Praseodymium(III,IV) oxide

Repeated dose toxicity: oral

Administrative data

Endpoint:

short-term repeated dose toxicity: oral

Remarks:

combined repeated dose and reproduction / developmental screening

Type of information:

experimental study

Adequacy of study:

key study

Study period:

6 August 2012 - 1 November 2012

Reliability:

1 (reliable without restriction)

Rationale for reliability incl. deficiencies:

other: Study conducted to GLP in compliance with agreed protocols, with no or minor deviations from standard test guidelines and/or minor methodological deficiencies, which do not affect the quality of the relevant results.

Data source

Reference

Reference Type:

study report

Title:

Unnamed

Year:

2013

Materials and methods

Test guideline

Qualifier:

according to guideline

Guideline:

OECD Guideline 422 (Combined Repeated Dose Toxicity Study with the Reproduction / Developmental Toxicity Screening Test)

Deviations:

no

GLP compliance:

yes

Limit test:

no

Test material

Test material information

Constituent 1

Reference substance name:

Praseodymium(III,IV) oxide

EC Number:

234-857-9

EC Name:

Praseodymium(III,IV) oxide

Cas Number:

12037-29-5

Molecular formula:

O11Pr6

IUPAC Name:

tetrakis(λ⁴-praseodymium(4+)) dipraseodymium(3+) undecaoxidandiide

Test material form:

solid: particulate/powder

Details on test material:

- Appearance: Brown powder
- Storage Conditions: Ambient, protected from light

Test animals

Species:

rat

Strain:

Sprague-Dawley

Sex:

male/female

Details on test animals or test system and environmental conditions:

TEST ANIMALS
- Strain: Crl:CD (SD)
- Age at study initiation: On dispatch from the supplier, the males were approximately 5 - 6 weeks old and the females were approximately 4 - 6 weeks old. At initiation of dosing, the animals were approximately 8 - 10 weeks old.
- Weight at study initiation: On dispatch from the supplier, the males weighed 158 - 200 g and the females weighed 112 - 156 g. At initiation of dosing, the animals weighed 330 - 460 g for males and 192 - 269 g for females.
- Housing: Animals were housed in cages, suspended on a series of racks. Male and female cages were racked separately. Animals were housed in polycarbonate cages with stainless steel grid tops and solid bottoms, with approximate dimensions of 61 x 43.5 x 24 cm. Cages were suspended on moveable racks and fitted with water bottles and integrated stainless steel food hoppers. The animals were initially housed 2 or 3 per cage. A few days prior to pairing for mating, males were transferred to individual cages with a stainless steel grid insert. After mating, the males were re-housed with their original cage mates. Mated females were transferred to individual solid bottom cages. White paper tissue was supplied as nesting material from Day 20 of gestation.
- Acclimation period: 27 days.
- Source: Charles River Limited, Margate, Kent, UK.
- Diet (e.g. ad libitum): Rat and Mouse Breeder Diet No. 3 (Expanded), supplied by Special Diets Services Limited, Witham, Essex, UK.
- Water (e.g. ad libitum): Water taken from the public supply (Scottish Water, Edinburgh, UK).

ENVIRONMENTAL CONDITIONS
- Temperature (°C): 19 - 23 °C
- Humidity (%): 40 - 70 %
- Air changes (per hr): A minimum of 10 air changes per hour.
- Photoperiod (hrs dark / hrs light): A 12 hour light/dark cycle was in operation.

IN-LIFE DATES: From: 27 August 2012 To: 12 October 2012

Administration / exposure

Route of administration:

oral: gavage

Vehicle:

other: 0.5 % w/v Methylcellulose

Details on oral exposure:

PREPARATION OF DOSING SOLUTIONS
The dosing formulations were prepared weekly, stored at ambient temperature and dispensed daily. All formulations were used within the 8 day stability period that was established previously at the testing laboratory.
The dosing formulations were stirred for at least 30 minutes prior to and throughout dosing.

VEHICLE
- Concentration in vehicle: 10, 30 or 100 mg/mL for the 100, 300 and 1000 mg/kg/day dose levels, respectively.
- Dose volume: 10 mL/kg. The volume administered to each animal was determined on each day by the weight of that animal recorded immediately prior to dosing, except during late gestation; from Day 16 of gestation until when parturition was complete, the dose volume was determined by the weight of the animal on Day 16 of gestation.

Analytical verification of doses or concentrations:

yes

Details on analytical verification of doses or concentrations:

ANALYTICAL METHOD
Analyses were performed by ICP-Optical Emission Spectroscopy using a validated analytical procedure. Samples to be analysed were transferred at ambient temperature to the analytical laboratory.

CONCENTRATION AND HOMOGENEITY ANALYSIS
Duplicate 0.5 mL samples were collected from the top, middle and bottom (duplicate middle only for control) from each formulation at each sampling time point (control samples not collected at Weeks 5 and 6) and were sent to the analytical laboratory.
Additional 0.5 mL triplicate samples were collected from the top, middle and bottom (triplicate middle only from control) from each formulation at each sampling time point (control samples not collected at Weeks 5 and 6) and were retained as back-up samples.
The results of the sample concentration were considered acceptable if they were within ± 10 % of theoretical concentration. For homogeneity, the criterion for acceptability was a relative standard deviation (RSD) of concentration of ≤10 % for each group.

Duration of treatment / exposure:

The males were dosed for 4 weeks, starting 2 weeks prior to mating. The females were dosed 2 weeks prior to mating, throughout mating, gestation and through to at least day 4 of lactation.
Several females were not dosed on their respective days of parturition due to the animals starting to give birth prior to the commencement of dosing on that day.

Frequency of treatment:

Once daily

Doses / concentrations

Doses / concentrations 1

Dose / conc.:

100 mg/kg bw/day (actual dose received)

Doses / concentrations 2

Dose / conc.:

300 mg/kg bw/day (actual dose received)

Doses / concentrations 3

Dose / conc.:

1 000 mg/kg bw/day (actual dose received)

No. of animals per sex per dose:

10 animals per sex per dose.

Control animals:

yes, concurrent vehicle

Details on study design:

- Dose selection rationale: The dose levels were agreed after a review of existing relevant toxicological data, including a 14 day dose range finding study conducted at the testing facility in which dose levels up to 1000 mg/kg/day produced no adverse reaction to treatment.

Examinations

Observations and examinations performed and frequency:

CAGE SIDE OBSERVATIONS: Yes
- Time schedule: Mortality/Moribundity checks on all animals were carried out early morning and as late as possible each day.

DETAILED CLINICAL OBSERVATIONS: Yes
- Time schedule: Once each week starting in pre-trial, all animals received a detailed clinical examination. All animals were examined for reaction to treatment at approximately hourly intervals up to 4 hours post dose on each day of dosing.

DETAILED FUNCTIONAL OBSERVATIONS (PARTIAL EXAMINATIONS)
Once during the pre-treatment period (week -1) and weekly thereafter, a more detailed examination was made on all animals.
- Cageside Observations: prostration, lethargy, writhing, circling, breathing abnormalities, gait abnormalities, tremor, fasciculation, convulsions, biting (of cage components or self mutilating), vocalisations, piloerection, ease of removal from the cage, body temperature, condition of the coat, presence of salivation and overall ease of handling.
- Condition of the eyes, checked for: pupillary function (reaction to visual stimulus), miosis, mydriasis, exophthalmos, encrustation and lacrimation.

Observations in a standardised area (2 min observation): latency (time to first locomotory movement), level of mobility, rearing, grooming, urination/defecation, arousal (level of alertness), posture (tremor/convulsions, vocalisation, piloerection), palpebral closure, gait abnormalities and stereotypy (excessive repetition of behaviours) and/or unusual behaviours.

BODY WEIGHT: Yes
- Time schedule for examinations: Bodyweights were recorded weekly for the three weeks preceding the start of treatment. From the start of treatment, the individual bodyweights were recorded daily.

FOOD CONSUMPTION: Yes
- Time schedule for examinations: Food consumption was measured for both sexes weekly, starting 3 weeks prior to dosing until pairing for mating. After pairing, the female food consumption was measured over Days 0 - 7, 7 - 14 and 14 - 20 of gestation and Days 0 - 4 of lactation. Recording of male food consumption did not recommence after pairing for mating.

FOOD EFFICIENCY: No

WATER CONSUMPTION: No

OPHTHALMOSCOPIC EXAMINATION: Yes
- Time schedule for examinations: Ophthalmoscopic examinations were taken pre-trial for all animals, in Week 4 for all male animals and shortly prior to sacrifice for all female animals.
The eyes were examined using an indirect ophthalmoscope after the application of a mydriatic agent (1 % Tropicamide, Mydriacyl). The following areas were evaluated: anterior, lenticular and fundic areas.

HAEMATOLOGY: Yes
- Time schedule for collection of blood: Samples were obtained during Week 4 for males and on or after Day 4 of lactation for females. Samples were collected via the tail vein. 0.5 mL of blood was then transferred into plastic tubes containing EDTA as an anticoagulant.
- Anaesthetic used for blood collection: No data
- Animals fasted: No
- How many animals: 5 per group; males, the first 5 animals per group and females the first 5 animals in each group to rear their litter to Day 3 of lactation.
- Parameters evaluated: red blood cell count, haemoglobin, haematocrit, mean cell volume, red cell distribution width, mean cell haemoglobin concentration, mean cell haemoglobin, reticulocyte count, platelet count, white blood cell count, neutrophil count, lymphocyte count, monocyte count, eosinophil count, basophil count and large unclassified cells.

COAGULATION: Yes
- Time schedule for collection of blood: Samples were obtained during Week 4 for males and on or after Day 4 of lactation for females. Samples were collected via the tail vein. Blood samples (0.9 mL) were taken into tubes containing 0.1 mL trisodium citrate (3.8 % (w/v)). The final sample volume was as close as possible to 1.0 mL to give a final concentration of 0.38 % (blood to citrate ratio of 9:1). The citrated blood samples were centrifuged and the plasma separated into plastic tubes and analysed.
- Anaesthetic used for blood collection: No data
- Animals fasted: No
- How many animals: 5 per group; males, the first 5 animals per group and females the first 5 animals in each group to rear their litter to Day 3 of lactation.
- Parameters evaluated: activated partial thromboplastin time, fibrinogen and prothrombin time.

CLINICAL CHEMISTRY: Yes
- Time schedule for collection of blood: Samples were obtained during Week 4 for males and on or after Day 4 of lactation for females. Samples were collected via the tail vein. 1.0 mL of blood was transferred into tubes containing lithium heparin and the samples centrifuged. The plasma was then examined for the appropriate parameters.
- Anaesthetic used for blood collection: No data
- Animals fasted: No
- How many animals: 5 per group; males, the first 5 animals per group and females the first 5 animals in each group to rear their litter to Day 3 of lactation.
- Parameters evaluated: urea, glucose, aspartate aminotransferase, alanine aminotransferase, alkaline phosphatase, creatine phosphokinase, lactate dehydrogenase, sodium, potassium, chloride, total protein, albumin, globulin, albumin/globulin ratio, cholesterol, creatinine, total bilirubin, calcium and phosphate.

URINALYSIS: No

NEUROBEHAVIOURAL EXAMINATION: Yes
FUNCTIONAL TESTS (FULL EXAMINATION)
The following additional functional assessments were performed on 5 males per group during Week 4 and on 5 females per group during lactation prior to blood sampling.
Reaction to sudden sound (click above the head), reaction to touch on the rump with a blunt probe, grip strength, pain perception, landing foot splay, motor activity and any other abnormality not already recorded in the above screening battery.

Sacrifice and pathology:

GROSS PATHOLOGY: Yes
- Terminal Procedures: The males were killed when mating was completed and the animals had been dosed for at least 4 weeks. The females and litters were killed between Day 5 and 7 of lactation. Animals were killed by exposure to carbon dioxide followed by exsanguination.
- Necropsy: All adult animals were subjected to a complete necropsy examination which included evaluation of the carcass and musculoskeletal system, all external surfaces and orifices, cranial cavity and external surfaces of the brain and thoracic, abdominal, and pelvic cavities with their associated organs and tissues. The reproductive tracts of all females were examined.
The following organs were weighed at necropsy for all adult animals before sampling and preservation: brain, epididymis, adrenal gland, pituitary gland, prostate gland, thyroid gland, heart, kidney, liver, lung, ovary, spleen, testis, thymus and uterus.

HISTOPATHOLOGY: Yes
Representative samples of the following tissues were collected from all adult animals and preserved as appropriate: animal identification (microchip), aortic artery, bone marrow smear, bone marrow (femur and sternum), femur (bone), rib (bone), sternum (bone), brain, cervix, epididymis, eye, adrenal gland, harderian gland, lacrimal gland, mammary gland, parathyroid gland, pituitary gland, prostate gland, salivary gland, seminal vesicle gland, thyroid gland, gross lesions, gut-associated lymphoid tissue, heart, kidney, large intestine (caecum, colon and rectum), larynx, liver, lung, lymph node (mandibular and mesenteric), skeletal muscle, nasal cavity, optic nerve, sciatic nerve, oesophagus, ovary, oviduct, pancreas, pharynx, skin, small intestine (duodenum, ileum and jejunum), spinal cord, spleen, stomach, testis, thymus, tongue, trachea, ureter, urinary bladder, uterus and vagina.
Histopathological evaluation of all tissues were undertaken for the 5 selected males and females in the control and high dose groups (the same animals that were used for the laboratory investigations).

Other examinations:

As part of the reproductive / developmental portion of the study, observations were also carried out on mating procedure, reproductive and viability indices were calculated, females with litters were observed during lactation and at the end of the study, pups were sacrificed and necropsied.

Statistics:

Where required to assist with interpretation, tests were applied to determine the statistical significance of observed differences between control and groups receiving test material. Unless otherwise stated, all statistical tests were two-sided and performed at the 5 % significance level using in-house software. Pairwise comparisons were only performed against the control group.

Bodyweight and food consumption data, haematology, coagulation, clinical chemistry and selected FOB and motor activity data was analysed for homogeneity of variance using the ‘F Max' test. If the group variances appeared homogeneous, a parametric ANOVA was used and pairwise comparisons were made using Fisher’s F protected LSD method via Student's t test; i.e., pairwise comparisons were made only if the overall F test was significant. If the variances were heterogeneous, log or square root transformations were used in an attempt to stabilise the variances. If the variances remained heterogeneous, then a Kruskal-Wallis non-parametric ANOVA was used and pairwise comparisons were made using chi squared protection (via z tests, the non-parametric equivalent of Student's t test).

Organ weight data was analysed as above, and by analysis of covariance (ANCOVA) using terminal bodyweight as the covariate.

Results and discussion

Results of examinations

Clinical signs:

effects observed, treatment-related

Description (incidence and severity):

see "Details on Results"

Mortality:

mortality observed, treatment-related

Description (incidence):

see "Details on Results"

Body weight and weight changes:

effects observed, treatment-related

Description (incidence and severity):

see "Details on Results"

Food consumption and compound intake (if feeding study):

effects observed, treatment-related

Description (incidence and severity):

see "Details on Results"

Food efficiency:

not examined

Water consumption and compound intake (if drinking water study):

not examined

Ophthalmological findings:

no effects observed

Haematological findings:

effects observed, treatment-related

Description (incidence and severity):

see "Details on results".

Clinical biochemistry findings:

effects observed, treatment-related

Description (incidence and severity):

see "Details on results".

Urinalysis findings:

not examined

Behaviour (functional findings):

no effects observed

Organ weight findings including organ / body weight ratios:

no effects observed

Gross pathological findings:

no effects observed

Histopathological findings: non-neoplastic:

no effects observed

Histopathological findings: neoplastic:

no effects observed

Details on results:

CLINICAL SIGNS AND MORTALITY
There were no unscheduled deaths during the course of this study.
There were no clinical signs during the course of the study that were considered to be related to administration of the test material.
Animal 60 (female, 100 mg/kg/day) was not dosed on Days 2 - 4 of lactation, prior to scheduled euthanasia on Day 5 of lactation, due to displaying signs of pale skin and eyes, walking on tiptoes, dark/red discharge from the vagina, weight loss, staining on fur (ventral abdomen), body hunched and piloerection.
Animal 68 (female: 300 mg/kg/day) was not dosed on Day 1 of lactation due to displaying signs of pale skin and eyes, walking on tiptoes, staining on fur (ventral abdomen), abnormal vocalisation (in hand of technician) and body hunched. At Day 2 of lactation, the animal suffered a total litter loss, however these signs were no longer present and the animal was dosed Days 2 - 4 of lactation, prior to scheduled euthanasia on Day 5 of lactation.
These signs were considered to be related to difficulties experienced by these animals during parturition and/or the early part of lactation, and were considered to be incidental given their low absolute incidence and the absence of similar signs in animals receiving 1000 mg/kg/day.

BODYWEIGHTS AND BODYWEIGHT CHANGES
Bodyweight gains were similar between control animals and animals receiving the test material.
Mean bodyweight gain was slightly higher than controls in females receiving 1000 mg/kg/day over gestation Day 0 - 20 (approximately 13 % more than the control group mean); however this was considered to be incidental given the considerable inter-animal variation within both the control and 1000 mg/kg/day dose groups.

FOOD CONSUMPTION
Food consumption was similar between control animals and animals receiving the test material.
Mean food consumption was noted to be higher than the controls in all dose groups between Days 0 - 4 of lactation; however this was considered incidental and a consequence of a slightly low mean control group value.

OPHTHALMIC EXAMINATIONS
There were no ophthalmic findings that were considered to be related to administration of the test material.

FUNCTIONAL OBSERVATION BATTERY PARAMETERS
Neurotoxicity Clinical Observations:
There were no neurotoxicity clinical observations during the course of the study that were considered to be related to administration of the test material.

Detailed Functional Observations:
Detailed functional observations in animals receiving the test material were similar to control animals throughout the course of the study. Any slight intergroup differences were considered to be too minor in magnitude to be attributed to the test material.

Motor activity:
Motion activity values in animals receiving the test material were similar to control animals. Any slight intergroup differences were considered to be too minor and/or transient to be attributed to the test material.

HAEMATOLOGY AND COAGULATION
In females during lactation, mean haemoglobin concentration, red blood cell counts and haematocrit were slightly lower than concurrent control at all dose levels (as low as 88 % of control mean at 1000 mg/kg/day), with a dose response relationship apparent in mean haemoglobin and haematocrit values. In addition, mean red cell distribution width values and reticulocyte counts were slightly higher than the concurrent control in these animals (reticulocytes as high as 18 % more than the control mean at 1000 mg/kg/day), with a dose response relationship apparent in mean red cell distribution width values. Although the changes were small in magnitude, many of the mean values were outside of the historical background control range, including all of these parameters at the highest dose level of 1000 mg/kg/day. The selected data is summarised in Table 1.
These changes were not apparent in males receiving the test material.
All other apparent differences in haematological or coagulation parameters were considered to be incidental due to large intra-group variation and/or the absence of a clear dose-response relationship.

CLINICAL CHEMISTRY
Mean lactate dehydrogenase activities were lower than the concurrent control in a dose dependent manner in males at Week 4 (82, 79 and 65 % of control mean, at 100, 300 and 1000 mg/kg/day, respectively), and in females during lactation (87, 78 and 63 % of control mean, at 100, 300 and 1000 mg/kg/day, respectively), attaining statistical significance in males receiving 300 and 1000 mg/kg/day. These values were lower than the historical background control range, at all dose levels in females and at 1000 mg/kg/day in males.
In females during lactation, mean phosphate levels were higher than the concurrent control at 300 and 1000 mg/kg/day (36 and 64 % higher than the control mean, respectively), attaining statistical significance and being higher than the historical background control range at 1000 mg/kg/day. Mean calcium levels were also very slightly high at 300 and 1000 mg/kg/day (5 and 8 % higher than the control mean, respectively), achieving statistical significance at 1000 mg/kg/day, however all groups (including the control) were above the historical background control range.
The selected data is summarised in Table 2.
All other apparent differences in clinical chemistry parameters were considered to be incidental due to large intra-group variation and/or the absence of a clear dose response relationship.

ORGAN WEIGHTS
No test-material related organ weight changes were noted. There were isolate organ weight values that were different from their respective controls. There were, however, no patterns, trends or correlating data to suggest these values were toxicologically relevant. Thus, the organ weight differences observed were considered incidental and unrelated to administration of the test material.

GROSS PATHOLOGY
No test-material related gross findings were noted. The gross findings observed were considered incidental, of the nature commonly observed in this strain and age of rat on this type of study, and/or were of similar incidence in control and treated animals and, therefore, were considered unrelated to administration of the test material.

HISTOPATHOLOGY
Minimal granulomas containing pigmented material were observed in the lungs of two animals given 1000 mg/kg/day (32 Male and 71 Female). This was considered to be some type of foreign inhaled material, presumably due to reflux of test material during the dosing procedure.
Other microscopic findings observed were considered incidental, of the nature commonly observed in this strain and age of rat on this type of study, and/or were of similar incidence and severity in control and treated animals and, therefore, were considered unrelated to administration of the test material.
Thymic atrophy and gastritis in females were considered to be secondary to stress associated with parturition.

Effect levels

Dose descriptor:

NOAEL

Effect level:

1 000 mg/kg bw/day (nominal)

Based on:

test mat.

Sex:

male/female

Basis for effect level:

other: see 'Remark'

Target system / organ toxicity

Critical effects observed:

not specified

Any other information on results incl. tables

Table 1: Selected Haematology Values For Females During Lactation - Group Mean Values

Dose level (mg/kg/day)		Hb (g/dL)	RBC (x 10¹²/L)	Hct (L/L)	RDW (%)	Ret (x 10⁹/L)
0	Mean SD N	12.9 0.3 5	7.01 0.38 5	0.411 0.007 5	14.9 0.5 5	440 38 5
100	Mean SD N	12.3 0.4 5	6.53 0.77 5	0.395 0.017 5	15.6 1.8 5	464 125 5
300	Mean SD N	12.2 0.6 5	6.58* 0.17 5	0.386 0.016 5	16.2 0.9 5	454 78 5
1000	Mean SD N	11.4** 0.9 5	6.31* 0.56 5	0.363*** 0.028 5	16.9 1.6 5	520 112 5

Significantly different from the controls: * = p<0.05; ** = p<0.01; *** = p< 0.001

Hb = Haemoglobin; RBC = Red Blood Cell Count; Hct = Haematocrit; RDW = Red Cell Distribution Width; Ret = Reticulocyte Count.

 

Table 2: Selected Clinical Chemistry Data - Group Mean Values

Dose level (mg/kg/day)		Males (Week 4)			Females (During Lactation)
		LDH (U/L)	Phos (mmol/L)	Ca (mmol/L)	LDH (U/L)	Phos (mmol/L)	Ca (mmol/L)
0	Mean SD N	157 24 5	2.42 0.28 5	2.72 0.04 5	202 26 5	1.51 0.49 5	2.82 0.07 5
100	Mean SD N	129 27 5	2.43 0.23 5	2.72 0.07 5	176 65 5	1.42 0.36 5	2.84 0.05 5
300	Mean SD N	124* 22 5	2.53 0.08 5	2.75 0.03 5	157 23 5	2.05 0.51 5	2.96 0.08 5
1000	Mean SD N	102** 21 5	2.52 0.17 5	2.75 0.09 5	127 33 5	2.48** 0.52 5	3.05** 0.21 5

Significantly different from the controls: * = p<0.05; ** = p<0.01

LDH = Lactate Dehydrogenase; Phos = Phosphate; Ca = Calcium

Applicant's summary and conclusion

Conclusions:

Under the conditions of this study the No Observed Adverse Effect Level (NOAEL) in Sprague-Dawley rats was considered to be 1000 mg/kg/day.

Executive summary:

A combined repeated dose toxicity study with reproduction/developmental toxicity screening test was carried out in order to assess the test material in accordance with the standardised guideline OECD 422.

Three groups of 10 male and 10 female Sprague-Dawley rats of the Crl:CD(SD) strain were dosed once daily, by oral gavage, with the test material at dose levels of 100, 300, or 1000 mg/kg/day. Another group of 10 male and 10 female rats were dosed with the vehicle (0.5 % w/v methylcellulose) following the same dosing regimen as the treated animals and were used as controls.

The males were treated for 2 weeks prior to mating, then through mating, until the day prior to necropsy (ca 4 weeks of treatment). Females were treated for 2 weeks prior to mating, then through mating, gestation and until at least Day 4 of lactation (ca. 6 weeks of treatment). 

The following parameters and end points were evaluated: clinical signs, bodyweights, bodyweight changes, food consumption, ophthalmology, detailed functional tests and observations, clinical pathology parameters (haematology, coagulation and clinical chemistry), gross necropsy findings, organ weights, and histopathological examinations, mating and pregnancy performance, fertility, maternal care and pup performance (litter survival and pup weights).

Treatment at levels up to 1000 mg/kg/day was not associated with any changes in clinical observations, body weight gain, food consumption, ophthalmoscopy findings, neurotoxicity findings, coagulation, organ weights, gross necropsy findings or histopathogical findings.

At all dose levels in females during lactation, mean haemoglobin concentration, red blood cell counts and haematocrit were slightly lower than the concurrent control, whilst mean red cell distribution width values and reticulocyte counts were slightly higher. 

Mean lactate dehydrogenase activities were lower than the control at all dose levels in males at Week 4 of study and in females during lactation. Mean phosphate and calcium levels were also higher than the control during lactation in females receiving 300 or 1000 mg/kg/day. 

Given that these clinical pathology changes were without pathological correlates and that the changes did not manifest themselves clinically, they were considered not to be adverse.

Under the conditions of this study the No Observed Adverse Effect Level (NOAEL) in Sprague-Dawley rats was considered to be 1000 mg/kg/day for both males and females.