Registration Dossier

Administrative data

Description of key information

Read- across: WoE, rat, OECD 422, GLP, NOEL = 300 mg/kg bw/d (Longobardi 2013)

Read-across: WoE, rat, OECD 407, GLP, NOAEL = 1000 mg/kg bw/d (Müller 2000)

Read-across: WoE, rat and mice, 13 week feeding study, NOAEL = 322.57 mg/kg bw/d (lowest value, male rat) (Maita 1981)

Key value for chemical safety assessment

Toxic effect type:
dose-dependent

Repeated dose toxicity: via oral route - systemic effects

Link to relevant study records

Referenceopen allclose all

Endpoint:
short-term repeated dose toxicity: oral
Type of information:
read-across from supporting substance (structural analogue or surrogate)
Adequacy of study:
weight of evidence
Reason / purpose for cross-reference:
read-across source
Reason / purpose for cross-reference:
assessment report
Clinical signs:
effects observed, treatment-related
Description (incidence and severity):
Four high dose females (nos. 92510061, 92510067, 92510071 and 92510075) were sacrificed for humane reasons on Day 21/22 of gestation. At despatch to necropsy these animals had pale appearance, decreased activity, piloerection and/or were cold to touch. At necropsy examination they were all pregnant.
At post mortem examination, all females showed pale colour of the kidneys and reduced size of the spleen and thymus. In addition some females also showed multiple depressed dark areas in the glandular region of the stomach with associated yellow mucoid contents in the ileum and jejunum and/or distended duodenum with gas contents and/or pale colour of the liver.
Such changes were considered to be treatment-related.
Treatment-related changes were noted in the kidneys, spleen, thymus and stomach. The observed renal injury involved the nephrons, unit of the kidney, morphologically characterised by degeneration of some glomeruli (sclerosis) and the majority of renal tubules, sometimes associated with subacute inflammatory reactions. The renal tubules showed lined cells, in some instances basophilic with presence of exfoliated cells and/or amorphous material (crystal-like) in their lumen. Focal tubular necrosis in the papilla, associated with pelvic dilatation (hydronephrosis) and hyperplasia of the pelvic lining epithelium were also reported in female no. 92510075.
Moreover, mild to marked lymphoid depletion or red pulp depletion and atrophy were noted in the spleen and in the thymus and ulceration or erosion in the glandular mucosa of the stomach was also reported in the four unscheduled dead females. These changes were considered stress-related.
All females mated. Only one control female (no. 92510005) was sacrificed 26 days post coitum and found not pregnant at necropsy.
The number of females with live pups on Day 4 postpartum was: 8 in the control group, 10 in each of the low and mid-dose groups, 6 in the high dose group. Total litter loss was observed in a single female from the control group (no. 92510019).
Male animals did not show any signs of toxicological significance during the whole study. No significant clinical signs were seen in the female animals killed at the end of the study.
Mortality:
mortality observed, non-treatment-related
Description (incidence):
Four females dosed at 1000 mg/kg/day were sacrificed for humane reasons on Day 21/22 of gestation due to the presence of a number of clinical signs (pale appearance, decreased activity, piloerection and/or cold to touch).
Body weight and weight changes:
no effects observed
Description (incidence and severity):
No significant changes in body weight were observed in the treated animals when compared to controls. Reductions in body weight and body weight gain (these being also statistically significant) were observed in the treated males during the mating treatment period. These reductions, which were not dose-related, were not considered to be toxicologically significant. No significant changes were observed in the females.
Food consumption and compound intake (if feeding study):
no effects observed
Description (incidence and severity):
No effects on food consumption were observed in the males. Slight and occasionally also significant at statistical analysis reductions were seen in the high dose females on Day 20 post coitum (-13%) and on Day 4 post partum (-19%).
Food efficiency:
not examined
Water consumption and compound intake (if drinking water study):
not examined
Ophthalmological findings:
not examined
Haematological findings:
not examined
Clinical biochemistry findings:
not examined
Urinalysis findings:
not examined
Behaviour (functional findings):
no effects observed
Description (incidence and severity):
Four high dose females (nos. 92510061, 92510067, 92510071 and 92510075) were sacrificed for humane reasons on Day 21/22 of gestation. At despatch to necropsy these animals had pale appearance, decreased activity, piloerection and/or were cold to touch.
Immunological findings:
not examined
Organ weight findings including organ / body weight ratios:
no effects observed
Description (incidence and severity):
Terminal body weight as well as absolute and relative weight of testes, epididymides and ovaries were similar between groups.
Gross pathological findings:
no effects observed
Description (incidence and severity):
Macroscopic observations:
No relevant changes were detected at post mortem examination in treated animals, when compared with controls.
Neuropathological findings:
not examined
Histopathological findings: non-neoplastic:
no effects observed
Description (incidence and severity):
No treatment-related changes were seen in selected organs/tissues evaluated in high dose males or females receiving the test substance, sacrificed at the end of treatment period, nor in the abnormalities detected at post mortem.
Seminiferous tubules were evaluated with respect to their stage in the spermatogenic cycle and to the integrity of the various cell types within the different stages; regular layering in the germinal epithelium was noted.
Histopathological findings: neoplastic:
no effects observed
Details on results:
Reproductive parameters: see IUCLID chapter 7.8
Dose descriptor:
NOEL
Effect level:
300 mg/kg bw/day (nominal)
Based on:
test mat.
Sex:
male/female
Basis for effect level:
clinical signs
Remarks on result:
other: systemic toxicity
Critical effects observed:
no
Conclusions:
Treatment-related effects indicating systemic toxicity were observed in some pregnant females dosed at 1000 mg/kg/day.
No changes were observed in males at any dose and in females dosed at 100 and 300 mg/kg/day.
No adverse effects on sexual function and fertility or in developmental parameters and lactation were observed at any of the dose levels investigated.
The dose level of 300 mg/kg/day is considered to be the NOEL for systemic toxicity, while 1000 mg/kg/day is the NOAEL for fertility and reproduction parameters.
Executive summary:

A study according to OECD Guideline 422 and GLP was performed to investigate the repeated dose toxicity of the test item when administered orally to male and female Sprague Dawley rats. Groups of 10 males and 10 females received the test item, by gavage at dosages of 100, 300 and 1000 mg/kg/day. A similarly constituted group of animals received the vehicle alone (purified water) and acted as a control. All doses were administered at a constant volume of 10 mL/kg body weight.

Males were treated for 2 weeks prior to pairing and during pairing of all females until the day before necropsy, for a total of 4 consecutive weeks. Females were treated for 2 weeks prior to pairing, during pairing and throughout the gestation and lactation periods until Day 3 post partum.

The following investigations were performed on all groups: body weight, clinical signs, food consumption, oestrous cycle, mating performance, litter observation, macroscopic observations, organ weights and histopathological examination of abnormalities. Histopathological examination of testes, epididymides and ovaries was performed only on control and high dose groups.

Mortality and fate of females

Four high dose females were sacrificed for humane reasons on Day 21/22 of gestation. At despatch to necropsy these animals had pale appearance, decreased activity, piloerection and/or were cold to touch. At necropsy examination they were all pregnant. Pale colour of the kidneys, reduced size of the spleen and thymus, multiple depressed dark areas in the glandular region of the stomach with associated yellow mucoid contents in the ileum and jejunum and/or distended duodenum with gas contents and/or pale colour of the liver were also observed.

Histopathological examination revealed treatment-related changes in the kidneys (nephrons, unit of the kidney, morphologically characterised by degeneration of some glomeruli and the majority of renal tubules, sometimes associated with subacute inflammatory reactions), spleen and thymus (lymphoid depletion or red pulp depletion and atrophy) and stomach (ulceration or erosion in the glandular mucosa). Only one control female was found not pregnant at necropsy.

The number of females with live pups on Day 4 post partum was: 8 in the control group, 10 in each of the low and mid-dose groups, 6 in the high dose group.

Clinical signs

No signs of toxicological significance were observed in animals from the control, low and mid-dose groups and in the surviving female animals from the high dose group.

Body weight and body weight gain

No changes of toxicological significance were observed in body weight and body weight gain in treated males and females when compared to controls.

Food consumption

No effects on food consumption were observed in the males and in low and mid-dose females. Reduced food consumption was seen in the high dose females during the post coitum period.

Oestrus cycle, mating performance and reproductive parameters

Mean oestrus cycle was slightly reduced in the females dosed at 1000 mg/kg/day (approximately 2 in the 15 day pre-mating period) when compared to controls (approximately 3).

No significant differences between groups were observed in pre-coital interval and copulation plugs, as well as in the reproductive parameters (copulatory index and fertility index).

Implantation, pre-birth loss data and gestation length

No treatment-related differences were noted in implantation number, pre-birth loss data and gestation length between control and treated groups.

Litter data and sex ratios

Litter data and sex ratios were unaffected by treatment.

Pre-weaning clinical signs of pups

Clinical signs were comparable between treated and control groups.

Necropsy findings in decedent pups and in pups sacrificed on Day 4 post partum No treatment-related effects were seen.

Organ weights

No toxicologically relevant changes were observed in organ weights.

Macroscopic observations

No relevant changes were detected at post mortem examination in treated animals killed at end of treatment period, when compared with controls.

Microscopic observations

No treatment-related changes were seen in selected organs/tissues evaluated in high dose males or females receiving the test item, sacrificed at the end of treatment period, or in the abnormalities detected at post mortem.

Seminiferous tubules were evaluated with respect to their stage in the spermatogenic cycle and to the integrity of the various cell types within the different stages; regular layering in the germinal epithelium was noted.

The source substance contains the major organic moieties of the target substance. Therefore, the results are also applicable for the target substance.

Endpoint:
short-term repeated dose toxicity: oral
Type of information:
read-across from supporting substance (structural analogue or surrogate)
Adequacy of study:
weight of evidence
Reason / purpose for cross-reference:
read-across source
Reason / purpose for cross-reference:
assessment report
Clinical signs:
no effects observed
Description (incidence and severity):
Only two clinical observations (slight diarrhoea and plough through bedding with nose) were registered.
The slight diarrhoea was observed only in one male control animal (No. 1) only on day 19, this sign is not to connect with the administration of the test article. The plough through bedding with nose was observed only briefly (approximately 5 minutes) immediately after the administration in all animals of the high dose group (all 10 male and all 10 female animals) only on days 13 until 16. These animals did not show this sign during the further administration or recovery period. For a tabular presentation of the onset and duration of the clinical observations was disclaimed in this study report because only these two temporary clinical signs were observed. None of the animals showed further alterations of their general state of well-being and behaviour.
Mortality:
no mortality observed
Description (incidence):
No mortality occurred during the study period.
Body weight and weight changes:
no effects observed
Description (incidence and severity):
Neither the body weights nor the body weight gain of the animals were influenced by administration of the test article. The few significant differences seem to be incidental, because the differences were not related to dose or time course.
Food consumption and compound intake (if feeding study):
no effects observed
Description (incidence and severity):
no influence by administration of test item
Food efficiency:
no effects observed
Water consumption and compound intake (if drinking water study):
not examined
Ophthalmological findings:
not examined
Haematological findings:
no effects observed
Description (incidence and severity):
The hemoglobin concentration and the packed cell volume were statistically significantly increased in the female animals of the high dose group. This was the reason for the determination of all hematological parameters in the satellite groups. In the satellite groups these parameters were not more increased. All these values are in the range of the of the historical control data in the testing facility for the strain used (confidence interval of the hemaglobin concentration in female control animals: 14.3 - 16.4 g/dl; of the packed cell volume in female control animals: 37.8 - 50.3 %).
Therefore it is deemed these changes are incidental and not caused by the test article administration.
None of the other haematological parameters investigated was affected by the administration of the test article.
Clinical biochemistry findings:
no effects observed
Description (incidence and severity):
The activity of aspartate arninotransferase (AST) was statistically significantly decreased in the high dose group of males and in the low and high dose group of females. Although there is no clear dose-dependence, this decrease is deemed to be caused by the test article adrninistration because all values in the treated animals are smaller than the control values, except of the middle dose group in female animals. A statistically significant decreased activity of aspartate e aminotransferase was also observed in the animals of satellite groups. The Change of activity of AST remains of uncertain toxicological relevance as liver cell damage usually is associated with an increase in enzyme levels. The Serum glucose level was increased in the male animals of all dose groups, statistically significantly in the middle dose group. In the satellite group this Parameter was not changed. However all values are in the range of the historical control data in the testing facility for the strain used (confidence interval of the fasting glucose level in male control animals: 4.0 - 8.8 mmol/l). It is to assume, that the control data were determined incidental to low. Therefore it is deemed these changes are incidental and not caused by the test article adrninistration. The statistically significant decreased Urea level in the female animals of the low dose group is also incidental (no dose dependence, confidence interval of the Urea level in female control animals: 3 5.6 - 60.1 mg/dl).

The sodium level was increased in female animals, statistically significantly in the high dose group. In the satellite group this parameter was not more increased. The potassium level was increased in male animals, statistically significantly in the middle and in the high dose group. A statistically significant increase of potassium level in male animals was also observed in the satellite group. However, the differences to the control data are only small in both cases and all the values are in the ranges of historical control data in the testing facility for the strain used (confidence interval of the sodium level in female control animals: 122.3 - 157.4 mmol/l; confidence interval of the potassium level in male control animals: 3.6 - 8.1 mmol/I). Thereforeit seems also to be that these changes are incidental and not caused by the test article administration. None of the other clinical-biochemical parameters investigated was affected by the administration of the test article.
Urinalysis findings:
not specified
Behaviour (functional findings):
no effects observed
Description (incidence and severity):
The examination of sensory response, grip strength or motor activity did not show any alterations prior to the administration of the test article or at the end of the study.
Immunological findings:
not examined
Organ weight findings including organ / body weight ratios:
no effects observed
Description (incidence and severity):
Organ weight determinations revealed some statistically significant decreased or increased values of the absolute and relative organ weight. These statistically significant changes were in all cases in the range of the historical control data in the testing facility for the strain used.
Gross pathological findings:
no effects observed
Description (incidence and severity):
No test substance-related macroscopic and histological findings were observed.
Neuropathological findings:
not examined
Histopathological findings: non-neoplastic:
no effects observed
Histopathological findings: neoplastic:
no effects observed
Dose descriptor:
NOAEL
Effect level:
1 000 mg/kg bw/day (nominal)
Based on:
test mat.
Sex:
male/female
Basis for effect level:
body weight and weight gain
clinical biochemistry
clinical signs
gross pathology
haematology
histopathology: non-neoplastic
mortality
organ weights and organ / body weight ratios
Dose descriptor:
NOEL
Effect level:
1 000 mg/kg bw/day (nominal)
Based on:
test mat.
Sex:
male/female
Basis for effect level:
behaviour (functional findings)
body weight and weight gain
clinical biochemistry
clinical signs
gross pathology
haematology
histopathology: non-neoplastic
mortality
organ weights and organ / body weight ratios
Critical effects observed:
not specified

A part of the haematological and clinical-biochemical parameters and of the absolute and relative weights of organs showed statistically significant effects, which are however in the ranges of historical control data in the testing facility for the strain used and therefore they were evaluated as incidental and not caused by the test article administration. It is conspicuous

that these incidental statistically significant effects occur in the high dose group in the most cases. It is to assume that this dose (1000 mg/kg b.w.) is the limit dose for the repeated administration of the test article without damage.

The following effects can be deemed as caused by the test article administration:

• The briefly plough through bedding with nose of the animals of the high dose group in a short period in the middle of the whole administration period as a sign of a temporary not general well-being.

• The activity of aspartate aminotransferase was statistically significantly decreased in the high dose group of males and in the low and high dose group of females. This effect was irreversible in the course of the recovery period. This changed enzyme activity remains of uncertain toxicological relevance as liver cell damage usually is associated with an increase in enzyme levels

Conclusions:
The daily oral administration of the test item at doses of 100, 300 and 1000 mg/kg body weight to rats for a period of 28 days was tolerated without any marked effects on physical condition, even at the high dose of 1000 mglkg body weight (maximum limit dose for repeated studies).
Executive summary:

A study according to OECD Guideline 407 and GLP was performed to provide information on the toxicity and reversibility of possible effects of the test article when administered orally (by gavage) to the rat, once daily for 28 days with a treatment-free recovery period of further 14 days.

The daily oral administration of the test article at doses of 100, 300 and 1000 mg/kg body weight to rats for a period of 28 days was tolerated without any marked effects on physical condition.

None of the animals died during the course of investigation.

The following effects can be deemed as caused by the test article administration:

• The briefly plough through bedding with nose of the animals of the high dose group in a short period in the middle of the whole administration period as a sign of a temporary not general well-being.

• The activity of aspartateaminotransferase was statistically significantly decreased in the . high dose group of males and in the low and high dose group of females. This effect wasirreversible in the course of the recovery period. This changed enzyme activity remains of uncertain toxicological relevance as liver cell damage usually is associated with an increase in enzyme levels.

The body weight, the food consumption, and the coagulation parameters were not influenced by administration of the test article. No substance-dependent pathological macroscopic and histological findings were observed.

A part of the haematological and clinical-biochemical parameters and of the absolute and relative weights of organs showed statistically significant effects, which arehowever in the ranges of historical control data in the testing facility for the strain used and therefore they were evaluated as incidental and not caused by the test article administration. It is conspicuous that these incidental statistically significant effects occur in the high dose group in· the most cases.

It is to assume that this dose (1000 mg/kg b.w.) is the limit dose for the repeated administration of the test article without damage.

Based on the findings the dose of 1000 mglkg body weight / day (dose of the high dose group) can be declared as the NOAEL (No Observed Adverse Effect Level).

The results of this study do not allow the determination of a target organ.

The source substance contains the major organic moieties of the target substance. Therefore, the NOAEL of 1000 mg/kg body weight/day is also applicable for the target substance.

Endpoint:
sub-chronic toxicity: oral
Type of information:
read-across from supporting substance (structural analogue or surrogate)
Adequacy of study:
weight of evidence
Reason / purpose for cross-reference:
read-across source
Reason / purpose for cross-reference:
assessment report
Clinical signs:
effects observed, treatment-related
Description (incidence and severity):
In mice, dead or moribund animals in the 30000 ppm group showed depressed spontaneous motility and unkempt fur a little while before death or sacrifice.
Mortality:
mortality observed, treatment-related
Description (incidence):
In mice, four males and one female in the 30000 ppm group were found dead or killed in extremis during the study. Mortality was 33.3% in males and 8.3% in females
Body weight and weight changes:
effects observed, treatment-related
Description (incidence and severity):
Mice of both sexes in the 30000 ppm group showed a more prominently retarded growth resulting in smaller body size than did those of other groups. A significant but very slight depression of weight gain was seen in females of the 300 ppm group for a week after commencement, followed by a rapid recovery to the control level.
In rats of the 30000 ppm group a depressed weight gain and dwarfism similar to that observed in mice was seen in males. Weight gain of females in this group was slightly depressed during the study with significant differences to control animals in the 1st to 5th weeks of the study.
Food consumption and compound intake (if feeding study):
effects observed, treatment-related
Description (incidence and severity):
The food intake of male and female mice in the 30000 ppm group was depressed during the first week of the study in comparison to that of the controls but showed a tendency to recover afterwards. Average intake of these animals then remained at only a slightly lower level than that of the control group. In rats, the food intake of males decreased after the third week of the study in the 30000 ppm group. A similar reduction was seen in females of this group during the 1st to 6th weeks but then disappeared. A slightly lower value of average food intake was disclosed in only males.
Food efficiency:
effects observed, treatment-related
Description (incidence and severity):
Food efficiencies of of male and female mice in the 30000 ppm group were markedly lower than those of the control group during the first week of the study, corresponding to the decrease in food intake. The overall average food efficiency in the 30000 ppm group was much lower than the control group. While there were some fluctuations in food efficiency of rats in each group, a slight reduction in overall average value was shown only in males of the 30000 ppm group.
Ophthalmological findings:
not examined
Haematological findings:
effects observed, treatment-related
Description (incidence and severity):
Male and female mice in the 30000 ppm group showed moderately lower values in hematocrit and hemoglobin concentration than those of the control group; the leukocyte count in males also decreased moderately.
In rats, a moderate reduction in leukocyte count was shown in both sexes in the 30000 ppm group; males also disclosed a slight decrease in hematocrit and hemoglobin concentration. There were no remarkable changes in animals in the less than 3000 ppm group, but there was a slight increment of hemoglobin concentration in females in the 3000 ppm group.
Clinical biochemistry findings:
effects observed, treatment-related
Description (incidence and severity):
Mice of both sexes in the 30000 ppm group showed a slight to moderate decrease in total protein, glucose and cholesterol and a moderate to marked increase in alkaline phosphatase and urea nitrogen. Additional significant changes occurring in these animals were depression of GPT level and increase in calcium level in females, and an increase of GOT level in males.
Significant reductions or reductive tendencies were seen in rats in the following parameters: GOT and GPT in all male chemically fed groups, total protein, cholesterol and calcium levels in males in the 30000 ppm group and calcium level in females in both the 3000 and 30000 ppm groups.
Urinalysis findings:
not examined
Behaviour (functional findings):
effects observed, treatment-related
Description (incidence and severity):
Male and female rats in the 30000 ppm group showed a symptom of discarding the diet from the food jar by picking it out with their fore-limbs. This symptom began a week after commencement of the experiment and persisted throughout the study.
Immunological findings:
not examined
Organ weight findings including organ / body weight ratios:
effects observed, treatment-related
Description (incidence and severity):
Organ weight analysis of the mice revealed that coincidental increase in absolute and relative weight was found in the thyroids of males and the kidneys of females in the 30,000 ppm group.
In the organ weights of the rats, a slight or moderate decrease in absolute and relative weights was seen in the liver and kidney among males in the 30000 ppm group.
Gross pathological findings:
effects observed, treatment-related
Description (incidence and severity):
In mice, marked emaciation, ischemic discoloration of the kidney and thyroid, strophy of the pancreas, edematous thickening of the upper small intestine and slight splenomegaly were recorded in the 30000 ppm group at necropsy, in addition to several cases of forestomach ulcer.
Neuropathological findings:
not examined
Histopathological findings: non-neoplastic:
effects observed, treatment-related
Description (incidence and severity):
Mice (30000 ppm): impairment of the urinary tract and regressive changes in the exocrine gland of the pancreas. Morphological changes of erythrocyte, anisocytosis, polychromatophilia and poikilocytosis, were seen in 6 males and 4 females which were reported by necropsy or microscopic observation to have fore-stomach ulcers. There were lesions attributable to the treatment in the pancreas, upper intestine, stomach, spleen and kidney from mice in the 30000 ppm group. The pancreatic acinar cells had swollen nuclei with an increased number of clarified nucleoii and whirl-like profiles in their cytoplasm which were more basophilically stained than the controls. Single cell necrosis of the acinar cells was also a common feature in these animals. Moreover, a decrease in the number of acinus and ductulelike metaplasia of acinar cells was demonstrated. There were mucosal catarrh in the upper intestine with proliferation of epithelial cells and edema at lamina propria, slight to moderate ulcerative lesions in the boundary of the fore-stomach and proliferation of erythropoietic immature cells in the splenic red pulp of these animals. Regressive changes of the renal cortex were observed in the females.
In rats, pancreatic lesions similar to those in mice were seen in the 30, 000 ppm group, as well as degeneration and necrosis of the acinar cells, clarification of centroacinar cells and interstitial fibrosis. No other lesions attributable to the treatment were found in rats.
Histopathological findings: neoplastic:
no effects observed
Dose descriptor:
NOEL
Remarks:
mice and rats
Effect level:
4 135 ppm
Based on:
test mat.
Sex:
male/female
Basis for effect level:
clinical biochemistry
gross pathology
haematology
histopathology: non-neoplastic
organ weights and organ / body weight ratios
Dose descriptor:
NOEL
Remarks:
mice
Effect level:
631.35 mg/kg bw/day (nominal)
Based on:
test mat.
Sex:
male
Basis for effect level:
clinical biochemistry
gross pathology
haematology
organ weights and organ / body weight ratios
Dose descriptor:
NOEL
Remarks:
mice
Effect level:
660.29 mg/kg bw/day (nominal)
Based on:
test mat.
Sex:
female
Basis for effect level:
clinical biochemistry
gross pathology
haematology
histopathology: non-neoplastic
organ weights and organ / body weight ratios
Dose descriptor:
NOEL
Remarks:
rats
Effect level:
322.57 mg/kg bw/day (nominal)
Based on:
test mat.
Sex:
male
Basis for effect level:
clinical biochemistry
gross pathology
haematology
histopathology: non-neoplastic
organ weights and organ / body weight ratios
Dose descriptor:
NOEL
Remarks:
rats
Effect level:
334.9 mg/kg bw/day (nominal)
Based on:
test mat.
Sex:
female
Basis for effect level:
clinical biochemistry
gross pathology
haematology
histopathology: non-neoplastic
organ weights and organ / body weight ratios
Critical effects observed:
no

The target substance contains 16.5 % Zn ions. Therefore, the hazard value for Zn ion is converted to the target substance considering a Zn content of 16.5 %.

NOEL (mice & rat, m/f) (ZnSO4.7H2O) = 3000 ppm

NOEL (mice & rat, m/f) (Zn; ZnSO4.7H2O*0.2274) = 682.35 ppm

NOEL (mice & rat, m/f) (target substance; Zn/0.165) = 4135.45 ppm

NOEL (mice; m) (ZnSO4.7H2O) = 458 mg/kg bw/day

NOEL (mice; m) (Zn; ZnSO4.7H2O*0.2274) = 104.17 mg/kg bw/day

NOEL (mice; m) (target substance; Zn/0.165) = 631.35 mg/kg bw/day

NOEL (mice; f) (ZnSO4.7H2O) = 479 mg/kg bw/day

NOEL (mice; f) (Zn; ZnSO4.7H2O*0.2274) = 108.95 mg/kg bw/day

NOEL (mice; f) (target substance; Zn/0.165) = 660.29 mg/kg bw/day

NOEL (rat; m) (ZnSO4.7H2O) = 234 mg/kg bw/day

NOEL (rat; m) (Zn; ZnSO4.7H2O*0.2274) = 53.22 mg/kg bw/day

NOEL (rat; m) (target substance; Zn/0.165) = 322.57 mg/kg bw/day

NOEL (rat; f) (ZnSO4.7H2O) = 243 mg/kg bw/day

NOEL (rat; f) (Zn; ZnSO4.7H2O*0.2274) = 55.26 mg/kg bw/day

NOEL (rat; f) (target substance; Zn/0.165) = 334.9 mg/kg bw/day

Conclusions:
The maximum no-effect level of the target substance was determined to be 4135.45 ppm, which is approximately equivalent to the following milligram doses: mice; male 631.35 mg/kg/day, female 660.29 mg/kg/day, rats; male 660.29 mg/kg/day, female 660.29 mg/kg/day.
Executive summary:

ICR mice and Wistar rats of both sexes were fed a diet containing ZnSO4 at 0, 300, 3000 and 30000 ppm for 13 weeks. Animals in the 30000 ppm group showed retarded growth along with low food intake, abnormal values in a few hematological parameters and regressive changes of the pancreatic exocrine gland. In addition, mice had decreased water intake and significant deviations in biochemical parameters, toxic lesions appeared in the stomach, intestine and spleen of both sexes and in the kidney of females. Four males and one female mouse were found dead or moribund during the study. The maximum no-effect level of ZnSO4 was determined to be 3000 ppm, which is approximately equivalent to the following milligram doses: mice: male 458 mg/kg/day, female 479 mg/kg/day, rats: male 234 mg/kg/day, female 243 mg/kg/day.

Considering the Zn concentration of max. 16.5 % in the target substance, the maximum no-effect level of the target substance was determined to be 4135.45 ppm, which is approximately equivalent to the following milligram doses: mice: male 631.35 mg/kg/day, female 660.29 mg/kg/day, rats: male 322.57 mg/kg/day, female 334.9 mg/kg/day.

Endpoint conclusion
Endpoint conclusion:
no adverse effect observed
Dose descriptor:
NOAEL
300 mg/kg bw/day
Study duration:
subacute
Species:
rat

Repeated dose toxicity: inhalation - systemic effects

Endpoint conclusion
Endpoint conclusion:
no study available

Repeated dose toxicity: inhalation - local effects

Endpoint conclusion
Endpoint conclusion:
no study available

Repeated dose toxicity: dermal - systemic effects

Endpoint conclusion
Endpoint conclusion:
no study available

Repeated dose toxicity: dermal - local effects

Endpoint conclusion
Endpoint conclusion:
no study available

Additional information

OECD 422 (Longobardi 2013)

A study according to OECD Guideline 422 and GLP was performed to investigate the repeated dose toxicity of the test item when administered orally to male and female Sprague Dawley rats. Groups of 10 males and 10 females received the test item, by gavage at dosages of 100, 300 and 1000 mg/kg/day. A similarly constituted group of animals received the vehicle alone (purified water) and acted as a control. All doses were administered at a constant volume of 10 mL/kg body weight.

Males were treated for 2 weeks prior to pairing and during pairing of all females until the day before necropsy, for a total of 4 consecutive weeks. Females were treated for 2 weeks prior to pairing, during pairing and throughout the gestation and lactation periods until Day 3 post partum.

The following investigations were performed on all groups: body weight, clinical signs, food consumption, oestrous cycle, mating performance, litter observation, macroscopic observations, organ weights and histopathological examination of abnormalities. Histopathological examination of testes, epididymides and ovaries was performed only on control and high dose groups.

Mortality and fate of females

Four high dose females were sacrificed for humane reasons on Day 21/22 of gestation. At despatch to necropsy these animals had pale appearance, decreased activity, piloerection and/or were cold to touch. At necropsy examination they were all pregnant. Pale colour of the kidneys, reduced size of the spleen and thymus, multiple depressed dark areas in the glandular region of the stomach with associated yellow mucoid contents in the ileum and jejunum and/or distended duodenum with gas contents and/or pale colour of the liver were also observed.

Histopathological examination revealed treatment-related changes in the kidneys (nephrons, unit of the kidney, morphologically characterised by degeneration of some glomeruli and the majority of renal tubules, sometimes associated with subacute inflammatory reactions), spleen and thymus (lymphoid depletion or red pulp depletion and atrophy) and stomach (ulceration or erosion in the glandular mucosa). Only one control female was found not pregnant at necropsy.

The number of females with live pups on Day 4 post partum was: 8 in the control group, 10 in each of the low and mid-dose groups, 6 in the high dose group.

Clinical signs

No signs of toxicological significance were observed in animals from the control, low and mid-dose groups and in the surviving female animals from the high dose group.

Body weight and body weight gain

No changes of toxicological significance were observed in body weight and body weight gain in treated males and females when compared to controls.

Food consumption

No effects on food consumption were observed in the males and in low and mid-dose females. Reduced food consumption was seen in the high dose females during the post coitum period.

Oestrus cycle, mating performance and reproductive parameters

Mean oestrus cycle was slightly reduced in the females dosed at 1000 mg/kg/day (approximately 2 in the 15 day pre-mating period) when compared to controls (approximately 3).

No significant differences between groups were observed in pre-coital interval and copulation plugs, as well as in the reproductive parameters (copulatory index and fertility index).

Implantation, pre-birth loss data and gestation length

No treatment-related differences were noted in implantation number, pre-birth loss data and gestation length between control and treated groups.

Litter data and sex ratios

Litter data and sex ratios were unaffected by treatment.

Pre-weaning clinical signs of pups

Clinical signs were comparable between treated and control groups.

Necropsy findings in decedent pups and in pups sacrificed on Day 4 post partum No treatment-related effects were seen.

Organ weights

No toxicologically relevant changes were observed in organ weights.

Macroscopic observations

No relevant changes were detected at post mortem examination in treated animals killed at end of treatment period, when compared with controls.

Microscopic observations

No treatment-related changes were seen in selected organs/tissues evaluated in high dose males or females receiving the test item, sacrificed at the end of treatment period, or in the abnormalities detected at post mortem.

Seminiferous tubules were evaluated with respect to their stage in the spermatogenic cycle and to the integrity of the various cell types within the different stages; regular layering in the germinal epithelium was noted.

OECD 407 (Müller 2000)

A study according to OECD Guideline 407 and GLP was performed to provide information on the toxicity and reversibility of possible effects of the test article when administered orally (by gavage) to the rat, once daily for 28 days with a treatment-free recovery period of further 14 days.

The daily oral administration of the test article at doses of 100, 300 and 1000 mg/kg body weight to rats for a period of 28 days was tolerated without any marked effects on physical condition.

None of the animals died during the course of investigation.

The following effects can be deemed as caused by the test article administration:

• The briefly plough through bedding with nose of the animals of the high dose group in a short period in the middle of the whole administration period as a sign of a temporary not general well-being.

• The activity of aspartateaminotransferase was statistically significantly decreased in the . high dose group of males and in the low and high dose group of females. This effect wasirreversible in the course of the recovery period. This changed enzyme activity remains of uncertain toxicological relevance as liver cell damage usually is associated with an increase in enzyme levels.

The body weight, the food consumption, and the coagulation parameters were not influenced by administration of the test article. No substance-dependent pathological macroscopic and histological findings were observed.

A part of the haematological and clinical-biochemical parameters and of the absolute and relative weights of organs showed statistically significant effects, which arehowever in the ranges of historical control data in the testing facility for the strain used and therefore they were evaluated as incidental and not caused by the test article administration. It is conspicuous that these incidental statistically significant effects occur in the high dose group in· the most cases.

It is to assume that this dose (1000 mg/kg b.w.) is the limit dose for the repeated administration of the test article without damage.

Based on the findings the dose of 1000 mglkg body weight / day (dose of the high dose group) can be declared as the NOAEL (No Observed Adverse Effect Level).

13 weeks feeding study (Maita 1981)

ICR mice and Wistar rats of both sexes were fed a diet containing ZnSO4 at 0, 300, 3000 and 30000 ppm for 13 weeks. Animals in the 30000 ppm group showed retarded growth along with low food intake, abnormal values in a few hematological parameters and regressive changes of the pancreatic exocrine gland. In addition, mice had decreased water intake and significant deviations in biochemical parameters, toxic lesions appeared in the stomach, intestine and spleen of both sexes and in the kidney of females. Four males and one female mouse were found dead or moribund during the study. The maximum no-effect level of ZnSO4 was determined to be 3000 ppm, which is approximately equivalent to the following milligram doses: mice: male 458 mg/kg/day, female 479 mg/kg/day, rats: male 234 mg/kg/day, female 243 mg/kg/day.

Considering the Zn concentration of max. 16.5 % in the target substance, the maximum no-effect level of the target substance was determined to be 4135.45 ppm, which is approximately equivalent to the following milligram doses: mice: male 631.35 mg/kg/day, female 660.29 mg/kg/day, rats: male 322.57 mg/kg/day, female 334.9 mg/kg/day.

Justification for classification or non-classification

Due to the findings in the available studies, the test item needs not to be classified according to Regulation (EC) No 1272/2008.