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Description of key information

In an OECD Test Guideline 407 study, to GLP, rats were administered diammonium hexachloroplatinate by gavage for 28 days. The systemic toxicity NOAEL was 10 mg/kg bw/day on the basis of clinical signs of toxicity (including reduced body weight and growth) in the mid and high dose groups (30 and 100 mg/kg bw/day), resulting in morphological changes to the kidneys and stomach (Hansen, 2015a).

 

In support, in an OECD Test Guideline 421 reproductive/developmental toxicity screening study, rats (12/sex/group) were administered diammonium hexachloroplatinate by oral gavage at doses of 0, 10, 30 or 100 mg/kg bw/day for at least 35 days. The study NOAEL was established to be 30 mg/kg bw/day for systemic toxicity (Hansen, 2015b).

 

No repeated dose toxicity studies by the inhalation or dermal route were identified, or are required.

 

Key value for chemical safety assessment

Repeated dose toxicity: via oral route - systemic effects

Link to relevant study records
Reference
Endpoint:
short-term repeated dose toxicity: oral
Type of information:
experimental study
Adequacy of study:
key study
Study period:
12 August 2014 - 11 June 2015
Reliability:
1 (reliable without restriction)
Rationale for reliability incl. deficiencies:
guideline study
Remarks:
Conducted according to GLP
Qualifier:
according to
Guideline:
OECD Guideline 407 (Repeated Dose 28-Day Oral Toxicity in Rodents)
Deviations:
no
Qualifier:
according to
Guideline:
EU Method B.7 (Repeated Dose (28 Days) Toxicity (Oral))
Deviations:
no
GLP compliance:
yes (incl. certificate)
Limit test:
no
Species:
rat
Strain:
Crj: CD(SD)
Sex:
male/female
Details on test animals and environmental conditions:
TEST ANIMALS
- Source: Charles River Laboratories, Research Models and Services, Germany GmbH, Sandhofer Weg 7, 97633 Sulzfeld, Germany.
- Age at study initiation: 60 days.
- Weight at study initiation: males, 314.4-342.7g; females, 221.6-243.4g.
- Fasting period before study: no data.
- Housing: animals were housed singly.
- Diet (e.g. ad libitum): certified commercial diet, offered ad libitum.
- Water (e.g. ad libitum): tap water, offered ad libitum.
- Acclimation period: 6 days.

ENVIRONMENTAL CONDITIONS
- Temperature (°C): room temperature of 22degC +/- 3degC (maximum range).
- Humidity (%): relative humidity of 55% +/- 15% (maximum range).
- Air changes (per hr): no data.
- Photoperiod (hrs dark / hrs light): 12 hours dark/12 hours light, about 150 lux.

IN-LIFE DATES:
From: June 2014.
To: 09 September 2014.
Route of administration:
oral: gavage
Vehicle:
corn oil
Details on oral exposure:
PREPARATION OF DOSING SOLUTIONS: test item formulations were prepared once weekly and stored in a tightly closed container at room temperature (10-25degC) until use.

DIET PREPARATION
- Rate of preparation of diet (frequency): not applicable.
- Mixing appropriate amounts with (Type of food): not applicable.
- Storage temperature of food: not applicable.

VEHICLE
- Justification for use and choice of vehicle (if other than water): the test substance was found to be unstable in water and other aqueous vehicles. Stability for 7 days in corn oil was demonstrated in an identity and stability investigation, and thus corn oil was taken forward as the vehicle for this study.
- Concentration in vehicle: 2, 6 or 20 mg test item/mL vehicle.
- Amount of vehicle (if gavage): 5 mL/kg bw/day.
- Lot/batch no. (if required): Batch no. 13249003, Caesar & Loretz GmbH, 40721 Hilden, Germany.
- Purity: no data.
Analytical verification of doses or concentrations:
yes
Details on analytical verification of doses or concentrations:
Samples of approximately 10 mL were collected once weekly, at the time of preparation of the formulation, and analysed for homogeneity and concentration.
Duration of treatment / exposure:
28 days.
Frequency of treatment:
Once daily.
Remarks:
Doses / Concentrations:
10 mg/kg bw/day
Basis:
actual ingested
Remarks:
Doses / Concentrations:
30 mg/kg bw/day
Basis:
actual ingested
Remarks:
Doses / Concentrations:
100 mg/kg bw/day
Basis:
actual ingested
No. of animals per sex per dose:
5
Control animals:
yes, concurrent vehicle
Details on study design:
- Dose selection rationale: dose levels were selected based on available toxicological data and a 14-day dose range finding study conducted at the same testing laboratory. In the dose range finding study, dose levels of 100, 125 and 200 mg/kg bw/day were administered; whilst all rats treated with the higher two groups died prematurely or were prematurely sacrificed due to pronounced clinical signs of toxicity, all animals treated with 100 mg/kg bw/day survived until scheduled sacrifice.
- Rationale for animal assignment (if not random): computer-generated randomisation programme.
- Rationale for selecting satellite groups: not applicable.
- Post-exposure recovery period in satellite groups: not applicable.
- Section schedule rationale (if not random): not applicable.
Positive control:
Not used.
Observations and examinations performed and frequency:
CAGE SIDE OBSERVATIONS: Yes.
- Time schedule: before and after dosing and, additionally, regularly throughout the working day (7:00 to 15:45 on Monday to Friday; 7:00 to 11:00 with a final check at 15:00 on Saturday and Sunday).
- Cage side observations included any behavioural changes, signs of illness or reaction to treatment. Skin/fur, eyes, mucous membranes, respiratory and circulatory systems, somatomotor activity and behaviour patterns were recorded, together with the onset, intensity and duration of any signs observed.

DETAILED CLINICAL OBSERVATIONS: Yes.
- Time schedule: once weekly, starting one week before the first administration and then on day 1 of test weeks 1-4 (inclusive).
- Signs recorded included: changes in skin, fur, eyes, mucous membranes; occurrence of secretions, excretions and autonomic activity; changes in gait, posture and response to handling; behavioural changes.

BODY WEIGHT: Yes.
- Time schedule for examinations: the weight of each rat was recorded at the time of group allocation (6 days prior to first test item administration), on test day 1 and once weekly thereafter (i.e. day 1 of weeks 2-4, inclusive).

FOOD AND WATER CONSUMPTION:
- Food consumption for each animal determined: Yes, on a weekly basis. Food intake per animal was determined using the total amount of food given to and left by each animal.
- Drinking water consumption for each animal determined: Yes, by daily visual appraisal throughout the study.

FOOD EFFICIENCY: No data.

OPHTHALMOSCOPIC EXAMINATION: No.

HAEMATOLOGY: Yes.
- Time schedule for collection of blood: samples were collected on the day of dissection (test day 29) from all study animals.
- Anaesthetic used for blood collection: Yes (isoflurane).
- Animals fasted: Yes (overnight).
- How many animals: all animals.
- Parameters checked in Table 1 were examined.

CLINICAL CHEMISTRY: Yes.
- Time schedule for collection of blood: samples were collected on the day of dissection (test day 29) from all study animals.
- Animals fasted: Yes (overnight).
- How many animals: all animals.
- Parameters checked in Table 2 were examined.

URINALYSIS: No.

NEUROBEHAVIOURAL EXAMINATION: Yes.
- Time schedule for examinations: Test day 23, approximately one to two hours after dosing.
- Dose groups that were examined: all animals.
- Battery of functions tested: see Table 3.
Sacrifice and pathology:
GROSS PATHOLOGY: Yes. The weights of the organs in Table 4 of all animals were recorded before fixation. The organs or parts of organs in Table 5 of all animals were fixed in 7% buffered formalin (apart from eyes, fixed in Davidson's solution, and testes, fixed in Bouin's solution).
HISTOPATHOLOGY: Yes. The organs in Table 5 of all control and high-dose animals, and the kidneys and stomach of all animals, were examined histologically.
Other examinations:
The myeloid:erythroid ratio of bone marrow cells was determined by cell differentiation. Samples were taken from the control and high-dose groups.
Statistics:
Test item-treated groups were compared to the control group using the following statistical methods:
All numerical function tests: Student's t-test
Body weight/food consumption/haematology/clinical chemistry/absolute and relative organ weights: Multiple t-test (Dunnett)
Bone marrow: Chi-squared test
Histology: Exact test (Fisher)
Clinical signs:
effects observed, treatment-related
Description (incidence and severity):
No test item-related deaths were observed. Clinical signs of toxicity reported in the high-dose females.
Mortality:
mortality observed, treatment-related
Description (incidence):
No test item-related deaths were observed. Clinical signs of toxicity reported in the high-dose females.
Body weight and weight changes:
effects observed, treatment-related
Description (incidence and severity):
Reduced body weight gain in mid-dose males. Reduced body weights, compared to control, in high-dose males and females.
Food consumption and compound intake (if feeding study):
effects observed, treatment-related
Description (incidence and severity):
Food consumption was reduced in high-dose males and females.
Food efficiency:
not examined
Water consumption and compound intake (if drinking water study):
no effects observed
Ophthalmological findings:
not examined
Haematological findings:
effects observed, treatment-related
Description (incidence and severity):
Test item-related increase in absolute number of leucocytes (WBC) and lymphocytes in high-dose males and females.
Clinical biochemistry findings:
effects observed, treatment-related
Description (incidence and severity):
Test item-related increase in ALAT activity in high-dose males and females.
Urinalysis findings:
not examined
Behaviour (functional findings):
effects observed, treatment-related
Description (incidence and severity):
Salivation seen in high-dose males. In high-dose females, neurobehavioural signs included piloerection, reduced motility, salivation, and hunched posture. All signs began to be observed within the last week of the study.
Organ weight findings including organ / body weight ratios:
effects observed, treatment-related
Description (incidence and severity):
High-dose males and females had increased relative and absolute kidney weights.
Gross pathological findings:
effects observed, treatment-related
Description (incidence and severity):
One male and one female high-dose animal had macroscopic alterations to the kidneys and stomach which were considered test item-related. One low-dose female displayed the same effect in the stomach.
Histopathological findings: non-neoplastic:
effects observed, treatment-related
Description (incidence and severity):
Test item-related morphological changes in the kidneys and stomach were observed in mid- and high-dose males and females.
Histopathological findings: neoplastic:
no effects observed
Details on results:
CLINICAL SIGNS AND MORTALITY
No test item-related deaths were observed. Clinical signs of toxicity reported in the high-dose females, including reduced amount of faeces, haemorrhagic nose and/or snout and/or a decreased body temperature (when assessed by touch).

BODY WEIGHT AND WEIGHT GAIN
Body weight gain in mid-dose males was reduced by 21% from test day 1 to 15 and by 18% from test day 1 to 18 compared with controls. In the high-dose group, body weight gain was reduced by 32% in males and 27% in females on test day 28, compared to the control group. Body weights of the high-dose animals, again compared to controls, were reduced by 7% (males) and 12% (females).

FOOD CONSUMPTION
Food consumption at the end of test week 1 was statistically significantly reduced, by 18% in males and 48% in females compared to controls. Food intake improved during the course of the study, and at test week 4 the reduction was not statistically significant, 11% in males and 6% in females.

WATER CONSUMPTION
The daily visual appraisal of the drinking water consumption did not reveal any differences to the control at any dose level.

HAEMATOLOGY
The only test item-related alteration to the assessed haematology parameters was an increase in the absolute number of leucocytes (WBC) and lymphocytes in the high-dose group. These were increased by 34 and 56% respectively in males, and by 84 and 73% respectively in females.

Statistically significant decreases in HGB and MCHC in high-dose females, and significant decreases in aPTT and significant increases in MCV and MCH in high-dose males, when compared to the controls, were considered to be within the normal range of natural variation and thus not related to test item treatment. A statistically significant increase in TPT in mid-dose males was also not considered to be compound related, as it lacked dose dependency.

CLINICAL CHEMISTRY
The activity of ALAT was more than 2-fold that of the control group in high-dose males, and nearly 3-fold the control group value in high-dose females. No other test item-related changes to the assessed clinical chemistry parameters were reported.

The statistically significant decreases in albumin and total protein in high-dose females, compared to controls, were considered to be within the normal range of natural variation, and thus not related to test substance administration.

NEUROBEHAVIOUR
Slight salivation was observed in high-dose males and females, starting on test day 23 or 24. In high-dose females, reduced motility and a hunched posture were also reported.

ORGAN WEIGHTS
Relative and absolute kidney weights were increased, compared to controls, in high-dose males and females (see Table 6). These findings correlate with the macroscopic and histopathological findings.

Statistically significant but not test item-related changes were reported to the weights of the adrenal gland, brain, epididymes, liver and spleen in various tested groups.

GROSS PATHOLOGY
One male animal in the high-dose group was found to have enlarged kidneys and adrenal glands. Another male in the same group had a stomach ulcer.

A high-dose female had yellowish discoloured kidneys; another had pale adrenal glands. A third animal in this group had a white adhesion to the mucosa of the cardiac region of the stomach.

One female animal in the low-dose group also presented with a white adhesion to the mucosa of the cardiac region of the stomach.

HISTOPATHOLOGY: NON-NEOPLASTIC
Histopathological examination revealed test item-related morphological lesions in the kidneys and stomach of males and females in the mid- and high-dose groups (See Table 7).

OTHER FINDINGS
The myeloid:erythroid ratio of high-dose males and females was not altered by treatment.
Dose descriptor:
NOAEL
Effect level:
10 mg/kg bw/day (actual dose received)
Based on:
test mat.
Sex:
male/female
Basis for effect level:
other: No general systemic effects seen at 10 mg/kg bw/day
Dose descriptor:
LOAEL
Effect level:
30 mg/kg bw/day (actual dose received)
Based on:
test mat.
Sex:
male
Basis for effect level:
other: Reduction in body weight gain compared to control group.
Critical effects observed:
not specified

Table 6: test item-related changes to kidney weights in high-dose animals, compared to the control group on test day 29.

     Males  Females
 Kidney, left  Relative  +35% (p≤0.05)  +45%  (p≤0.01)
   Absolute  +28%  +36%  (p≤0.05)
 Kidney, right  Relative  +44%  (p≤0.01)  +45%  (p≤0.01)
   Absolute  +36%  (p≤0.05)  +37%  (p≤0.01)

Table 7: histopathological findings

Kidney   
 Tubular basophilia  Mid-dose females (2/5)
   High-dose males and females (10/10)
 Hyaline casts  Mid-dose females (2/5)
   High-dose males and females (3/5, 2/5)
 Lymphocytic infiltration  High-dose males and females (4/5, 5/5)
 Tubular dilation  High-dose males and females (2/5, 2/5)
 Tubular necrosis  High-dose females (2/5)
 Fibrosis  High-dose males (1/5)
 Stomach  
 Lympho-histio-granulocytic infiltration  High-dose males and females (10/10)
 Eosinophilic infiltration  High-dose females (1/5)
 Chronic/subacute ulcerative inflammation of the glandular mucosa  High-dose males and females (1/5, 1/5)
Conclusions:
In an OECD Test Guideline 407 study, to GLP, rats (5/sex/group) were administered diammonium hexachloroplatinate by gavage for 28 days. The systemic toxicity NOAEL was 10 mg/kg bw/day on the basis of clinical signs of toxicity (including reduced body weight and growth) in the mid and high dose groups (30 and 100 mg/kg bw/day), resulting in morphological changes to the kidneys and stomach
Executive summary:

In a 28-day repeated dose oral toxicity study, conducted according to OECD Test Guideline 407 and to GLP, rats (5/sex/group) were gavaged with diammonium hexachloroplatinate via stomach tube at doses of 0 (vehicle control, given corn oil), 10, 30 or 100 mg/kg bw/day. Rats were observed for signs of toxicity and mortality, and changes in body weight. Neurobehaviour was assessed (using a functional observation battery). On day 29, blood samples were collected for the analysis of haematological parameters and clinical chemistry. This was immediately followed by sacrifice and scheduled necropsy, in which a comprehensive range of organs and tissues were examined macroscopically and microscopically.

Males in the mid-dose group had reduced body weight and body weight gain, compared to controls, and mid-dose females were found to have histopathological lesions in the kidneys (tubular basophilia and hyaline casts) and stomach. In the high-dose group, body weights and body weight gain were affected in both sexes, and lesions of the kidneys and stomach had increased in incidence and severity. Additional findings were noted in the kidney (lymphocytic infiltration, fibrosis, tubular dilation and tubular necrosis) and stomach (lympho-histio-granulocytic infiltration, lymphocytic or eosinophilic infiltration, and/or a chronic/subacute and ulcerative inflammation) at the high dose. Furthermore, high-dose animals were observed to be salivating after treatment. Other clinical signs of toxicity in high-dose females were piloerection, reduced motility, reduced amount of faeces, hunched posture, haemorrhagic nose and/or snout and a decreased body temperature.

Under the conditions of this study, the no-observed-adverse-effect level (NOAEL) for systemic toxicity was 10 mg/kg bw/day.

Endpoint conclusion
Endpoint conclusion:
adverse effect observed
Dose descriptor:
NOAEL
10 mg/kg bw/day
Study duration:
subacute
Species:
rat
Quality of whole database:
Overall, good-quality database which meets REACH Standard Information Requirements.
System:
urinary

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

Mode of Action Analysis / Human Relevance Framework

No data identified.

Additional information

No relevant human data were identified, or repeated dose toxicity laboratory animal data with dipotassium hexachloroplatinate.

 

However, a reliable 28-day oral gavage repeated dose toxicity and a reproduction/developmental screening toxicity study have been conducted with diammonium hexachloroplatinate. Diammonium hexachloroplatinate is considered to fall within the scope of the read-across category "hexachloroplatinate(IV) compounds". See section 13 in IUCLID for full read-across justification report.

 

In a 28-day repeated dose oral toxicity study, conducted according to OECD Test Guideline 407 and to GLP, rats (5/sex/group) were gavaged with diammonium hexachloroplatinate via stomach tube at doses of 0 (vehicle control, given corn oil), 10, 30 or 100 mg/kg bw/day. Rats were observed for signs of toxicity and mortality, and changes in body weight. Neurobehaviour was assessed (using a functional observation battery). On day 29, blood samples were collected for the analysis of haematological parameters and clinical chemistry. This was immediately followed by sacrifice and scheduled necropsy, in which a comprehensive range of organs and tissues were examined macroscopically and microscopically. Males in the mid-dose group had reduced body weight and body weight gain, compared to controls, and mid-dose females were found to have histopathological lesions in the kidneys (tubular basophilia and hyaline casts) and stomach. In the high-dose group, body weights and body weight gain were affected in both sexes, and lesions of the kidneys and stomach had increased in incidence and severity. Additional findings were noted in the kidney (lymphocytic infiltration, fibrosis, tubular dilation and tubular necrosis) and stomach (lympho-histio-granulocytic infiltration, lymphocytic or eosinophilic infiltration, and/or a chronic/subacute and ulcerative inflammation) at the high dose. Furthermore, high-dose animals were observed to be salivating after treatment. Other clinical signs of toxicity in high-dose females were piloerection, reduced motility, reduced amount of faeces, hunched posture, haemorrhagic nose and/or snout and a decreased body temperature. Under the conditions of this study, the no-observed-adverse-effect level (NOAEL) for systemic toxicity was 10 mg/kg bw/day (Hansen, 2015a).

 

In support, in a GLP OECD Test Guideline 421 reproductive/developmental toxicity screening study, rats (12/sex/group) were administered diammonium hexachloroplatinate by oral gavage at doses of 0, 10, 30 or 100 mg/kg bw/day for at least 35 days. Parental (F0) animals were observed for clinical signs of toxicity throughout the study, with body weights and food consumption monitored. At necropsy, animals were subjected to external and internal macroscopic examinations for any abnormalities or pathological changes. Special attention was paid to the reproductive organs. Two females in the high-dose group died prematurely. Surviving high-dose animals displayed slight to extreme salivation, piloerection, and/or pale faeces were reported. Increased water consumption was observed in high-dose males. Salivation and piloerection were also observed in low- and mid-dose animals. In the high-dose group, body weights were significantly reduced compared to controls, and cellular changes to the stomach and kidneys were also reported. No test item-related microscopic changes were noted in the reproductive organs at any dose level. The study NOAEL was established to be 30 mg/kg bw/day for systemic toxicity (Hansen, 2015b).

 

According to REACH Annex VIII (EC 1907/2006), repeated dose toxicity studies only need to be conducted on one species taking into consideration the most appropriate route of administration regarding human exposure.

Justification for classification or non-classification

Based on the histopathological effects on the kidney observed at 30 mg/kg bw/day and above in the 28-day rat study with diammonium hexachloroplatinate, dipotassium hexachloroplatinate should also be classified as STOT RE1 according to EU CLP criteria (EC 1272/2008). The observed effects on the stomach are likely the result of local irritancy.