle weight

F2 – no effects

14.4 mg Zn/kg bw/day:

P – abs./rel. Spleen/uterus weight;

F1 - body weight gain (PND21); abs/rel kidney weight; litter size and #surviving pubs until PND4;

F2 – body weight gain (PND21); abs/rel kidney weight; litter size and number surviving pubs until PND4;

Maternal toxicity at any dose level. The NOAEL for fertility and development toxicity is about 15 mg ZnCl2/kg bw/d, this corresponds to 7.2 mg Zinc/kg bw/day. No NOAEL for systemic toxicity could be derived.

2 (reliable with restrictions)

key study

Khan et al., 2007

Zinc sulphate

Charles foster rats fed with a diet containing 4000ppm Zn (in form of zinc sulphate); exposure equals 200 mg Zn/kg bw exposure started 30-32 days prior to mating.

200 mg Zn/kg bw/day

P – Zn-concentration in testis and sperm; sperm mobility; number of pregnancies

F1 – number of live births

2 (reliable with restrictions)

key study

Samanta et al., 1986

Effects on Fertility, Human information

In reviews by the World Health Organisation in the Environmental Health Criteria for Zinc (WHO, 2001) and by the US Agency for Toxic Substances and Disease Registry in the Toxicity Profile for Zinc (ATSDR, 2005), existing human studies which examined the responses of women to zinc supplementation during pregnancy have been summarised. Studies on large controlled trials that were conducted to investigate the effects of dietary zinc supplementation in healthy pregnant women were peer reviewed. The reviewers concluded that zinc at a rate of 20mg/day and 30 mg/day did not result in any adverse reproductive effects during pregnancy (Hunt et al.,1984; Kynast and Saling et al.,1986).Two exemplar studies are summarised in the following:

A double blind trial was conducted in 56 pregnant women at risk of delivering a small for gestational-age baby to determine the effects of dietary zinc supplementation during the last 15-25 weeks of pregnancy following administration of 22.5 mg zinc/day. No adverse reproductive effects were observed (Simmer et al.,1991).

Pregnant women who received 0.3 mg zinc/kgbw/day as zinc sulphate capsules during the last two trimesters did not exhibit any changes in maternal body weight gain, blood pressure, postpartum haemorrhage or infection, inidicating no adverse reproductive effects (Mahomed et al.,1989).


Short description of key information:
A range of studies have been conducted to assess the effects of zinc on fertility and reproductive performance, most of them with very soluble zinc chloride and zinc sulphate. A complete overview and review of available fertility studies is available in the EU risk assessment of zinc compounds (EU RAR, 2004), the review of the of health effects of zinc compounds by the US Agency for Toxic Substances and Disease Registry (ATSDR, 2005), the toxicological review of zinc and compounds by the US Environmental Protection Agency (US EPA, 2005) or the review by the WHO (WHO, 2001). The results of the key experimental studies addressing potential effects of zinc compounds on fertility are summarised in the CSR.

Effects on developmental toxicity

Description of key information
Several prenatal toxicity studies are available that examined the developmental toxicity of various zinc compounds in rats, mice, rabbit or hamsters up to dietary exposure levels of 200 mg Zn/kg bw/day or 50 mg Zn/kg bw/day by gavage (for details see Table developmental toxicity studies). No developmental toxicity has been observed in these studies and there NOAEL’s have been established at the highest doses tested.
Although some developmental effects such as decreases in body weights or decrease in individual organ weights were observed in F1 and/or F2 generations in the one or two generation reproductive toxicity studies conducted by Khan et al. (2007) at high exposure levels, these observations are, however, not suitable for risk assessment or hazard classifications as they were always accompanied with maternal toxicity. Moreover, no developmental toxicity was observed at non-maternally toxic doses in a teratogenicity study in which CF-1 albino mice were administered intraperitoneally 0, 12.5, 20.5 and 25 mg/kg on Day 11 of gestation (test 1) and at 20.5 mg/kg on Days 8 -11 of gestation (test 2) (Chang et al., 1979).
Effect on developmental toxicity: via oral route
Endpoint conclusion:
no adverse effect observed
Dose descriptor:
NOAEL
50 mg/kg bw/day
Species:
rat
Additional information

The developmental toxicity of zinc compounds can be assessed on the basis of prenatal toxicity studies that have been conducted with soluble zinc sulphate and zinc chloride and slightly soluble zinc carbonate in rats, mice, hamsters or rabbits. Moreover, a total of three (one or two generation) reproductive toxicity studies conducted by Khan et al.(2001, 2003, 2007) provide further information on potential teratogenic effects of zinc compounds.

No prenatal toxicity was observed with either zinc sulphate, zinc chloride or zinc carbonate at exposure levels up to 50 mg Zn/kg bw/day by oral gavage or 200 mg Zn/kg bw/day if the zinc was dosed via the diet. Established NOAELs in these studies were typically at highest dose tested and systemically tolerated by the dams. Developmental effects such as decrease in body or organ weights were, however, observed in F1 and/or F2 generations in the one or two generation reproductive toxicity studies conducted by Khan et al. (2001, 2003, 2007). These studies are not considered suitable for the assessment of teratogenic effects for hazard classification or risk assessment purposes since they were always observed in the presence of maternal toxicity.

Overview of experimental key studies on developmental toxicity

Test substance*

Species

Route

Method

Result

Remark

Reference

Zinc sulphate

Mouse

CD-1

Oral

Females received daily doses of 0, 0.3, 1.4, 6.5 and 30 mg ZnSO4(unspecified)/kg bw by oral gavage during days 6-15 of gestation.

No discernible effects were seen on or maternal or foetal survival. No difference in number of abnormalities found in foetuses.

NOAEL:

30 mg/kg bw/day equalling

12mg Zn/kg bw/d (anhydrate);

6.8mg Zn/kg bw/d (heptahydrate)

2 (reliable with restrictions)

Key study

Food and Drugs Research Labs., Inc, 1973*

Zinc sulphate

Rat

Wistar

Oral

Females received daily doses of 0, 0.4, 2.0, 9.1 and 42.5 mg ZnSO4(unspecified)/kg bw by oral gavage during days 6-15 of gestation.

No discernible effects were seen on or maternal or foetal survival. No difference in number of abnormalities found in foetuses.

NOAEL:

42.5 mg/kg bw/day equalling

17mg Zn/kg bw/d (anhydrate);

9.6 mg Zn/kg bw/d (heptahydrate)

2 (reliable with restrictions)

Key study

Food and Drugs Research Labs., Inc, 1973*

Zinc sulphate

Rat

Charles Foster

Oral

Females received daily doses of 0, and 200 mg Zn/kg bw (in form of ZnSO4) in diet during days 1-18 of gestation

No discernible effects were seen on or maternal or foetal survival. A reduced number of implantations observed. No difference in number of abnormalities found in foetuses.

NOAEL:

200 mg/kg bw/day

2 (reliable with restrictions)

Key study

EU RAR, 2004

Zinc sulphate

Hamster

Oral

Females received daily doses of 0, 0.9, 4.1, 19, and 88 mg ZnSO4(unspecified)/kg bw by oral gavage during days 6-10 of gestation.

No discernible effects were seen on or maternal or foetal survival. No difference in number of abnormalities found in foetuses.

NOAEL:

88 mg/kg bw/day

equalling

32.5mg Zn/kg bw/d (anhydrate);

19.9mg Zn/kg bw/d (heptahydrate);

2 (reliable with restrictions)

Key study

Food and Drugs Research Labs., Inc, 1973*

Zinc sulphate

Rabbit

Dutch

Oral

Females received daily doses of 0, 0.6, 2.8, 13 and 60 mg ZnSO4(unspecified)/kg bw during days 6-18 of gestation.

No discernible effects were seen on or maternal or foetal survival. No difference in number of abnormalities found in foetuses.

NOAEL:

60 mg/kg bw/day

equalling

24mg Zn/kg bw/d (anhydrate);

13.6mg Zn/kg bw/d (heptahydrate)

2 (reliable with restrictions)

Key study

Food and Drugs Research Labs., Inc, 1974*

Zinc carbonate

Rat

Sprague Dawley

Oral

Females received daily doses of 0, 2.5, and 50 mg Zn/kg bw (in form of ZnCO3) in diet during days 1-20 of gestation.

No discernible effects were seen on or maternal or foetal survival. No difference in number of abnormalities found in foetuses.

NOAEL:

50 mg/kg bw/day

 

2 (reliable with restrictions)

Key study

Uriu-Hare, 1989

* ZnSO4form is unspecified. The NOAEL, expressed as Zn cation, has been calculation for both anhydrate- and heptahydrate forms.

Developmental toxicity, Human information

In establishing the Environmental Health Criteria for Zinc, the World Health Organisation has reviewed and summarised existing human studies examining the responses of women to zinc supplementation during pregnancy. None of the studies indicated any significant effects on the developing foetus (WHO, 2001). Two exemplar studies are summarised in the following:

A study was conducted on pregnant women to determine the effects of nutrients during pregnancy on maternal and fetal outcome. Four hundred fifty women were observed during pregnancy and postpartum. Forty-three variables including 12 laboratory indices of maternal nutrient status were assessed. Maternal plasma zinc levels were inversely correlated with fetal weight. Blood examinations revealed a significant association between the total occurrence of fetomaternal complications or fetal distress, and lowest quartile zinc/albumin and highest quartile folate. Under the study conditions, plasma zinc was determined to be a discriminator for fetomaternal complications only in women in the lowest quartile for plasma zinc (Mukherjee et al., 1984).

A double blind trial was conducted on pregnant women to determine the effects zinc supplementation during pregnancy on maternal and fetal outcome. 494 women booking before 20 week of gestation in a hospital were prescribed either 66 mg zinc sulphate (equivalent to 20 mg elemental zinc, 0.3 mg zinc/kgbw/day) capsules or placebo for once daily use, starting from day of booking till delivery. Various adverse outcomes were tested, including maternal bleeding, hypertension, complications of labour and delivery, gestational age, Apgar scores, and neonatal abnormalities. The main outcome measure was birth weight. There were no differences between the mothers and neonates of the zinc supplemented and placebo group. Under the test conditions, zinc supplementation during pregnancy did not affect maternal or fetal outcome (Mahomed et al.,1989).

In summary, in studies with women receiving zinc supplementation during pregnancies at levels of approximately ≤ 0.3 mg Zn/kg bw/day, no reproductive or developmental effects were observed (WHO, 2001; SCF, 2003). Evidence of zinc toxicity during human pregnancy has not been reported, but this may be due to the fact that very high exposures to zinc in human pregnancy are unusual. In contrast, zinc is necessary for normal growth and development (e.g., gene expression, vitamin metabolism) and therefore it is not surprising that zinc deficiency during pregnancy can cause a variety of adverse effects to the foetus or may result in reduced fertility or delayed sexual maturation in animals as well as in humans (EU RAR, 2004; WHO, 2001).

Justification for classification or non-classification

There is no experimental evidence that would justify a classification of zinc compounds for hazardous effects for reproductive or developmental toxicity according under the Dangerous Substance Directive 67/548/EEC or Regulation (EC) 1272-2008 on the classification, labelling and packaging of substances and mixtures. The available reproductive and developmental toxicity information has been exclusively generated with soluble zinc compounds zinc chloride or zinc sulphate which ensure maximum bioavailable concentration of zinc and hence, allow the use of the information also for the assessment of the slightly soluble zinc compounds and insoluble zinc metal on a read across basis. No experimental fertility data were identified for these compounds.