Registration Dossier

Administrative data

Description of key information

NOAEL values of 250 mg/kg bw/day and 300 mg/kg bw/day were obtained from a 90 day study on a structural similar substance and from a 28 day study on the substance itself.

Key value for chemical safety assessment

Repeated dose toxicity: via oral route - systemic effects

Link to relevant study records
Reference
Endpoint:
sub-chronic toxicity: oral
Type of information:
read-across from supporting substance (structural analogue or surrogate)
Adequacy of study:
key study
Study period:
11-2011 until 03-2012
Reliability:
1 (reliable without restriction)
Rationale for reliability incl. deficiencies:
guideline study
Reason / purpose:
read-across source
Qualifier:
according to
Guideline:
OECD Guideline 408 (Repeated Dose 90-Day Oral Toxicity in Rodents)
Deviations:
no
GLP compliance:
yes
Limit test:
no
Species:
rat
Strain:
Wistar
Sex:
male/female
Details on test animals and environmental conditions:
TEST ANIMALS
- Source: Harlan Italy s.r.l., San Pietro al Natisone (UD), Italy
- Age at study initiation: 27-29 days
- Weight at study initiation: 94-112 g (males), 96-109 g (females)
- Fasting period before study: not applicable
- Housing: polycarbonate cages
- Diet (e.g. ad libitum): laboratory rodent diet (4 RF 21, Mucedola S.r.l., Via G. Galilei, 4, 20019, Settimo Milanese (MI), Italy)
- Water (e.g. ad libitum): ad libitum
- Acclimation period: 18 days

ENVIRONMENTAL CONDITIONS
- Temperature (°C): 22 ± 2
- Humidity (%): 55 ± 15
- Air changes (per hr): 15-25
- Photoperiod (hrs dark / hrs light): 12/12

IN-LIFE DATES: From: To:
Route of administration:
oral: gavage
Vehicle:
water
Details on oral exposure:
PREPARATION OF DOSING SOLUTIONS:

DIET PREPARATION
- Rate of preparation of diet (frequency):
- Mixing appropriate amounts with (Type of food):
- Storage temperature of food:

VEHICLE
- Justification for use and choice of vehicle (if other than water): not applicable
- Concentration in vehicle: 5, 25 and 100 mg/mL
- Amount of vehicle (if gavage): 10 ml/kg body weight
- Lot/batch no. (if required): not required
- Purity: purified water
Analytical verification of doses or concentrations:
yes
Details on analytical verification of doses or concentrations:
Prior to commencement of treatment, analysis was performed to confirm that the proposed formulation procedure was acceptable (content check).

Formulates were tested for the content in disodium sulfinatoacetate, disodium sulfonatoacetate and disodium sulphite.

Formulates of BRUGGOLITE FF6 in purified water tested in the study resulted corresponding to the theoretical doses ( 5 mg/mL, 25 mg/mL and 100 mg/mL respectively).
Duration of treatment / exposure:
13 weeks
Frequency of treatment:
once a day, 7 days a week
Dose / conc.:
0 mg/kg bw/day (nominal)
Remarks:
nominal in water
Dose / conc.:
50 mg/kg bw/day (nominal)
Remarks:
nominal in water
Dose / conc.:
250 mg/kg bw/day (nominal)
Remarks:
nominal in water
Dose / conc.:
1 000 mg/kg bw/day (nominal)
Remarks:
nominal in water
No. of animals per sex per dose:
10
Control animals:
yes
Details on study design:
- Post-exposure recovery period in satellite groups: 4 weeks
- Dose selection rationale: Dose levels were selected in consultation with the Sponsor based on information from preliminary studies.
- Rationale for animal assignment (if not random): not applicable
- 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 applicable
Observations and examinations performed and frequency:
CAGE SIDE OBSERVATIONS: Yes
- Time schedule: Starting from Day 37 of the study, observations of the cage tray were performed and recorded at least three times weekly.

DETAILED CLINICAL OBSERVATIONS: Yes
- Time schedule:
Once before commencement of treatment and once per week from the start of treatment, each animal was given a detailed clinical examination. Each animal was observed in an open arena. The test included observation of changes in gait and posture, reactivity to handling, presence of clonic or tonic movements, stereotypies or bizarre behaviour and effects on the autonomic nervous system (e.g. lachrymation, piloerection, unusual respiratory pattern). Changes in fur, skin, eyes, mucous membranes, occurrences of secretions and excretions were also recorded. Once during Week 12 of treatment and once during Week 4 of recovery, an evaluation of sensory reactivity to stimuli of different modalities (e.g. auditory, visual and proprioceptive stimuli) and an assessment of grip strength were also performed.

BODY WEIGHT: Yes
- Time schedule for examinations: Each animal was weighed on the day of allocation to treatment group, on the day that treatment commenced, weekly thereafter and just prior to necropsy.

FOOD CONSUMPTION AND COMPOUND INTAKE (if feeding study):
- Food consumption for each animal determined and mean daily diet consumption calculated as g food/kg body weight/day: Yes
- Compound intake calculated as time-weighted averages from the consumption and body weight gain data: No

FOOD EFFICIENCY:
- Body weight gain in kg/food consumption in kg per unit time X 100 calculated as time-weighted averages from the consumption and body weight gain data: No

WATER CONSUMPTION AND COMPOUND INTAKE (if drinking water study): not applicable

OPHTHALMOSCOPIC EXAMINATION: Yes
- Time schedule for examinations: pior to the study and in week 13 of the study
- Dose groups that were examined: all

HAEMATOLOGY: Yes
- Time schedule for collection of blood: in week 13 of the study and in week 4 of the recovery period
- Anaesthetic used for blood collection: Yes (isofluorane)
- Animals fasted: Yes, under conditions of food and water deprivation.
- How many animals: 10 male and 10 female of main study, all animals of recovery group
- Parameters checked:
Haematocrit
Haemoglobin
Red blood cell count
Reticulocyte count
Mean red blood cell volume
Mean corpuscular haemoglobin
Mean corpuscular haemoglobin concentration
White blood cell count
Differential leucocyte count - Neutrophils
- Lymphocytes
- Eosinophils
Report Page 20 of 480
RTC Study No.: 85850 TEXT - Page 21 of 31
- Basophils
- Monocytes
- Large unstained cells
Platelets

CLINICAL CHEMISTRY: Yes
- Time schedule for collection of blood: see haematology
- Animals fasted: see haematology
- How many animals: see haematology
- Parameters checked:
Alkaline phosphatase
Alanine aminotransferase
Aspartate aminotransferase
Gamma-glutamyltransferase
Urea
Creatinine
Glucose
Triglycerides
Inorganic Phosphorus
Total bilirubin
Total cholesterol
Total protein
Albumin
Globulin
Albumin/Globulin Ratio
Sodium
Potassium
Calcium
Chloride

URINALYSIS: No

NEUROBEHAVIOURAL EXAMINATION: Yes
- Time schedule for examinations: prior to examination and once a week
- Dose groups that were examined: all
Sacrifice and pathology:
GROSS PATHOLOGY: Yes
HISTOPATHOLOGY: Yes
GROSS PATHOLOGY: Yes (see table 1)

Organ weights:
From all animals completing the scheduled test period, the organs indicated in table 1 were dissected free of fat and weighed. The ratios of organ weight to body weight were calculated for each animal.

Tissues fixed and preserved:
Samples of all the tissues listed in table 1 were fixed and preserved in 10% neutral buffered formalin (except eyes, testes and epididymides which were fixed in Modified Davidson's fluid and preserved in 70% ethyl alcohol).

HISTOPATHOLOGY: Yes (see table 1)

The tissues required for histopathological examination are listed in table 1. After dehydration and embedding in paraffin wax, sections of the tissues were cut at 5 micrometre thickness and stained with haematoxylin and eosin. The examination was as detailed below:

a) Tissues specified in table 1 from all animals in the control and high dose groups dying during the treatment period or killed at the end of the 13 weeks of treatment
b) Tissues specified in table 1 from the animal dying during the treatment period
c) All abnormalities in all main phase groups

On the basis of the histopathological changes observed between control and high dose groups, the examination was extended to:
- kidneys and stomach: from male and female animals of Groups 2 and 3 of the main phase and from control and Group 4 animals (males and females) of the recovery phase;
- urinary bladder: from male animals of Groups 2 and 3 of the main phase and from control and Group 4 male animals of the recovery phase.
Other examinations:
not applicable
Statistics:
Standard deviations were calculated as considered appropriate. For continuous variables the significance of the differences amongst group means was assessed by analysis of variance. Differences between each treated group and the control group were assessed by Dunnett’s test using a pooled error variance. The homogeneity of the data was verified by Bartlett’s test. If the data were found to be heterogeneous, a modified t test (Cochran and Cox) was
applied. The mean values, standard deviations and statistical analysis were calculated from actual values in the computer without rounding off. Statistical analysis of histopathological findings was carried out by means of the non-parametric Kolmogorov-Smirnov test.
Clinical signs:
effects observed, treatment-related
Description (incidence and severity):
Clinical signs: Dosing phase: Decreased activity piloerection, hunched posture, rales, eye damage, soft faeces (incidence and explanation, see details on results). Recovery phase: No clinical signs were noted during the recovery period with the exception of one female previously dosed with 1000 mg/kg/day which showed hairloss on neck. Mortality: five unscheduled deaths (incidence and explanation, see details on results).
Mortality:
mortality observed, non-treatment-related
Description (incidence):
Clinical signs: Dosing phase: Decreased activity piloerection, hunched posture, rales, eye damage, soft faeces (incidence and explanation, see details on results). Recovery phase: No clinical signs were noted during the recovery period with the exception of one female previously dosed with 1000 mg/kg/day which showed hairloss on neck. Mortality: five unscheduled deaths (incidence and explanation, see details on results).
Body weight and weight changes:
no effects observed
Description (incidence and severity):
Dosing phase: No differences in body weight were observed between control and treated groups. Recovery phase: During the recovery phase, body weight of treated animals of both sexes was comparable to the control group.
Food consumption and compound intake (if feeding study):
no effects observed
Description (incidence and severity):
Dosing phase: No effects in food consumption were observed in treated groups when compared to the control group. Recovery phase: No food consumption differences were noted between control and treated groups.
Food efficiency:
not examined
Water consumption and compound intake (if drinking water study):
not examined
Ophthalmological findings:
no effects observed
Description (incidence and severity):
No significant findings were detected at the ophthalmoscopic examination performed during Week 13 of treatment.
Haematological findings:
effects observed, treatment-related
Description (incidence and severity):
Dosing phase: A slight decrease of erythrocytes, haemoglobin and haematocrit (approximately 10%) was recorded in some animals (see details on results). Recovery phase: No differences between control and treatment animals were observed.
Clinical biochemistry findings:
effects observed, treatment-related
Description (incidence and severity):
Statistically significant changes of some biochemical parameters were observed in treated animals (see details on results).
Urinalysis findings:
not examined
Behaviour (functional findings):
effects observed, treatment-related
Description (incidence and severity):
Dosing phase: Weekly detailed clinical examinations: No neurotoxicological significant alterations were recorded (see details on results).
Immunological findings:
not examined
Organ weight findings including organ / body weight ratios:
effects observed, treatment-related
Description (incidence and severity):
A statistically significant increase in absolute kidneys and liver weights was noted in females receiving 1000 mg/kg/day (+19% and +14%, respectively). see details on results
Gross pathological findings:
effects observed, treatment-related
Description (incidence and severity):
Final sacrifice: Treatment-related macroscopic changes were noted in the kidneys and in the stomach (see details on results).
Neuropathological findings:
no effects observed
Histopathological findings: non-neoplastic:
effects observed, treatment-related
Description (incidence and severity):
Final sacrifice: Treatment-related microscopic changes were noted in the kidneys and in the stomach in both sexes, and in the urinary bladder of the males receiving BRUGGOLITE FF6 M at 1000 mg/kg/day (high dose) (see details on results).
Histopathological findings: neoplastic:
no effects observed
Details on results:
CLINICAL SIGNS AND MORTALITY

CLINICAL SIGNS:

Dosing phase
Decreased activity was noted in 5 out of 15 males receiving 1000 mg/kg/day. Piloerection was observed in two females, one receiving 50 mg/kg/day and one receiving 1000 mg/kg/day. The female receiving 50 mg/kg/day also showed hunched posture. Rales were recorded in one control female. All these signs were noted only for a few days during the treatment period. In addition, damaged eye was observed in two females, one receiving 250 mg/kg/day and one receiving 1000 mg/kg/day. This sign was observed towards the end of the study and was correlated to the bleeding procedure performed for clinical pathology investigations. Soft faeces were recorded on the cage tray of all males (starting from Day 37 of the study) and in one cage tray of females (starting from Day 52 of the study) receiving 1000 mg/kg/day. After Day 66 of the study until termination no signs were observed.

Recovery phase
No clinical signs were noted during the recovery period with the exception of one female previously dosed with 1000 mg/kg/day which showed hairloss on neck.

MORTALITY:

Five unscheduled deaths during the study (see table 2).
All unscheduled dead animals did not show any clinical signs the day of death or shortly before.

Animal no.: 85850004: Macroscopic changes consisted of presence of a catheter in the esophagus and duodenum; and presence of dark red areas in the thymus. Histologically, the observed changes were: renal minimal focal nephropathy and thymic congestion. The cause of death is suggested to be due to a gavage error.

Animal no.: 85850031: Macroscopic changes consisted of presence of adhesions on the heart; dark kidneys, liver, lungs, mesenteric lymph nodes and ovaries. Histologically, the noted changes were: acute inflammation of the pericardium and pleura; congestion of the kidneys, liver, mesenteric lymph nodes, ovaries; atrophy and inflammation of the thymus. The cause of death is suggested to be due to a gavage error.

Animal no.: 85850085: Macroscopic changes consisted of red and/or dark colour of the meninges (brain), liver, lungs, pituitary, and thymus; presence of dark red fluid in the thoracic cavity. Histologically, the observed changes were: congestion of the meninges, thymus and liver; pulmonary alveolar hyperplasia and presence of histiocytes. The cause of death is not clear, but due to the presence of fluid in the thoracic cavity (without evidence of
inflammation in thoracic organs), it is potentially due to a gavage error.

Animal no.: 85850091: Macroscopic changes consisted of cannibalisation of multiple organs; red and/or dark color of the cervical lymph nodes, lungs and stomach. Histologically, the observed change was: congestion of the cervical lymph nodes. The cause of death is unknown.

Animal no.: 85850095: Macroscopic changes consisted of dark and/or red adrenals, brain, liver, lungs, and pituitary; renal pelvic dilation; and presence of dark red clear fluid in the thoracic cavity. Histologically, the noted change was: congestion of the lungs, liver and pituitary. The cause of death is not clear, but due to the presence of fluid in the thoracic cavity (without evidence of inflammation in thoracic organs), it is potentially due to a gavage error.


BODY WEIGHT AND WEIGHT GAIN

Dosing phase
No differences in body weight were observed between control and treated groups.

Recovery phase
During the recovery phase, body weight of treated animals of both sexes was comparable to the control group.


FOOD CONSUMPTION AND COMPOUND INTAKE (if feeding study)

Dosing phase
No effects in food consumption were observed in treated groups when compared to the control group.

Recovery phase
No food consumption differences were noted between control and treated groups.


FOOD EFFICIENCY

not determined


WATER CONSUMPTION AND COMPOUND INTAKE (if drinking water study)

not determined


OPHTHALMOSCOPIC EXAMINATION

No significant findings were detected at the ophthalmoscopic examination performed during Week 13 of treatment.


HAEMATOLOGY

Dosing phase
A slight decrease of erythrocytes, haemoglobin and haematocrit (approximately 10%) was recorded in some animals treated with 1000 mg/kg/day. Significant reticulocytosis was recorded in only one female (animal no. 85850075). Due to the low severity, the above mentioned changes were not considered to be of toxicological relevance. The other statistically significant changes recorded (mean corpuscular volume and leucocytes in males) were of minimal magnitude and/or not dose-related, therefore considered unrelated to treatment. Prothrombin time was decreased in two males treated with 1000 mg/kg/day (animal nos. 85850076 and 85850084). Due to the low incidence and severity (mean group value was 5% below controls), the above mentioned change was considered of no toxicological significance.

Recovery phase
No differences between control and treatment animals were observed.


CLINICAL CHEMISTRY

Dosing phase
Statistically significant changes of some biochemical parameters were observed in treated animals. Males dosed with 1000 mg/kg/day showed decrement of alkaline phosphatase (-25%), gamma-glutamyl transferase (-33%), creatinine (-23%), protein (-9%), albumin (-7%), globulin (-15%) and increment of alanine aminotransferase (+40%) and phosphorus (+17%). Females from the same group showed decrement of bilirubin (-42%), chloride (-3%), sodium (-4%) and potassium (-19%) and increment of triglycerides (+106%) and urea (+24%). Some of the above mentioned changes were also observed in animals treated with 250 mg/kg/day (gammma-glutamyl transferase, protein and globulin in males, chloride, sodium, potassium in females). Even though these findings did not represent a clear adverse effect, it cannot be concluded that they are unrelated to treatment. Sodium was significantly increased in males treated with 50 and 250 mg/kg/day. Due to the low severity (2%) and the lack of dose-relation, this change was considered incidental.

Recovery phase
At the end of the recovery phase, most of the observed findings showed complete reversibility and the remaining differences between control and treated animals mainly referred to only one treated animal. In particular, animal no. 85850092 showed elevated values of transaminases enzymes and gammma-glutamyl transferase and female no. 85850093 showed high aspartate aminotransferase and triglycerides. At the end of the recovery period, all changes showed a complete reversibility.


URINALYSIS

not determined


NEUROBEHAVIOUR

Dosing phase

Weekly detailed clinical examinations
No neurotoxicological significant alterations were recorded at the examinations (removal from the cage and open arena) performed during the treatment period.

Sensory reactivity to stimuli
A statistically significant increase in grip strength means was observed in males receiving the dose level ≥ 250 mg/kg/day, when compared to the control group. This tendency was also observed in treated females and was due to the fact that few animals showed intergroup outlier values.

Motor activity
No statistically significant alteration in motor activity was recorded in any treatment group at the examinations performed at the end of treatment period.

Recovery phase
Neurotoxicity assessment, including weekly detailed clinical examinations, sensory reactivity to stimuli and motor activity, did not show any alteration during the recovery period.


ORGAN WEIGHTS

Final sacrifice
No differences were noted in terminal body weight in treated animals of both sexes compared to the control group.

Absolute organ weights
A statistically significant increase in absolute kidneys and liver weights was noted in females receiving 1000 mg/kg/day (+19% and +14%, respectively).

Relative organ weights
Relative kidneys (+15% in males; +16% in females) and liver (+14% in males; +11% in females) weights were increased with a statistical significance in treated animals of both sexes receiving 1000 mg/kg/day, while relative spleen weight was increased in females receiving 50 mg/kg/day.

Recovery sacrifice
Terminal body weight of treated animals was comparable to the control groups.

Absolute and relative organ weights
No differences were observed between treated and control groups.


GROSS PATHOLOGY

Final sacrifice
Treatment-related macroscopic changes were noted in the kidneys and in the stomach. In the kidneys abnormal areas (i.e. depressed and/or pale) were increased in incidence in the males and females treated with the high dose. In the stomach, increased incidence of thickened limiting ridge (i.e., the non-glandular transition line of the forestomach, close to the glandular stomach) was noted in the males and females treated with the high dose. The above-mentioned macroscopic changes correlated with treatment-related histopathological findings. All other lesions seen had a comparable incidence in control and treated animals and were known to occur spontaneously and/or incidentally in Wistar Hannover rats of the same age, used at our laboratory, under similar experimental conditions, and therefore are not considered as related to treatment.

Recovery sacrifice
No treatment-related macroscopic changes were noted.


HISTOPATHOLOGY: NON-NEOPLASTIC

Final sacrifice
Treatment-related microscopic changes were noted in the kidneys and in the stomach in both sexes, and in the urinary bladder of the males receiving BRUGGOLITE FF6 M at 1000 mg/kg/day (high dose).

Kidneys: Increased incidence and severity of nephropathy were noted in male and female rats treated with the high dose, being more frequent and severe in the males. The severity of the nephropathy ranged from minimal (grade 1) to moderate (grade 3) in the males, and was mild (grade 2) in the females. The nephropathy was characterized by the presence of dilated tubules, extending in the cortex and medulla, and lined by basophilic, regenerated epithelium. The basement membrane was mostly thickened, occasional exfoliated cells or protein casts were noted in the lumen. Subchronic inflammation was noted in the interstitium. In some animals, the presence of crystal-like unstained aggregates was noted in the tubular lumen or in the pelvic cavity. Only in the males, pelvic dilation, associated with hyperplasia of the pelvic lining epithelium, or of the epithelium lining the papilla, was
observed.

Urinary bladder: In the males only, hyperplasia, ranging from minimal to mild, was noted. This was multifocal or diffuse.

Forestomach: Minimal to mild focal hyperplasia of the limiting ridge only was noted.

No treatment-related change was noted in the target organs in the low and intermediate dose animals. Minimal focal epithelial cell degeneration was noted in the limiting ridge of non grandular region of the stomach in a single male (no. 85850070) of intermediate dose group.

All other lesions seen had a comparable incidence in control and treated animals and were known to occur spontaneously and/or incidentally in Wistar Hannover rats of the same age, used at our laboratory, under similar experimental conditions.

Recovery sacrifice
Treatment-related changes were observed in the kidneys and in the urinary bladder of treated animals sacrificed after recovery period. Nephropathy was noted associated with interstitial inflammatory cell infiltration and fibroblastic reaction. Pelvic epithelial hyperplasia was also observed in the kidneys of one female. Epithelial hyperplasia was reported in the urinary bladder of one male.


HISTOPATHOLOGY: NEOPLASTIC (if applicable)

not applicable


HISTORICAL CONTROL DATA (if applicable)

not applicable


OTHER FINDINGS

none
Dose descriptor:
NOAEL
Effect level:
ca. 250 mg/kg bw/day (nominal)
Based on:
test mat.
Sex:
male/female
Basis for effect level:
clinical signs
mortality
body weight and weight gain
ophthalmological examination
haematology
behaviour (functional findings)
organ weights and organ / body weight ratios
gross pathology
Critical effects observed:
not specified

table 2 (mortality):

Animal number Group/Sex Day of death Description of death
85850004 1/M 29 Humane kill
85850031 2/F 38 Found dead
85850085 4/F 5 Found dead
85850091 4/F 6 Found dead
85850095 4/F 5 Found dead
Conclusions:
On the basis of the above mentioned results, the NOAEL (No Observed Adverse Effect Level) for males and females could be considered 250 mg/kg/day.
Executive summary:

The toxicity of BRUGGOLITE FF6 M (50, 250 and 1000 mg/kg/day) was investigated,

when given orally to rats for 13 weeks followed by a recovery period of 4 weeks.

One control male and four females (3 receiving 1000 mg/kg/day and one receiving 50

mg/kg/day) were unscheduled dead during the study between Days 5 and 38. No clinical

signs were observed in these animals the day of death or shortly before.

The findings noted at macroscopic and microscopic observations suggested that the cause of

death of these animals was due to a gavage error or to a potential gavage error, with the

exception of one animal which was cannibalised and therefore the cause of death was

unknown.

Dcreased activity and piloerection were noted in males and females receiving

1000 mg/kg/day, respectively. Hunched posture and piloerection were also noted in females

receiving 50 mg/kg/day. All these signs were observed for a few days.

Soft faeces were observed in all cage trays of males and in one cage tray of females

receiving 1000 mg/kg/day.

During the recovery phase, all clinical signs previously observed showed complete

reversibility.

During the treatment period, neurotoxicity assessment (including motor activity and sensory

reaction to stimuli), body weight, food consumption, and ophthalmoscopy of treated groups

did not show relevant signs, when compared to the control group.

Also during the recovery period, all the above parameters of the treated groups were

comparable to the control group.

At haematological evaluation, no changes of toxicological relevance were observed after

13 weeks of treatment.

No differences between control and treated animals were observed at the end of the recovery

period.

Increases in alanine aminotransferase (males), phosphorus (males), triglycerides (females)

and urea (females) were recorded at clinical chemistry evaluation in animals receiving the

highest dosage (1000 mg/kg/day).

In addition, a decrease in alkaline phoshatase, -glutamyl transferase, creatinine, protein,

albumin and globulin was noted in males, while a decrement of bilirubin, chloride,

potassium and sodium was observed in females, both receiving 1000 mg/kg/day.

At the end of the recovery period, all changes showed complete reversibility.

However one male previously dosed with 1000 mg/kg/day showed a high value of

-glutamyl transferase and one female had high values of aspartate aminotransferase and

triglycerides.

Terminal body weights of treated animals were comparable to the control group both in

dosing and recovery phase.

At the end of the treatment period, absolute kidneys and liver weights were increased in

females receiving 1000 mg/kg/day. In addition, relative kidneys and liver weights were also

increased in males and females receiving1000 mg/kg/day.

No treatment-related effect on organ weights was noted at the end of the recovery period.

Treatment-related macroscopic findings were recorded in animals of both sexes receiving

1000 mg/kg/day and sacrificed at termination. These findings were observed in the kidneys

and stomach and were correlated with the treatment-related findings noted at the

histopathological examinations.

In addition, treatment-related microscopic changes were noted in the kidneys (nephropathy,

more frequent and severe in the males, subchronic inflammation in the interstitium and

presence of crystal-like unstained aggregates in the tubular lumen or in the pelvic cavity), in

the stomach (hyperplasia of the limiting ridge) of males and females and in the urinary

bladder (hyperplasia, multifocal or diffuse) of the males receiving 1000 mg/kg/day.

Moreover minimal to mild epithelial hyperplasia in the urinary bladder was also noted in the

males only.

Most of the findings observed in males and females at the end of the treatment period were

still noted after the recovery period and were: nephropathy in the kidneys associated with

interstitial inflammatory cell infiltration and fibroblastic reaction.

Pelvic epithelial hyperplasia in the kidneys of one female as well as epithelial hyperplasia in

the urinary bladder of one male were also observed.

However, the treatment-related histopathological changes observed in the recovery group

indicated a clear trend of recovery when comparing their extent and severity with the high

dose main group.

On the basis of the above mentioned results, the NOAEL (No Observed Adverse Effect

Level) for males and females could be considered 250 mg/kg/day.

Endpoint conclusion
Dose descriptor:
NOAEL
250 ng/kg bw/day
Study duration:
subchronic
Species:
rat
Quality of whole database:
satisfying

Additional information

Justification for classification or non-classification

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