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Toxicological information

Repeated dose toxicity: oral

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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
Reliability:
1 (reliable without restriction)
Rationale for reliability incl. deficiencies:
other: Comparable to guideline study
Cross-reference
Reason / purpose for cross-reference:
reference to same study

Data source

Reference
Reference Type:
study report
Title:
Unnamed
Year:
2008
Report date:
2008

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)
GLP compliance:
yes (incl. QA statement)
Limit test:
no

Test material

Constituent 1
Details on test material:
- Name of test material (as cited in study report): Phosphoric acid
- Molecular formula (if other than submission substance): not applicable
- Molecular weight (if other than submission substance): not applicable
- Smiles notation (if other than submission substance): not applicable
- InChl (if other than submission substance): not applicable
- Structural formula attached as image file (if other than submission substance): not applicable
- Substance type: no data
- Physical state: no data
- Analytical purity: no data
- Impurities (identity and concentrations): no data
- Composition of test material, percentage of components: no data
- Isomers composition: no data
- Purity test date: no data
- Lot/batch No.: no data
- Expiration date of the lot/batch: no data
- Radiochemical purity (if radiolabelling): not applicable
- Specific activity (if radiolabelling): not applicable
- Locations of the label (if radiolabelling): not applicable
- Expiration date of radiochemical substance (if radiolabelling): not applicable
- Stability under test conditions: no data
- Storage condition of test material: no data
- Other: no data

Test animals

Species:
rat
Strain:
Sprague-Dawley
Sex:
male/female
Details on test animals or test system and environmental conditions:
TEST ANIMALS
- Source: ORIENTBIO Inc., Korea
- Age at study initiation: male rats: 9 weeks old; female rats: 8 weeks old
- Weight at study initiation: male: 288.6 - 336.7 g; female: 163.5 - 188.9 g
- Fasting period before study:
- Housing: Stainless wire cage, 260 W x 50 D x 10 H (mm3). Polycarbonate cage, 260 W x 20 L x 80 H (mm3): number of animals per cage: 1 animal per cage and during mating period: 1 male and 1 female; during lactation period: 1 female and neonate
- Use of restrainers for preventing ingestion (if dermal): not applicable
- Diet (e.g. ad libitum): no data
- Water (e.g. ad libitum): no data
- Acclimation period: 1 week
- Cages and fodder tub were replaced once every two weeks, and water bottles were replaced twice a week. Housing materials were washed with cage washer and sterilized by an autoclave.
- During the acclimation period, animals were marked on the tail using a red indelible pen in accordance with the method of identification and marked on the tail using a blue indelible pen during the experimental period. Each cage was compartmentalized by an identification card to enable identification of individual animals.

ENVIRONMENTAL CONDITIONS
- Temperature (°C): 19.2 - 23.4 °C
- Humidity (%): 27.6 - 64.3 %
- Air changes (per hr): 10 - 15 times/hour
- Photoperiod (hrs dark / hrs light): 12 hours dark/12 hours light (lights on at 7 a.m., lights off at 7 p.m.
- Light intensity: 150 - 300 Lux

Administration / exposure

Route of administration:
oral: gavage
Vehicle:
water
Details on oral exposure:
no data
Analytical verification of doses or concentrations:
no
Duration of treatment / exposure:
The males were administrated once daily during 2 weeks prior to mating, the 2 weeks of mating period, and 2 weeks after mating (total 6 weeks). The females of the main group were administrated once daily from 2 weeks before mating to Day 4 post partum (approximately 54 days). The females of the recovery group (without mating) were administrated approximately 54 days (applied equally main group females). Also, coitus was confirmed, but females did not show gestational signs, and were administrated until gestation Day 26.
Frequency of treatment:
Daily
Doses / concentrations
Remarks:
Doses / Concentrations:
0, 125, 250 and 500 mg/kg
Basis:
no data
No. of animals per sex per dose:
See 'Any other information on materials and methods including tables'
Control animals:
yes, concurrent vehicle
Details on study design:
- Group designation: see free text field 'Any other information on materials and methods including tables
Positive control:
No data

Examinations

Observations and examinations performed and frequency:
CAGE SIDE OBSERVATIONS: No data
- Time schedule: No data

DETAILED CLINICAL OBSERVATIONS: Yes
- Time schedule: once a week

BODY WEIGHT: Yes
- Time schedule for examinations: no data

FOOD CONSUMPTION AND COMPOUND INTAKE: Yes
- No data

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 data

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

OPHTHALMOSCOPIC EXAMINATION: No data
- Time schedule for examinations: No data
- Dose groups that were examined: No data

HAEMATOLOGY: Yes
- Time schedule for collection of blood: No data
- Anaesthetic used for blood collection: No data
- Animals fasted: No data
- How many animals: main group and recovery group compared with control group
- Parameters checked in table: eosinophils, prothrombin time, no further data on other parameters examined

CLINICAL CHEMISTRY: Yes
- Time schedule for collection of blood: No data
- Animals fasted: No data
- How many animals: main group and recovery group compared with control group
- Parameters checked: phosphorus (P), triglycerides, total proteins (TP), albumin, no further data on other parameters examined

URINALYSIS: Yes
- Time schedule for collection of urine: No data
- Metabolism cages used for collection of urine: No data
- Animals fasted: No data
- Parameters checked: No data

NEUROBEHAVIOURAL EXAMINATION: Yes
- Time schedule for examinations: No data
- Dose groups that were examined: all of the control and treatment groups
- Battery of functions tested: sensory reflex and motor function test: pinna reflex, auditory reflex, corneal reflex, papillary reflex, traction, rota rod, open field test

OTHER: organ weights
Sacrifice and pathology:
SACRIFICE
Remaining animals were euthanized under CO2 gas anesthesia

GROSS NECROPSY
- Spleen, kidney, epididymis, gastro-intestinal organs, uterus (horn)

HISTOPATHOLOGY / ORGAN WEIGHTS
- Kidney (tubulus), epididymis, uterus (horn)
Statistics:
Statistical analysis of this study was performed by the SAS program (SAS R 9.1.3, SAS Institute Inc., U.S.A.). For the data including body weights, food consumption, urine, hematology, blood biochemistry parameters, organ weights, mating results, birth and survival rates, sensory reflex, and motor function test, the Bartlett test was conducted to test for variance homogeneity at the 5.0% one-tailed probability level. One-way analysis of variance (ANOVA) test was employed on homogeneity, if significant, followed by Dunnett's t-test for multiple comparisons. Kruskal-wallis was employed on heterogeneity, if significant, followed by Mann-whitney u-test for multiple comparisons. Group comparison was performed at the 5.0 and 1.0% two-tailed probability level. For the data of recovery, Folded-F test was conducted to test for variance homogeneity at the 5.0% two-tailed probability level. Student t-test was employed on homogeneity, if overrule, Aspin-Welch t-test was performed at the 5.0 and 1.0% two-tailed probability level. Non pregant animals were excluded.

Results and discussion

Results of examinations

Clinical signs:
no effects observed
Mortality:
no mortality observed
Body weight and weight changes:
no effects observed
Food efficiency:
not specified
Ophthalmological findings:
not examined
Haematological findings:
effects observed, treatment-related
Clinical biochemistry findings:
effects observed, treatment-related
Urinalysis findings:
no effects observed
Behaviour (functional findings):
no effects observed
Organ weight findings including organ / body weight ratios:
effects observed, treatment-related
Gross pathological findings:
effects observed, treatment-related
Histopathological findings: non-neoplastic:
effects observed, treatment-related
Histopathological findings: neoplastic:
not specified
Details on results:
CLINICAL SIGNS AND MORTALITY: In the main group, death was observed in one female (2403) of the 500 mg/kg treatment group on Day 16 of administration. And In the recovery group, death was observed in one female (2416) of the 500 mg/kg treatment group on Day 52 of administration. In the main group, no clinical signs were observed in males and females of 125 mg/kg treatment group and in females of the 250 mg/kg treatment group. But, transient salivations which appeared immediately after administrations were sporadically observed in four males of the 250 mg/kg treatment group from Day 22 of administration to completion of administration. Also, transient salivations were observed in all males of the 500 mg/kg treatment group from Day 20 of administration to completion of administration and in four females of the 500 mg/kg treatment group from Day 2 of gestation to completion of administration, sporadically or persistently. In addition, mucous stool, soft stool, and dirty nose were observed in one male (1407) of the 500 mg/kg treatment group on Day 22 and 23. In the recovery group, transient salivations were observed in almost all males of the 500 mg/kg treatment group from Day 16 of administration to completion of administration and in almost all females of the 500 mg/kg treatment group from Day 20 of administration to completion of administration, sporadically or persistently.

DETAILED CLINICAL SIGNS: No detailed clinical signs were observed in all animals of the control and treatment groups.

BODY WEIGHT AND WEIGHT GAIN
In the main group, no treatment-related abnormalities in body weight gain were noted in all animals of the control and treatment groups. In females of the recovery group, body weights on Day 55 of administration were significantly decreased (p<0.05) compared with the control group in the 500 mg/kg treatment group. Besides, no treatment-related abnormalities in body weight gain were noted in all males.

FOOD CONSUMPTION AND COMPOUND INTAKE
In the main group, no treatment-related abnormalities in food consumption were noted in all animals of the control and treatment groups. In feamles of the recovery group, food consumtpion was significantly decreased (p<0.05) compared to with the control group on Day 13 and 34 of administration in the 500 mg/kg treatment group. Besides, no treatment-related abnormalities in food consumption were noted in all males.

FOOD EFFICIENCY
No data

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

OPHTHALMOSCOPIC EXAMINATION
Not examined

HAEMATOLOGY
In the main group, no treatment-related abnormalities in hematology were noted in all animals of the control and treated groups. In the recovery group, eosinophils (EOS) of males and phrotrombin time (PT) of females were significantly increased (p<0.05) compared with the control group in the 500 mg/kg treatment group.

CLINICAL CHEMISTRY
In the main group, phosphorus (P) was significantly decreased (p< 0.05) compared with the control group in females of the 125 mg/kg treatment group. In females of the recovery group, triglycerides (TG, p<0.05), total proteins (TP, p<0.05), and albumin (Alb, p<0.01) were significantly decreased compared with the control group in the 500 mg/kg treatment group.

URINALYSIS
In the main and recovery groups, no treatment-related abnormalities were noted in the urinalysis parameters

NEUROBEHAVIOUR
Sensory reflex and motor function test: No treatment-related abnormalities in the pinna reflex, auditory reflex, corneal reflex, papillary reflex, traction, rota rod, and open field test were observed in all animals of the control and treatment groups

ORGAN WEIGHTS
In the main group, no treatment-related abnormalities in absolute and relative organ weights of the males were noted compared with the control group. However, in females, absolute organ weights of kidney were significantly decreated (p<0.05, p<0.01) compared with the control group in all treatment groups, and relative organ weights of uterus were significantly decreased (p<0.05) compared with the control group in the 500 mg/kg treatment group. In the recovery group, no treatment-related abnormalities in absolute and relative organ weights of the males were noted. However, in females, fasted body weights (p<0.01) and absolute organ weights (p<0.05) were significantly decreased compared with the control group in all treatment groups.

GROSS PATHOLOGY
In the necropsy findings of the main group agenesis of spleen was observed in one male (1302) of the 250 mg/kg treatment group, and yellow spot of epididymis was observed in one male (1408) of the 500 mg/kg treatment group. The other abdominal signs were not observed, and the necropsy finding of the recovery group was not noted. In the necropsy findings for dead animals, gaseous distension was observed in one female (2403) of the 500 mg/kg treatment group in the main group on Day 16 of administration. In the recovery group, gaseous distension, severe hydronephrosis of left kidney and dilation of left uterine horn were observed in one female (2416) of the 500 mg/kg treatment group on Day 52 of administration.

HISTOPATHOLOGY: NON-NEOPLASTIC
In the main group, cast of kidney was observed in one male (1104) of the control group, and sper granuloma of epididymis was observed in one male (1408) of the 500 mg/kg treatment group. The other abnormal findings were not observed. In the recovery group, basophilic tubules of kidney was observed in one female (2119) of the control group. The other abnormal findings were not observed. In the 500 mg/kg treatment group of the main group female, no histopathological change in one dead animal (2403) was observed. In the 500 mg/kg treatment group of the recovery female, renal tissue in one dead animal (2416) was not observed by severe hydronephrosis of gross findings, and left uterine horn was dilated; however, no histopathological change was observed.

HISTOPATHOLOGY: NEOPLASTIC (if applicable)
No data

HISTORICAL CONTROL DATA (if applicable)
No data

OTHER FINDINGS
No data

Effect levels

Dose descriptor:
NOAEL
Effect level:
250 other: mg/kg
Based on:
not specified
Sex:
male/female
Basis for effect level:
other: see 'Remark'

Target system / organ toxicity

Critical effects observed:
not specified

Applicant's summary and conclusion

Conclusions:
Effects of the test substance for all males and females were not observed in less than 250 mg/kg in the toxicological change. However, two dead females in the 500 mg/kg treatment group had occurred, and findings of gaseous distension of gastrointestinal tract were observed. Mucous stool, soft stool, and dirty nose were observed in one male of the 500 mg/kg treatment group. Therefore, a NOAEL for repeated dose toxicity was determined at 250 mg/kg in all males and females.