Sodium chlorite

Repeated dose toxicity: oral

Administrative data

Endpoint:

sub-chronic toxicity: oral

Type of information:

experimental study

Adequacy of study:

key study

Reliability:

1 (reliable without restriction)

Rationale for reliability incl. deficiencies:

guideline study

Cross-reference

Reason / purpose for cross-reference:

other: Range-finding study

Reference

Endpoint:

short-term repeated dose toxicity: oral

Remarks:

14 day range-finding experiment

Type of information:

experimental study

Adequacy of study:

supporting study

Reliability:

1 (reliable without restriction)

Rationale for reliability incl. deficiencies:

other: Range-finding study for a 90-day oral toxicity study.

Reason / purpose for cross-reference:

other: The present study is the range finding for the main study:

Principles of method if other than guideline:

Range-finding study.

GLP compliance:

yes

Species:

rat

Strain:

other: Crl:CD(SD) BR strain (VAF plus)

Sex:

male/female

Details on test animals or test system and environmental conditions:

TEST ANIMALS
- Source: Charles River (UK) Limited, Margate, England.
- Weight at study initiation: At start of the main study the males weighted 131-176 g and females 122-158 g
.- Housing: In groups of 5, by sex, in grind bottomed stainless steel cages measuring 56x35x20 cm, suspended over cardboard lined extreta trays.
- Diet (e.g. ad libitum): With the exceptions of periods of deprivation associated with laboratory investigations, ad libitum.
- Water (e.g. ad libitum): With the exceptions of periods of deprivation associated with laboratory investigations, ad libitum.
- Acclimation period: 14 days.

DETAILS OF FOOD AND WATER QUALITY:Certificate of analysis for the batches of diet used during the study and a representative analytical report for the water supply are available.

ENVIRONMENTAL CONDITIONS
- Temperature (°C): 10- 22ºC. The room temperature was at the lower extreme for several very short periods only and was not associated with any health problems in the animals.
- Humidity (%): 29-55 %.
- Photoperiod (hrs dark / hrs light): 12 hours light/12 hours dark

Route of administration:

oral: gavage

Vehicle:

water

Details on oral exposure:

PREPARATION OF DOSING SOLUTIONS: The test article was formulated freshly each day in the vehicle

Analytical verification of doses or concentrations:

yes

Details on analytical verification of doses or concentrations:

Dosing solutions were assayed for sodium chlorite content by titration with standard sodium thiosulphate, of the iodine released by treatment with potassium iodide and hydrochloric acid.Samples of each formulation prepared on one day of dosing in week 1 (including the control formulation) were analyzed for test article. The analysis results demonstrated accurate preparation of the dosing formulations (99.5 to 107.6 % of theoretical).Moreover, a 14 day stability trial was conducted during the study. It confirmed that the test article is stable in the vehicle (100.2-107.6 % recoveries) . As the test article forms a true solution, homogeneity assesment was considered unnecessary.

Duration of treatment / exposure:

14 days

Frequency of treatment:

Once daily

Dose / conc.:

0 mg/kg bw/day (nominal)

Dose / conc.:

25 mg/kg bw/day (nominal)

Dose / conc.:

50 mg/kg bw/day (nominal)

Dose / conc.:

100 mg/kg bw/day (nominal)

Dose / conc.:

200 mg/kg bw/day (nominal)

No. of animals per sex per dose:

5/sex/dose group

Control animals:

yes

Details on study design:

- Dose selection rationale: A preliminary tolerance study was performed during acclimatisation period. Animals were dosed as for the main study as follows: One male and one female were dosed once at 1000 mg/kg-bw/day. Both animals were sacrificed in extremis approximately half and hour after dosing. Clinical signs were prostration, pale extremities, and slow laboured breathing. At necropsy the blood was found to be brown coloured. Two males and two females were dosed for up to three days. One female was found dead and one was female sacrificed in extremis on the second day. Clinical signs were pale extremities, piloerection, hutched posture, blood stained urogenital area/bedding. Necropsy findings were autolysis and fluid distension of stomach for the female found dead. Pale liver and blood in urinary bladder were recorded for the sacrificed female. Both males survived the 3 day dosing period. One male was observed with piloerection on the first and second days. Male bodyweight gain was normal and no abnormalities were found at necropsy.Two males and two females were dosed for three days. All animals survived. No clinical signs of toxicity were observed and bodyweights were normal. No abnormalities were observed at necropsy.Based on these effects, 25, 50, 100 and 200 were selected for the range finding study (14 days).

Observations and examinations performed and frequency:

Mortality checks were performed twice daily.

DETAILED CLINICAL OBSERVATIONS: Yes
- Time schedule: Daily for changes in condition or behaviour.

BODY WEIGHT: Yes
- Time schedule for examinations: On the first day of dosing, twice weekly thereafter and at necropsy.FOOD CONSUMPTION AND COMPOUND INTAKEThe food consumed by each cage of animals was recorded weekly.

OPHTHALMOSCOPIC EXAMINATION: No

HAEMATOLOGY: Yes
- Time schedule for collection of blood: Blood samples were taken from all animals in week 2. Blood samples were obtained twice, by puncture of the retro-orbital sinus, under light ether anaesthesia; one sample was taken 4 hours after dosing for mathaemoglobin level only and a second sample was taken 24 hours after dosing and after overnight fast.- Anaesthetic used for blood collection: Yes, ether- Animals fasted: Yes - How many animals: All.- The following parameters were analyzed: Haemoglobin, red blood cell count, packed cell volume, mean cell volume, mean cell haemoglobin concentration, total leucocyte count, platelet count, methaemoglobin level, cell morphology, leucocyte differential count.

CLINICAL CHEMISTRY: No

URINALYSIS: Yes
- Time schedule for collection of urine: Overnigth under water deprivation from all animals in week 2.
- Metabolism cages used for collection of urine: Not specified
- Animals fasted: Not specified.
- The following parameters were checked: Volume, colour, specific gravity, pH, protein, glucose, ketones, bilirubin, blood pigments, urobilinogen, erythrocytes, leucocytes, crystals, debris and casts.

NEUROBEHAVIOURAL EXAMINATION: No

IMMUNOLOGY: No

Sacrifice and pathology:

GROSS PATHOLOGY: Yes, all animals were weighed and examined externally, a macroscopic examination was then performed by opening the cranial, thoracic and visceral cavities and by observing the appearance of the tissues in situ. The location, colour, shape and size of any abnormalities were recorded. The following organs, dissected from fat and other contiguous tissue, were weighed: adrenals, brain, heart, kidneys, liver, ovaries, spleen, testes and thymus.

HISTOPATHOLOGY: No, either whole organs or samples of the following tissues were preserved in buffered formol saline:All gross lesions, kidneys, liver. In the first instance the above tissues were not processed further for histopathology.

Statistics:

For those responses that have historically been found to have normally distributed errors, Bartlett’s was used to check that the variances for all groups are homogenous. If the group variances appeared to increase in proportion to the group means, a log transformation was carried out and Bartlett’s test repeated. If significance was found at 5% level non-parametric methods was used. For those responses where Bartlett’s test was not significant a one-way analysis of variance (ANOVA) was used to test whether there was evidence of any difference in the group means. If a significant ANOVA was found the differences were determined using t-tests carried out between the control group and the treatment groups. Non-parametric testing was carried out using a Krustal-Wallis test for between group differences. If a significant between group differences was found, Wilcoxon rank sum test was used to compare the treatment group with the control group.

Clinical signs:

effects observed, treatment-related

Description (incidence and severity):

Treatment-related clinical signs were observed at 200mg/kg/day only. One male was found dead on day 3 was observed with pale eyes and extremities, a small amount of blood around the urogenital area, slight incoordination and piloerection. One female when found dead, was observed with blood around the vulva.

Mortality:

mortality observed, treatment-related

Description (incidence):

At 200mg/kg/day, one female was found dead on day 2 and one male and one female were found dead on day 3. Because of the high mortality in this group all survivors, although appearing healthy, were sacrificed on day 3; consequently bodyweights, food consumption, haematology and urinalysis were not assessed for this group.There were no deaths at 25, 50 or 100 mg/kg/day.

Body weight and weight changes:

not examined

Description (incidence and severity):

Bodyweight gain for both males and females at 100 mg/kg/day was statistically significantly less (Student's t test) than the control values. Male bodyweight gain at 50mg/kg/day was also less than the control values, but the difference was not statistically significant.Bodyweight gain for males and females at 25mg/kg/day and females at 50mg/kg/day was similar to the control values.

Food consumption and compound intake (if feeding study):

not specified

Description (incidence and severity):

The amount of food consumed by males at 50 and 100 mg/kg/day and females at 25, 50 and 100 mg/kg/day was slightly less than the control values, but still within normal ranges. Male food consumption at 25mg/kg/day was slightly greater than the control values.

Food efficiency:

not examined

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

not specified

Ophthalmological findings:

not specified

Haematological findings:

effects observed, treatment-related

Description (incidence and severity):

Changes considered to be treatment-related were observed for a number of parameters. At 100 mg/kg/day, red blood cell count, haemoglobin concentration and packed cell volume were decreased and total leucocyte count, neutrophil count and methaemoglobin levels (at both 4 and 24 hours after dosing) were increased in comparison with the control values. These changes occurred for both males and females and the majority of differences were statistically significant. At 100 mg/kg/day there was also a number of animals with gross or moderate red blood cell polychromasia and anisocytosis, Howell-Jolly bodies and punctate basophilia.At 25 and 50mg/kg/day, packed cell volume was reduced for both males and females; statistically significant differences (Student's t test) were observed for males at 25mg/kg/day and females at 50mg/kg/day. Occasionally differences for other parameters were observed but none of these was part of a dose-dependent pattern.

Clinical biochemistry findings:

not examined

Urinalysis findings:

effects observed, treatment-related

Description (incidence and severity):

Urine volume was statistically significantly increased (Student's t test) in comparison with the control group for both males and females at 100 mg/kg/day. Urine volume was also significantly increased for males at 25mg/kg/day, but this was not part of a dose-related trend as volume at 50mg/kg/day was similar to the control values. Urine volume for females at 25 and 50mg/kg/day was similar to the control values. Specific gravity was slightly reduced for males and females at 100 mg/kg/day. For individual animals with increased volume, the urine colour was pale.There were no differences for the remaining urinalysis parameters considered to be treatment-related.

Behaviour (functional findings):

not examined

Immunological findings:

not examined

Organ weight findings including organ / body weight ratios:

not specified

Description (incidence and severity):

There were no organ weight changes that followed a dose-related trend and were consistent for both males and females. Absolute kidney weight for males was statistically significantly less (Student's t test) than the control values at 50 and 100 mg/kg/day, but when analysed on a bodyweight-related basis, significance was achieved only for the 50mg/kg/day group. Bodyweight-related female kidney weight was, however, together with adrenal weight, significantly increased at 100 mg/kg/day. Male absolute liver weight was significantly less than the controls at 50 and 100 mg/kg/day and when liver was analysed on a bodyweight-related basis, significance was achieved at 25 and 100 mg/kg/day. A similar trend was not, however, apparent for the females.

Gross pathological findings:

effects observed, treatment-related

Description (incidence and severity):

Abnormalities considered to be treatment-related were observed at 200mg/kg/day. Brown coloured or brown tinged blood was observed for one mortality (female) and for two animals sacrificed prematurely (male female). Two female mortalities were found with blood stained fur around the urethral opening. Little or no food was present in the stomach of 2 females and, additionally, one of them had a pale coloured liver.Several animals from the other treated groups were observed with a swollen area and broken skin on the mouth. This minor lesion may have been caused by the dosing procedure, but its significance is not clear.

Neuropathological findings:

not examined

Histopathological findings: non-neoplastic:

not examined

Histopathological findings: neoplastic:

not examined

Other effects:

no effects observed

Key result

Dose descriptor:

LOAEL

Effect level:

100 mg/kg bw/day (nominal)

Based on:

test mat.

Sex:

male/female

Basis for effect level:

haematology

urinalysis

Remarks on result:

other:

Remarks:

At 25 and 50mg/kg/day there was little clear evidence of toxicity. Packed cell volume was slightly reduced at these levels and male bodyweight gain was slightly reduced at 50mg/kg/day, but these changes could not, in view of the small group size, be conclusively related to treatment.

Key result

Critical effects observed:

yes

Lowest effective dose / conc.:

100 mg/kg bw/day (nominal)

System:

haematopoietic

Treatment related:

not specified

Dose response relationship:

not specified

Relevant for humans:

not specified

Conclusions:

It was concluded that gavage adminsitration to the rat elicited toxicity at 200 and 100 mg/kg/day. At the highest dose level three out of ten animals died after two or three days and the survivors were sacrificed prematurely; necropsy findings included brown coloured blood and blood stained fur around the urethral opening. At 100mg/kg/day, bodyweight gain was reduced, certain haematological parameters were changed (red blood cell count, haemoglobin concentration and packed cell volume were decreased, total leucocyte count, neutrophil count and methaemoglobin levels were increased, gross or moderate red blood cell polychomasia and anisocytosis, Howell-Jolly bodies and punctate basophilia were observed) and urine volume was increased. At 25 and 50mg/kg/day there was little clear evidence of toxicity. Packed cell volume was slightly reduced at these levels and male bodyweight gain was slightly reduced at 50mg/kg/day, but these changes could not, in view of the small group size, be conclusively related to treatment.

Executive summary:

A range-finding study in rats was conducted for 14 days at the following doses 0, 25, 50, 100 and 200 mg/kg/day. These doses were selected based on a preliminary tolerance study. It was concluded that gavage adminsitration to the rat elicited toxicity at 200 and 100 mg/kg/day. At the highest dose level three out of ten animals died after two or three days and the survivors were sacrificed prematurely; necropsy findings included brown coloured blood and blood stained fur around the urethral opening. At 100mg/kg/day, bodyweight gain was reduced, certain haematological parameters were changed (red blood cell count, haemoglobin concentration and packed cell volume were decreased, total leucocyte count, neutrophil count and methaemoglobin levels were increased, gross or moderate red blood cell polychomasia and anisocytosis, Howell-Jolly bodies and punctate basophilia were observed) and urine volume was increased. At 25 and 50mg/kg/day there was little clear evidence of toxicity. Packed cell volume was slightly reduced at these levels and male bodyweight gain was slightly reduced at 50mg/kg/day, but these changes could not, in view of the small group size, be conclusively related to treatment.

Data source

Reference

Reference 1

Reference Type:

study report

Title:

Unnamed

Year:

1992

Reference 2

Reference Type:

publication

Title:

Subchronic Toxicity of Sodium Chlorite in the Rat

Author:

Harrington RM, Romano RR, Gates D and Ridgway

Year:

1995

Bibliographic source:

Journal of the American College of Toxicology, 14(1): 21-33

Materials and methods

Test guideline

Test guideline 1

Qualifier:

according to guideline

Guideline:

EPA OPP 82-1 (90-Day Oral Toxicity)

Deviations:

no

Test guideline 2

Qualifier:

equivalent or similar to guideline

Guideline:

OECD Guideline 408 (Repeated Dose 90-Day Oral Toxicity Study in Rodents)

Deviations:

no

GLP compliance:

yes

Limit test:

no

Test material

Test material information

Constituent 1

Reference substance name:

Sodium chlorite

EC Number:

231-836-6

EC Name:

Sodium chlorite

Cas Number:

7758-19-2

Molecular formula:

ClHO2.Na

IUPAC Name:

sodium chlorite

Test material form:

solid: flakes

Details on test material:

Lot/Batch number: ONF 28408B
Description: A flaky white material
Purity: 80.9 %
Stability: The test article is stable in the vehicle for at least 14 days

Test animals

Species:

rat

Strain:

Crj: CD(SD)

Sex:

male/female

Details on test animals or test system and environmental conditions:

- Source: Charles River (UK) Limited, Margate, England.
- Age at study initiation: 3-4 weeks old on arrival.
- Weight at study initiation: At the start of treatment for the main study the males weighed 120 - 164g and the females weighed 110 - 160 g.
- Housing: The animals were housed in groups of 5, by sex, in grid bottomed stainless steel cages measuring 56 x 35 x 20cm, suspended over cardboard lined excreta tray.
- Diet (e.g. ad libitum): With the exception of periods of deprivation associated with laboratory investigations, freely available throughout the study
- Water (e.g. ad libitum):With the exception of periods of deprivation associated with laboratory investigations, freely available throughout the study
- Acclimation period: 12 days

.DETAILS OF FOOD AND WATER QUALITY:The supplier's certificates of analysis for the batches of diet used during this study and a representative analytical report for the water supply are available.

ENVIRONMENTAL CONDITIONS
- Temperature (°C): 18 - 22ºC.
- Humidity (%): 36-67 %
- Photoperiod (hrs dark / hrs light): 12 hours light/12 hours dark.

Administration / exposure

Route of administration:

oral: gavage

Details on route of administration:

The test article was administered orally, by gavage, once daily, 7 days a week for 13 weeks until the day before necropsy. Dosing was performed using a rubber catheter attached to a plastic disposable syringe. Animals were dosed in group order.

Vehicle:

water

Details on oral exposure:

PREPARATION OF DOSING SOLUTIONS: The test item was formulated weekly.

Analytical verification of doses or concentrations:

yes

Details on analytical verification of doses or concentrations:

Dosing supensions were assayed for sodium chlorite content by titration, with standard sodium thiosulphate, of iodine released on treatment with iodine/hydrochloric acid.Prior to the start of the study, the batch of sodium chloride to be used was assayed in triplicate for purity. The mean concentration of sodium chlorite was 80.9 % (0.023 % CV) in the bulk material.A 14 day stability trial was conducted during the range-finding study, which confirmed that the test article is stable in the vehicle for at least 14 days.Analysis of formulations prepared for weeks 1, 6 and 13 of dosings gave recoveries, for all groups, of between 98 and 103 % of theoretical concentrations (corrected for purity at 80.9 %) indicating that formulations were accuterately prepared.

Duration of treatment / exposure:

13 weeks

Frequency of treatment:

Daily

Doses / concentrations

Doses / concentrations 1

Dose / conc.:

0 mg/kg bw/day (nominal)

Doses / concentrations 2

Dose / conc.:

10 mg/kg bw/day (nominal)

Doses / concentrations 3

Dose / conc.:

25 mg/kg bw/day (nominal)

Doses / concentrations 4

Dose / conc.:

80 mg/kg bw/day (nominal)

No. of animals per sex per dose:

15/sex/group

Control animals:

yes, concurrent vehicle

Details on study design:

- Dose selection rationale: Based on a range finding study. - Rationale for animal assignment (if not random): The required number of animals was obtained by weighing all animals and discarding those at the extremes of the weight range. Selected animals were then allocated randomly to groups using a stratified bodyweight procedure. - Section schedule rationale (if not random): Cage positions on the battery were allocated by starting at the top left hand corner of the battery and working left to right and top to bottom placing a cage from each group of males in ascending group order. The sequence was repeated until all the males were placed and was then performed similarly for females.

Examinations

Observations and examinations performed and frequency:

CLINICAL OBSERVATIONS: Yes
- Time schedule: Morbidity was checked twice daily at intervals of a minimum of 5 h on checks performed in day.

BODY WEIGHT: Yes
- Time schedule for examinations: On the first day of dosing, weekly thereafter and at necropsy.

FOOD CONSUMPTION:
- Time schedule: Weekly

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: Yes / No / No data

WATER CONSUMPTION: No

OPHTHALMOSCOPIC EXAMINATION: Yes - Time schedule for examinations: Both eyes of all animals were examined before treatment started. In week 12 the eyes of control and high dose animals were examined. Examinations were performed by a veterinary ophthalmologist using an indirect ophthalmoscope and then, if necessary, a direct ophthalmoscope after previous instillation of a mydriatic agent.

HAEMATOLOGY: Yes
- Time schedule for collection of blood: Week 13
- How many animals: All
- Parameters: haemoglobin concentration, red blood cell count, packed cell volume, mean cell volume, mean cell haemoglobin, mean cell haemoglobin concentration, total leucocyte count, platelet count, methaemoglobin level, leucocyte differential count, reticulocyte count, cell morphology.

CLINICAL CHEMISTRY: Yes
- Time schedule for collection of blood: Week 13
- How many animals: All- Parameters: Blood urea nitrogen, glucose, alanine aminotransferase, aspartate amino transferase, total protein, albumin, albumin/globulin ratio, sodium, potassium, calcium, chloride, inorganic phosphorous, bilirubin, creatinine, cholesterol, gamma glutamyl transferase.

URINALYSIS: Yes
- Time schedule for collection of urine: Week 13
- Parameters: volume, colour, specific gravity, pH, protein, glucose, ketones, bilirubin, urobilinogen, blood pigments, microscopic examination of sediment deposit: erythrocytes, leucocytes, crystals, debris and casts.

NEUROBEHAVIOURAL EXAMINATION: No

Sacrifice and pathology:

ORGAN WEIGHTS:- Organs studied: adrenals, kidneys, ovaries, testes, brain, liver, pituitary, thymus, heart, lungs, spleen, thyroids (weighed after fixation)

GROSS AND HISTOPATHOLOGY- Tissues: Adrenals, caecum, epidiymides, exorbital lacrimal gland*, jejunum, lungs, oesophagus, pituitary, salivary glands, skeletal muscle*, spleen, testes, thyroids, (incl. parathyroids) vagina*, aorta, colon, eyes* (incl. optic nerve) heart, kidneys, mammary gland (female only) ovaries, prostate*, seminal vesicles*, skin*, sternum (incl. marrow) thymus, urinary bladder, brain (4 levels) duodenum, femur* with articular surface, ileum, liver, mesenteric lymph nodes, pancreas, rectum, sciatic nerve, spinal cord* (3 levels) stomach, trachea*, uterus.*not examined in the first instance.

Statistics:

For those responses that have historically been found to have normally distributed errors, Bartlett’s was used to check that the variances for all groups are homogenous. If the group variances appeared to increase in proportion to the group means, a log transformation was carried out and Bartlett’s test repeated. If significance was found at 5% level non-parametric methods was used. For those responses where Bartlett’s test was not significant a one-way analysis of variance (ANOVA) was used to test whether there was evidence of any difference in the group means. If differences were found Dunnet’s multiple comparison T-test was also applied. For all data, William’s test was applied where, from an inspection of the means, a monotonic trend was apparent. Non-parametric testing was carried out using a Kruskall-Wallis test for between group differences. If differences were found, Dunn’s multiple comparison tests were also applied. For all data, Shirley’s test was applied where, from an inspection of the means, a monotonic trend was apparent. For Shirley’s and William’s tests significance levels were set at p< 0.05 and 0.01. For Dunnet’s and Dunn’s multiple comparison tests significance levels were set at p < 0.05, 0.01 and 0.001 for a two-sided risk.

Results and discussion

Results of examinations

Clinical signs:

effects observed, treatment-related

Description (incidence and severity):

CLINICAL SIGNS AND MORTALITYPoor clinical condition was observed in three animals at 80 mg/kg/day. One male had a hunched posture, ptosis and peribuccal staining during week 8. A further male was observed with a hunched posture on the day of terminal necropsy. From weeks 9 to 13, one female was observed with swollen hind limbs, hunched posture, hypoactivity and piloerection. Salivation either immediately before or after dosing was observed room the third week of dosing in all animals at 80 mg/kg/day. Salivation was also observed very occasionally for 2 males at 25 mg/kg/day. No treatment related clinical signs were observed 10 mg/kg/day.

Mortality:

mortality observed, treatment-related

Description (incidence):

There were 5 premature deaths during the study, one at 25 mg/kg/day and 4 at 80 mg/kg/day. The mortality at 25 mg/kg/day was accidental. One was killed during week 6 after biting and swallowing the dosing catheter. The mortalities at 80 mg/kg/day were considered likely to be treatment related. On the day of bleeding, shortly after sampling, one male was found dead and one was killed in extremis. Signs immediately before death included pale eyes and extremities and hypoactivity. One animal was observed in poor clinical condition during week 7. In both cases the animals were severely anaemic. Blood sampling may have exacerbated this problem and been a contributory factor to the cause of death.

Body weight and weight changes:

no effects observed

Description (incidence and severity):

Body weights were not affected by treatment.

Food consumption and compound intake (if feeding study):

no effects observed

Description (incidence and severity):

Food consumption was not affected by treatment.

Food efficiency:

not examined

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

not examined

Ophthalmological findings:

no effects observed

Description (incidence and severity):

There were no treatment related ocular changes.

Haematological findings:

effects observed, treatment-related

Description (incidence and severity):

At 80 mg/kg/day the group mean erythrocyte count in both males and females was statistically significantly less than the control values. In males, only haematocrit and haemoglobin levels were statistically significantly less and methaemoglobin levels and neutrophil counts were statistically significantly greater than the control values. The reticulocyte count was increased but was not statistically significant. The group mean changes reflected marked changes in these parameters for 2 individual males which died shortly after bleeding. Morphological changes included polychromasia, anisocytosis, poikilocytosis and presence of Howell-Jolly bodies and normoblasts were also observed. Morphological changes were also observed in the blood smears of three females at 80 mg/kg/day, including polychromasia, poikilocytosis, macrocytosis and neutrophilia. Most probably as a consequence of the increased neutrophil count in males at 80 mg/kg/day, lymphocyte counts were statistically significantly less than the control values. Where the primary red cell parameters (mean erythrocyte count, heamoglobin and haematocrit) were affected, associated changes were seen in the calculated parameters (mean cell volume, mean cell haemoglobin, Mean cell haemoglobin concentration). In both males and females at 25 mg/kg/day and males only at 10 mg/kg/day statistical trends highlighted a dose dependent downward trend for erythrocyte counts. Statistical significance was not confirmed by direct comparison with the control group and group mean values were within the background range, therefore a conclusive association with treatment could not be established.

Clinical biochemistry findings:

effects observed, treatment-related

Description (incidence and severity):

Total protein levels were decreased for both sexes at 80 mg/kg/day and males only at 25 mg/kg/day, but values were within the background range and an association with treatment was considered unlikely. The aspartate aminotransferase levels were elevated in 2 males which died or were killed in extremis and one female at 80 mg/kg/day. Additionally, one male had elevated bilirubin levels and the female, cholesterol levels were also elevated.

Urinalysis findings:

effects observed, treatment-related

Description (incidence and severity):

Urine volume was unusually high in 4 females at 80 mg/kg/day, urinary specific gravity was reduced in the affected animals. These changes were not associated with histopathological changes in the kidney.

Behaviour (functional findings):

not examined

Immunological findings:

not examined

Organ weight findings including organ / body weight ratios:

effects observed, treatment-related

Description (incidence and severity):

The weight of the spleen and adrenals occurred in both sexes. In males at 80 mg/kg/day spleen weight was statistically significantly greater than the control values when analysed on both an absolute and body weight related basis. In females, spleen weights were significantly greater than the control values at 10 and 80 mg/kg/day when analysed on an absolute basis and 25 and 80 mg/kg/day on a body weight related basis. Adrenal weights were significantly greater than the control values in males at 80 mg/kg/day only when analysed on a body weight related basis. Female adrenal weight was statistically significantly greater than the controls at 80 mg/kg/day when analysed on an absolute basis and at 25 and 80 mg/kg/day when analysed on a body weight related basis. It is possible that spleen weight increases at 80 mg/kg/day were secondary to the changes in erythrocyte characteristics. Other statistically significant differences were considered not to be treatment related.

Gross pathological findings:

effects observed, treatment-related

Description (incidence and severity):

In the stomach, changes including squamous epithelial hyperplasia with hyperkeratosis and ulceration, chronic inflammation and oedema, were seen in 7 males and 8 females at 80 mg/kg/day. Ulceration, chronic inflammation and oedema were seen in the stomach of two males at 25 mg/kg/day. Extramedullary haemopoiesis were seen in the spleen of 2 males and 2 females at 80 mg/kg/day and 1 animal from each of the 10 and 25 mg/kg/day. Although there was no correlation between the observation of extremedullary haemopoiesis and severity of anaemia or changes in organ weights, the increased incidence at 80 mg/kg/day was considered likely to be treatment related, but as this is relatively common finding and can occur spontaneously in the laboratory rat, the presence in single animals at the intermediate and low level dose levels were considered unlikely to be treatment related.

Neuropathological findings:

not examined

Histopathological findings: non-neoplastic:

effects observed, treatment-related

Histopathological findings: neoplastic:

no effects observed

Other effects:

no effects observed

Details on results:

OTHER FINDINGSThe formulation analysis results were all within the range of 98 to 103% of nominal concentration demonstrating that the formulations were accurately prepared.

Effect levels

1

Key result

Dose descriptor:

LOAEL

Effect level:

25 other: mg sodium chlorite/kg bw/day

Based on:

test mat.

Sex:

male/female

Basis for effect level:

clinical signs

histopathology: non-neoplastic

2

Dose descriptor:

LOAEL

Effect level:

19 other: mg chlorite/ kg bw/day

Based on:

act. ingr.

Sex:

male/female

Basis for effect level:

clinical signs

histopathology: non-neoplastic

3

Key result

Dose descriptor:

NOAEL

Effect level:

10 other: mg sodium chlorite/kg bw/day

Based on:

test mat.

Sex:

male/female

Basis for effect level:

other: See remarks:

4

Dose descriptor:

NOAEL

Effect level:

7.5 other: mg chlorite/kg bw/day

Based on:

act. ingr.

Sex:

male/female

Basis for effect level:

other: See remarks:

Target system / organ toxicity

Key result

Critical effects observed:

yes

Lowest effective dose / conc.:

25 mg/kg bw/day (nominal)

System:

other: Gastrointestinal and haematopoietic

Organ:

spleen

stomach

Treatment related:

yes

Relevant for humans:

not specified

Applicant's summary and conclusion

Conclusions:

Sodium chlorite treatment by the oral (gavage) route for 13 weeks elicited toxicity at 80 and 25 mg/kg/day. At the highest dose level this was characterised by changes in clinical condition, mortality, haematological changes, histopathological abnormalities in the spleen and evidence of irritation of the gastric mucosa. Certain individual animals appeared more sensitive to treatment than others. Minor clinical signs and occasional histopathological abnormalities in the stomach mucosa were observed at 25 mg/kg/day. At the lowest dose level there were no changes considered to be treatment-related and it was therefore, concluded that the no observed effect level was 10 mg/kg/day.

Executive summary:

A sub-chronic repeated dose toxicity study was conducted according to EPA guideline 82.1 which is equivalent to OECD guideline 408 under GLP conditions. The test article (sodium chlorite) was administered by gavage daily for 13 weeks to the rat. Doses were selected based on a previous range-finding study. The doses for the main study were 10, 25 and 80 mg/kg/day. Animals were observed daily and bodyweight and food consumption were recorded weekly. Ophthalmic examinations were performed on all animals before dosing started and on high dose and control animals during week 12. Blood samples for haematological and blood chemical investigations were obtained during week 13. Urine samples were analysed during week 13. At the end of the treatment period animals were sacrificed and necropsied. Several organs were weighed and a range of tissues was preserved for subsequent histopathology. Sodium chlorite treatment elicited toxicity at 80 and 25 mg/kg/day. At the highest dose level this was characterised by changes in clinical condition, mortality, haematological changes, histopathological abnormalities in the spleen and evidence of irritation of the gastric mucosa. Certain individual animals appeared more sensitive to treatment than others. Minor clinical signs and occasional histopathological abnormalities in the stomach mucosa were observed at 25 mg/kg/day. At the lowest dose level there were no changes considered to be treatment-related and it was therefore, concluded that the no observed effect level (NOEL) was 10 mg/kg/day.