Registration Dossier

Administrative data

Key value for chemical safety assessment

Effects on fertility

Description of key information

Systemic toxicity of zinc lactate can be understood in terms of the systemic toxicity of lactic acid and zinc(II). Lactic acid is not reprotoxic. According to the zinc chloride dossier, no effects on fertility and reproduction were observed for zinc(II) in the absence of maternal effects. In a two-generation study with zinc chloride the NOAEL for reproduction toxicity was 15 mg/kg bw/d, which corresponds to 26.6 mg/kg bw/d zinc lactate.

Link to relevant study records
Reference
Endpoint:
two-generation reproductive toxicity
Type of information:
experimental study
Adequacy of study:
key study
Study period:
Not reported
Reliability:
2 (reliable with restrictions)
Rationale for reliability incl. deficiencies:
study well documented, meets generally accepted scientific principles, acceptable for assessment
Qualifier:
equivalent or similar to
Guideline:
OECD Guideline 416 (Two-Generation Reproduction Toxicity Study)
Principles of method if other than guideline:
Not applicable
GLP compliance:
no
Limit test:
no
Species:
rat
Strain:
Sprague-Dawley
Sex:
male/female
Details on test animals and environmental conditions:
TEST ANIMALS
- Source: Harlan Sprague-Dawley Breeding Laboratories, Harlan Sprague-Dawley, Inc., Indianapolis, IN, USA
- Age at study initiation: 30-35 d
- Housing: Polycarbonate cages with stainless-steel wire lids
- Diet: Rodent chow (Lab Diet, Richmond Standard, PMI Feeds, Inc., St. Louis, MO), ad libitum
- Water: Deionized water, ad libitum
- Acclimation period: 2 week

ENVIRONMENTAL CONDITIONS
- Temperature: 21.1 to 25.5 °C
- Humidity: 50-55 %
- Air changes: 1/10 min
- Photoperiod : 12 h light/12 h dark cycle


Route of administration:
oral: gavage
Vehicle:
water
Details on exposure:
PREPARATION OF DOSING SOLUTIONS: 97% ZnCl2 was dissolved in milli-Q water.


Details on mating procedure:
- Length of cohabitation: 21 d
- Proof of pregnancy: Conception (day 0 of gestation)was checked daily in the mornings by looking for the presence or absence of copulatory plugs.
Analytical verification of doses or concentrations:
no
Details on analytical verification of doses or concentrations:
Not applicable
Duration of treatment / exposure:
2 generations
Frequency of treatment:
7 d/wk
Details on study schedule:
Dosing (7 days/week) started after two weeks of acclimation and was continued for males and females for 77 days prior to cohabitation. Dosing was continued throughout the periods of cohabitation (21 days) for both sexes. Dosing of female rats was continued throughout the gestation (21 days) and lactation (21 days) periods.
The doses for both sexes were adjusted weekly according to
changes in body weight.
Dose / conc.:
7.5 mg/kg bw/day (nominal)
Dose / conc.:
15 mg/kg bw/day (nominal)
Dose / conc.:
30 mg/kg bw/day (nominal)
No. of animals per sex per dose:
25
Control animals:
yes, concurrent vehicle
Details on study design:
- Dose selection rationale: The dosage levels were derived from a 14-day dose range finding study. The maximum tolerated dose (MTD) of ZnCl2 was set at 60 mg/kg/day in rats. In order to prevent a large effect of zinc-induced toxicity on non-reproductive tissues interfering with the interpretation of pure reproductive toxicity, the high-dose group (group 4) was set at 1/2 (30.00 mg of ZnCl2/kg bw/d) of the established MTD. Likewise, the middose group (group 3) was at 1/4 (15.00 mg of ZnCl2/kg of bw/d) of the established MTD and the lowest dose group (group 2) was 1/8 (7.50 mg of ZnCl2/kg bw/d) of the established MTD.
Positive control:
No data
Parental animals: Observations and examinations:
CAGE SIDE OBSERVATIONS: Yes
- Time schedule: Daily


DETAILED CLINICAL OBSERVATIONS: Yes


BODY WEIGHT: Yes


OTHER:
Hematology and clinical chemistry: Prior to necropsy, the Fo males were anesthetized with a combination of intraperitoneal Pentothal and Isoflo via inhalation. While the male rats were still under anesthesia, blood samples for hematology and clinical chemistries were collected in heparinised 3mL syringes via cardiac puncture. Following sample collection and while still under anesthesia, the animals were exsanguinated and necropsied. All plasma samples were analysed for alanine aminotransferase (ALT), aspartate aminotransferase (AST), alkaline phosphatase (ALK), amylase (Amyl), blood urea nitrogen (BUN), creatinine (Crea), cholesterol (Chol), sodium (Na), potassium (K), chloride (Cl), calcium (Ca), phosphorus (Phos), albumin (ALB), total protein (TP), total bilirubin (Tbil), and glucose (Glu) using Roche Cobas Mira S Chemistry Analyser (Roche Diagnostic System, Inc., Somerville, NJ).

Oestrous cyclicity (parental animals):
No data
Sperm parameters (parental animals):
No data
Litter observations:
STANDARDISATION OF LITTERS
- Performed on day 4 postpartum: yes
- Maximum of 8 pups/litter (4sex/litter); excess pups were killed and discarded.


PARAMETERS EXAMINED
The following parameters were examined in [F1 / F2 / F3] offspring: Total litter size, number of stillborn pups per sex, sex distribution, pup body weight and the presence of any obvious external congenital anomalies


GROSS EXAMINATION OF DEAD PUPS:
No
Postmortem examinations (parental animals):
SACRIFICE
- Male animals: All surviving animals, as soon as possible after the last litters in each generation were produced
- Maternal animals: All surviving animals, after the last litter of each generation were weaned


HISTOPATHOLOGY / ORGAN WEIGHTS:

Organ weights: During the necropsy, organ weights were recorded for the kidneys, liver, brain, pituitary, adrenals, pancreas, thymus, spleen, testes, epididymides, prostate, and seminal vesicles. Fo male organ weights were also adjusted to body weight for statistical analysis.

Histopathology: Tissue samples collected from organs listed above for histopathologic evaluation were fixed in either Bouins solution (all reproductive tissues) or 10% neutral buffered formalin (all other tissues). After fixation, the tissue samples were trimmed, processed, embedded in paraffin, cut at 6 μm and stained with hematoxylin and eosin.
Postmortem examinations (offspring):
At the end of cohabitation for the parental F1 males and lactation for the F1 females, the animals were anesthesized, sacrificed and their organ weights
were recorded like their Fo parents.
Statistics:
- Kruskal-Wallis test followed by the Mann-Whitney U test for pair-wise comparisons to detect the difference between treatment group and control means
- ANOVA for analysing body-weight change, fertility, litter size, pups’ viability, pups’ body weight, postpartum dam weight and organ weight data between different treatment groups
- Dunnett’s and/or Duncan’s multiple comparison procedures
Reproductive indices:
The reproductive parameters were expressed in terms of indices, weights, ratios and efficiencies that considered all stages from conception to weaning. The parameters were:
- Fertility index (%) = (number of females delivering/number of females cohabited) × 100
- Live birth index (%) = (number of live pups at Day 0/number of pups born) × 100
- 4-d survival index (%) = (number of live pups on Day 4/number of pups alive on day 0) × 100
- Body weights of pups = the body weight of pups were recorded on days 0, 4, 7, 14 and 21
- Sex ratio (%) = (the total number of males on the day of weaning)/ (the total number of females on the day of weaning) × 100
- Food efficiency = (body weight gain/amount of diet consumed) × 100
Offspring viability indices:
- 21-d (weaning) survival index (%) = (number of pups alive on Day 21/number of pups alive on Day 4) × 100
- Litter Size = Number of pups/number of pregnant females
Clinical signs:
effects observed, treatment-related
Description (incidence and severity):
see "Details on results (P0)"
Body weight and weight changes:
effects observed, treatment-related
Food consumption and compound intake (if feeding study):
effects observed, treatment-related
Organ weight findings including organ / body weight ratios:
effects observed, treatment-related
Histopathological findings: non-neoplastic:
effects observed, treatment-related
Other effects:
not examined
Reproductive function: oestrous cycle:
not examined
Reproductive function: sperm measures:
not examined
Reproductive performance:
effects observed, treatment-related
CLINICAL SIGNS AND MORTALITY (PARENTAL ANIMALS): Aggression/hyperactivity throughout the study in both males and females, hair loss behind the ears in males, vaginal discharges in low and high dose females; 0-20 and 12-24 % mortality in males and females respectively.

BODY WEIGHT AND FOOD CONSUMPTION (PARENTAL ANIMALS): All ZnCl2-treated F0 males experienced significant reduction in body weight after the 1st week of dosing and this trend continued up to the end of the experiment. The total weight gain of males was significantly reduced (dose dependent) in the low-, mid- and high-dose groups. The males experienced 0, 8, 20, and 12% mortality in control, low-, mid- and high-dose groups, respectively. In the F0 females, total weight gain and percent reduction in the low- mid- and high-dose groups were not significantly different from the control.

HEMATOLOGY AND CLINICAL CHEMISTRY: None of the hemogram or leukogram values of both Fo and F1 males and females among the ZnCl2-treated groups were different from those of the control groups. However, there was a trend toward decreased values of Packed Cell Volume (PCV). The clinical chemistry findings in males and females of both generations did not show any significant difference from those of their controls. However, in mid- and high-dose males of both generations, there seemed to a trend toward elevated values of Amyl, ALK, and GLu.

REPRODUCTIVE PERFORMANCE (PARENTAL ANIMALS): In F0 rats, ZnCl2 treatment caused a significant reduction on the fertility, litter size, and the viability indices (Days 0 and 4) were significantly reduced at the high-dose group compared to control.

ORGAN WEIGHTS (PARENTAL ANIMALS): In F0 males, the unadjusted weights of the brain in the mid and high-dose groups, the liver and kidney in all ZnCl2-treated groups, the spleen in the high-dose group, and the seminal vesicles in the mid- and high-dose groups were significantly different from the control. When organ weights of F0 males were adjusted for body weight, the brain in the mid- and high-dose groups, the liver and kidney in all ZnCl2-treated groups, the spleen in the high-dose group, and the seminal vesicles in the mid- and high-dose groups remained significantly different from their controls. The unadjusted organ weights of F0 females revealed significant differences for the spleen and uterus in the high-dose group. Following the adjustments of F0 female organ weights for body weight, the spleen and the uterus in the high-dose group remained significantly different from their controls.

GROSS PATHOLOGY (PARENTAL ANIMALS): Gross findings related to ZnCl2-treatment in males were primarily seen in the target organ systems (digestive, hematopoietic-lymphoreticular, and reproductive) already established for zinc. Digestive system lesions in the gastrointestinal tract (GIT) (distention, discoloration/hemorrhage and ulceration) and pancreas (smaller than usual) were mostly seen in rats given the two highest doses. Hematopoietic-lymphoreticular system lesions (small spleens and thymuses) were also scattered among the groups of ZnCl2-treated males. In the reproductive tract of the males, the only gross changes noted were small prostates and small seminal vesicles (one each) in the high-dose group. Gross lesions in ZnCl2-treated females generally paralleled those observed in their male counterparts.

HISTOPATHOLOGY (PARENTAL ANIMALS): In males, the most biologically meaningful lesions were found in the reproductive system (prostatic acinar atrophy and inflammation) and the hematopoietic-lymphoreticular system (splenic lymphoid depletion and hemosiderosis and thymic atrophy) of ZnCl2-treated groups. No significant changes in clinical pathology values or organ weights correlated with these lesions. None of the microscopic changes in target organs were of great magnitude. All unscheduled deaths were confined to the ZnCl2-treated groups, the majority of them probably being related to toxicity, but histomorphologic confirmation of this was not noted. The histopathology observed among the ZnCl2-treated females was similar to that seen in the males, except that no lesions were seen in the reproductive system. The correlations and biological interpretations were also very similar.

OTHER FINDINGS (PARENTAL ANIMALS): Postpartum dam body weight: The F0 and F1 post-partum dam weights in all dose groups were significantly different from their control groups.
Dose descriptor:
NOAEL
Effect level:
7.5 mg/kg bw/day (nominal)
Based on:
test mat.
Sex:
male/female
Basis for effect level:
other: overall toxicity
Dose descriptor:
NOAEL
Effect level:
15 mg/kg bw/day (nominal)
Sex:
male/female
Basis for effect level:
other: NOAEL for fertility and developmental toxicity
Clinical signs:
effects observed, treatment-related
Mortality / viability:
mortality observed, treatment-related
Body weight and weight changes:
effects observed, treatment-related
Sexual maturation:
not examined
Organ weight findings including organ / body weight ratios:
effects observed, treatment-related
Gross pathological findings:
effects observed, treatment-related
Histopathological findings:
effects observed, treatment-related
VIABILITY (OFFSPRING): The F1 males in the mid- and high-dose groups experienced a significant reduction in body weight after the 1st week of dosing and the low-dose group experienced a similar reduction after the 2nd week of dosing. These trends continued up to the end of the experiment. The total weight gain of F1 males was significantly reduced (dose dependent) in the low, mid-, and high-dose groups. The males experienced 0, 12, 8, and 4% mortality in the control, low-, mid- and high-dose groups, respectively. The mortality among the F1 females was 0, 8, 12, and 20% in the control, low-, mid- and high-dose groups, respectively.

CLINICAL SIGNS (OFFSPRING): Aggression/hyperactivity was observed throughout the study in both F1 males and females of ZnCl2-treated groups.

BODY WEIGHT (OFFSPRING): The body weights of F1 and F2 pups at Day 21 in the high-dose group were significantly lower compared to their control.

ORGAN WEIGHTS (OFFSPRING): In F1 males, the unadjusted weights of the brain, spleen, and prostate in all ZnCl2-treated groups, the liver, adrenal, testis and seminal vesicles in mid-dose and the kidney in high-dose were significantly different from their controls. When the organ weights of F1 males were adjusted for body weight, the brain, spleen, and prostate in all ZnCl2-treated groups, the liver, adrenal and seminal vesicles in mid-dose group, and kidney in high-dose group remained significantly different from their controls. The unadjusted organ weights of F1 females that were different from their controls included the brain and spleen in low- mid- and high-dose groups and the kidneys in the high-dose group. Following the adjustments of F1 female organ weights for body weight, the brain and spleen in all dose groups and kidneys in high dose groups were significantly different from controls.

GROSS PATHOLOGY (OFFSPRING): Gross findings related to ZnCl2-treatment in males were primarily seen in the target organ systems (digestive, hematopoietic-lymphoreticular, and reproductive) already established for zinc. Digestive system lesions in the gastrointestinal tract (GIT) (distention, discoloration/hemorrhage and ulceration) and pancreas (smaller than usual) were mostly seen in rats given the two highest doses. Hematopoietic-lymphoreticular system lesions (small spleens and thymuses) were also scattered among the groups of ZnCl2-treated males. In the reproductive tract of the males, the only gross changes noted were small prostates and small seminal vesicles (one each) in the high-dose group. Gross lesions in ZnCl2-treated females generally paralleled those observed in their male counterparts.

HISTOPATHOLOGY (OFFSPRING): In males, the most biologically meaningful lesions were found in the reproductive system (prostatic acinar atrophy and inflammation) and the hematopoietic-lymphoreticular system (splenic lymphoid depletion and hemosiderosis and thymic atrophy) of 30.00 mg/kg/day ZnCl2-treated groups. These results indicated that ZnCl2 exposure has only mild effects on the reproductive performance of rats. No significant changes in clinical pathology values or organ weights correlated with these lesions. None of the microscopic changes in target organs were of great magnitude. All unscheduled deaths were confined to the ZnCl2-treated groups, the majority of them probably being related to toxicity, but histomorphologic confirmation of this was not noted. The histopathology observed among the ZnCl2-treated females was similar to that seen in the males, except that no lesions were seen in the reproductive system. The correlations and biological interpretations were also very similar.

OTHER FINDINGS (OFFSPRING): Reproductive performance: F1: No significant difference was seen in the weaning index and sex ratios in F1 pups. In F1 generation rats, ZnCl2 treatment resulted in a significant reduction on fertility, viability (Day 0) and litter size in the high-dose group compared to control. However, ZnCl2 treatment showed no effect on viability index, weaning index and sex ratios of F2 pups.
Dose descriptor:
NOAEL
Generation:
F1
Effect level:
7.5 mg/kg bw/day (nominal)
Sex:
male/female
Basis for effect level:
other: overall effects
Dose descriptor:
NOAEL
Generation:
F1
Effect level:
15 mg/kg bw/day (nominal)
Based on:
test mat.
Sex:
male/female
Basis for effect level:
other: NOAEL for fertility and developmental toxicity
Reproductive effects observed:
not specified

None

Conclusions:
Under the test conditions, administration of test material to adult male and female rats throughout maturation, mating, gestation and early lactation resulted in significant effects on adults and offspring at 30 and 15 mg/kg/d. Although effects were seen at 7.5 mg/kg/d, these were considered to be toxicologically non significant and is therefore considered to be the "No Observed Adverse Effect Level" (NOAEL).
Executive summary:

A study was conducted to evaluate the reproductive toxicity potential of the test material in rats for two generations. Male and female rats were administered the test material at the doses of 7.50, 15.00 and 30.00 mg/kg/d over two successive generations. Control group animals received deionised water. Exposure of F0 and F1 parental rats to test material showed significant reduction in fertility, viability (days 0 and 4), and the body weight of F1 and F2 pups from the high-dose group but caused no effects on litter size, weaning index, and sex ratio. Significant reduction in body weights of F0 and F1 parental males and postpartum dam weights female rats. Exposure of test material to F0 and F1 generation parental animals resulted in non significant change in clinical pathology parameters (except the ALK level). Reduction of brain, liver, kidney, spleen and seminal vesicles weights of males and in the spleen and uterus of females was observed in F0 and F1 rats. Gross lesions were observed in gastro-intestinal (GI) tract, lymphoreticular/ hematopoietic and reproductive tract in parental rats in both generations. Reduced body fat was also recorded in F1 parental rats. Under the test conditions, administration of test material to adult male and female rats throughout maturation, mating, gestation and early lactation resulted in significant effects on adults and offspring at 30 and 15 mg/kg/d. Although effects were seen at 7.5 mg/kg/d, these were considered to be toxicologically non significant and is therefore considered to be the "No Observed Adverse Effect Level" (NOAEL).

Effect on fertility: via oral route
Endpoint conclusion:
no adverse effect observed
Dose descriptor:
NOAEL
15 mg/kg bw/day
Study duration:
subchronic
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

Lactic acid is a major and essential element in mammalian primary metabolism, and a ubiquitous ingredient in all kinds of food. Reproductive toxicity is not a relevant endpoint for such a substance since there is no possibility of lowering exposure below the minimum required levels or normal (or even abnormal) internal levels.

Effects on developmental toxicity

Description of key information

Systemic toxicity of zinc lactate can be understood in terms of the systemic toxicity of lactic acid and zinc(II). Lactic acid is a major and essential element in mammalian primary metabolism, and a ubiquitous ingredient in all kinds of food. Developmental toxicity is not a relevant endpoint for such a substance since there is no possibility of lowering exposure below the minimum required levels or normal (or even abnormal) internal levels.

To assess the toxicity of zinc(II) several developmental toxicity studies are available on the read-across substance zinc sulphate, which were already subject to peer-review in the EU risk assessment report on zinc sulphate (2004). Developmental toxicity studies, according to study design similar to OECD 414 with mice, rats, hamsters and rabbits were described. In these studies neither reproductive nor developmetal or maternal effects were observed, not even at the highest dose.

Lactic acid is a major and essential element in mammalian primary metabolism, and a ubiquitous ingredient in all kinds of food. Reproduction toxicity is not a relevant endpoint for such a substance since there is no possibility of lowering exposure below the minimum required levels or normal (or even abnormal) internal levels.

Systemic toxicity of zinc lactate can be understood in terms of the systemic toxicity of lactic acid and zinc(II). Lactic acid is not teratogenic. The NOAEL of zinc(II) is in accordance with the EU risk assessment report on zinc sulphate. The report stated that, when it is assumed (worst case) that the heptahydrate was administered from the study with hamsters it can be calculated that the NOAEL for both maternal and and developmental effects is 20 mg Zn(II)/kg bw/d. This corresponds to 74.5 mg/kg bw/d zinc lactate.

Link to relevant study records
Reference
Endpoint:
developmental toxicity
Type of information:
migrated information: read-across based on grouping of substances (category approach)
Adequacy of study:
weight of evidence
Study period:
Not reported
Reliability:
2 (reliable with restrictions)
Rationale for reliability incl. deficiencies:
other: Used in risk assessment report for zinc sulphate, acceptable for assessment.
Qualifier:
equivalent or similar to
Guideline:
OECD Guideline 414 (Prenatal Developmental Toxicity Study)
Principles of method if other than guideline:
Female hamsters were administered daily doses of 0.9, 4.1, 19 and 88 mg/kg bw by gavage during days 6-10 of gestation to evaluate the developmental toxicity of zinc sulphate.
GLP compliance:
not specified
Limit test:
no
Species:
hamster
Strain:
not specified
Route of administration:
oral: gavage
Vehicle:
not specified
Analytical verification of doses or concentrations:
no
Details on mating procedure:
No data
Duration of treatment / exposure:
Days 6-10 of gestation
Frequency of treatment:
Daily
Duration of test:
No data
Dose / conc.:
0.9 mg/kg bw/day
Remarks:
during days 6-15 of gestation
Dose / conc.:
4.1 mg/kg bw/day
Remarks:
during days 6-15 of gestation
Dose / conc.:
19 mg/kg bw/day
Remarks:
during days 6-15 of gestation
Dose / conc.:
88 mg/kg bw/day
Remarks:
during days 6-15 of gestation
No. of animals per sex per dose:
23-25 animals/group
Control animals:
yes
Dose descriptor:
NOAEL
Effect level:
88 mg/kg bw/day
Basis for effect level:
other: maternal toxicity
Details on embryotoxic / teratogenic effects:
Embryotoxic / teratogenic effects: no effects
Dose descriptor:
NOAEL
Effect level:
88 mg/kg bw/day
Basis for effect level:
other: teratogenicity
Abnormalities:
not specified
Developmental effects observed:
not specified

- No clearly discernible effects on maternal survival, body weight gains, number of corpora lutea, implantations and resorptions were observed.

- The number of live litters, live and dead foetuses, the foetus weights and sex ratio were not affected by treatment.

- No difference in number or kind of abnormalities (in either soft or skeletal tissues) was found between exposed and control groups.

Conclusions:
Under the conditions of the test, administration of up to 88 mg/kg bw/day of unspecified zinc sulphate (ca. 35.2 mg or 19.9 mg Zn2+/kg bw, for anhydrate and heptahydrate, respectively) had no adverse effects on adult hamsters and their foetuses.
Executive summary:

Female hamsters (23-25 animals/group; outbred strain of golden hamster) received daily doses of 0.9, 4.1, 19 and 88 mg unspecified ZnSO4/kg bw by gavage during days 6-10 of gestation. A control group was included. All animals were observed daily for appearance and behaviour with particular attention to food consumption and body weight. Body weights were recorded on day 0, 8, 10 and 14 of gestation. The females were sacrificed at day 14. The urogenital tract of each animal was examined in detail. Between 21 and 24 females were pregnant at term in all groups. No clearly discernible effects on maternal survival, body weight gains, number of corpora lutea, implantations and resorptions were observed. The number of live litters, live and dead foetuses, the foetus weights and sex ratio were not affected by treatment. No difference in number or kind of abnormalities (in either soft or skeletal tissues) was found between exposed and control groups. Under the conditions of the test, administration of up to 88 mg/kg bw/day of unspecified zinc sulphate (ca. 35.2 mg or 19.9 mg Zn2+/kg bw, for anhydrate and heptahydrate, respectively) had no adverse effects on adult hamsters and their foetuses.

Effect on developmental toxicity: via oral route
Endpoint conclusion:
no adverse effect observed
Dose descriptor:
NOAEL
20 mg/kg bw/day
Study duration:
subacute
Species:
hamster
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

The chosen NOAEL is in accordance with the EU risk assessment report on zinc sulphate (2004) of 20 mg Zn(II)/kg bw/d. This corresponds to 74.5 mg/kg bw/d zinc lactate.

Justification for classification or non-classification

Based on the available data, the target substance does not warrant classification for reproductive toxicity.