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Toxicological information

Repeated dose toxicity: oral

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Administrative data

Endpoint:
short-term repeated dose toxicity: oral
Type of information:
experimental study
Adequacy of study:
supporting study
Study period:
1996
Reliability:
1 (reliable without restriction)
Rationale for reliability incl. deficiencies:
other: Well documented report of a guideline study conducted according to GLP.

Data source

Reference
Reference Type:
study report
Title:
Unnamed
Year:
1996
Report Date:
1996

Materials and methods

Test guideline
Qualifier:
according to
Guideline:
OECD Guideline 407 (Repeated Dose 28-Day Oral Toxicity in Rodents)
Deviations:
no
GLP compliance:
yes (incl. certificate)
Limit test:
no

Test material

Reference
Name:
Unnamed
Type:
Constituent
Type:
Constituent
Type:
Constituent
Details on test material:
- Name of test material (as cited in study report): OS 114454
- Molecular formula (if other than submission substance): C7H16N2O4S
- Molecular weight (if other than submission substance): 225.29
- Smiles notation (if other than submission substance): [NH4+].O=S(=O)(O)CC(C)(C)/N=C(\[O-])C=C
- InChl (if other than submission substance): 1/C7H13NO4S.H3N/c1-4-6(9)8-7(2,3)5-13(10,11)12;/h4H,1,5H2,2-3H3,(H,8,9)(H,10,11,12);1H3
- Substance type: organic
- Physical state: liquid (aqueous solution)
- Analytical purity:
- Impurities (identity and concentrations):
- Composition of test material, percentage of components: 58.4% 2-Acrylamido-2-methylpropanesulfonic acid, ammonium salt in water
- Purity test date: no data
- Lot/batch No.: 1
- Expiration date of the lot/batch: 12 January 1998
- Stability under test conditions: stable
- Storage condition of test material: room temperature

Test animals

Species:
rat
Strain:
Sprague-Dawley
Sex:
male/female
Details on test animals and environmental conditions:
TEST ANIMALS
- Source: Charles River Laboratories, Kingston, NY, USA
- Age at study initiation: 44 days
- Weight at study initiation: Males: Mean = 219 g, Range = 196-241 g; Females: Mean = 165 g, Range = 143-191 g
- Fasting period before study: no
- Housing: 2 per cage during first week of acclimation, singly thereafter
- Diet: ad libitum
- Water: ad libitum
- Acclimation period: 17 days

ENVIRONMENTAL CONDITIONS
- Temperature (°C): 20-23°C
- Humidity (%): 20-68%
- Air changes (per hr): no data
- Photoperiod (hrs dark / hrs light): 12-hour light/dark cycle

IN-LIFE DATES: From: 16/021996 To:29/03/1996

Administration / exposure

Route of administration:
oral: gavage
Vehicle:
water
Details on oral exposure:
PREPARATION OF DOSING SOLUTIONS:

The test material (a 58.4% aqueous solution) was diluted with water to achieve the desired dose volumes. The appropriate amount of test material was weighed into a calibrated beaker. The vehicle (distilled water) was added to achieve the desired total volume. The dose solution was then stirred for at least 2 minutes or until mixed well. Fresh dosing solutions were prepared once weekly. Prepared dose solutions were stored at room temperature. The dose volume was 5 ml/kg.
Analytical verification of doses or concentrations:
yes
Details on analytical verification of doses or concentrations:
ANALYSIS OF DOSE SOLUTIONS
Analyses to determine homogeneity, stability, and concentration of the test and/or control materials with carriers under the conditions of this study were performed by the Testing Facility.
HOMOGENEITY
Prior to initiation of the study, batches of low-concentration and high-concentration dose solutions were prepared. Three samples each from the top, middle and bottom portion of each mixture were taken for analysis.
STABILITY
Duplicate samples pf the low- and high-concentration dose solutions were assayed 4, 7, and 14 days after preparation (samples for homogeneity assays, evaluated on the day of preparation, were used to establish concentration at time of preparation).
CONFIRMATION OF ANALYSIS
All 4 dose levels were assayed weekly (1 sample per concentration was taken and 2 sub-samples were analyzed).
Duration of treatment / exposure:
28 days
Frequency of treatment:
7 days/week
Doses / concentrations
Remarks:
Doses / Concentrations:
0, 50, 150, 400 and 1000 mg/kg
Basis:
actual ingested
No. of animals per sex per dose:
10 males and 10 females rats per group
Control animals:
yes, concurrent vehicle
Details on study design:
SELECTION
More animals than required for the study were purchased and acclimated. Animals considered unsuitable for the study on the basis of pretest physical examinations or outlying body weight data were eliminated prior to random selection for group assignment. Individual weights of animals placed on test for the main study were within ±20% of the mean weight for each sex.
GROUP ASSIGNMENT
Animals considered suitable for the study were distributed into 3 groups of 10 animals per sex (Groups I, IV and V) and 2 groups of 5 animals per sex (Groups II and III) by a computerized random sort program so that body weight for each group were comparable.
Positive control:
No

Examinations

Observations and examinations performed and frequency:
CAGE SIDE OBSERVATIONS: Yes
- Time schedule: twice daily
- Cage side observations: mortality, genral appearance and signs of severe toxic or pharmacologic effects

DETAILED CLINICAL OBSERVATIONS: Yes
- Time schedule: twice pretest and weekly thereafter

BODY WEIGHT: Yes
- Time schedule for examinations: twice pretest, weekly during treatment and recovery perios, and terminally (after fasting)

FOOD CONSUMPTION: Yes
- Time schedule: weekly, beginning one week prior to test

OPHTHALMOSCOPIC EXAMINATION: No

HAEMATOLOGY: Yes
- Time schedule for collection of blood: Termination: test dat 29; Recovery: test day 43
- Anaesthetic used for blood collection: Yes, CO2/O2
- Animals fasted: Yes, overnight
- How many animals: Termination: 10 animals/sex/groups I, IV and V, 5 animals/sex/group II and III; Recovery: 5 animals/sex/group I, IV and V
- Parameters checked: haemoglobin concentration, hematocrit, erythrocyte count, platelet count, mean corpuscular volume, mean corpuscular haemoglobin, mean corpuscular haemoglobin, total leukocyte count, differential leukocyte count, absolute lymphocytes, absolute segmented neuttophils, prothrombin time, activated partial thromboplastin time, erythrocyte morphology

CLINICAL CHEMISTRY: Yes
- Time schedule for collection of blood: Termination: test day 29; Recovery: test day 43
- Animals fasted: Yes, overnight
- How many animals: : Termination: 10 animals/sex/groups I, IV and V, 5 animals/sex/group II and III; Recovery: 5 animals/sex/group I, IV and V
- Parameters checked: Aspartate aminotransferase, alanine aminotransferase, alkaline phosphatase, blood urea nitrogen, creatinine, glucose, cholesterol-enzymatic, triglycerides, total protein, albumin, flobulin, albumin/globulin ratio, total bilirubin, sodium, potassium, chloride, calcium, inorganic phosphorus, gamma-glutamyl transferase

URINALYSIS: Yes
- Time schedule for collection of urine: Termination: test day 28: Recovery: test day 42
- Metabolism cages used for collection of urine: Yes
- Animals fasted: Yes, 4 hours prior
- How many animals: Termination: 10 animals/sex/groups I, IV and V, 5 animals/sex/group II and III; Recovery: 5 animals/sex/group I, IV and V
- Parameters checked: specific gravity, protein, glucose, ketones, occult blood, pH, bilirubon, urobilinogen, sediment

NEUROBEHAVIOURAL EXAMINATION: No
Sacrifice and pathology:
GROSS PATHOLOGY: Yes (see table of results below)

HISTOPATHOLOGY: Yes
Statistics:
Statistical evaluation was made by the appropriate one way analysis of variance technique followed by a post-hoc comparison procedure (e.g., Dunnett’s test or Kruskal-Wallis test).

Results and discussion

Results of examinations

Clinical signs:
no effects observed
Mortality:
no mortality observed
Body weight and weight changes:
no effects observed
Food consumption and compound intake (if feeding study):
no effects observed
Food efficiency:
no effects observed
Water consumption and compound intake (if drinking water study):
not examined
Ophthalmological findings:
not examined
Haematological findings:
no effects observed
Clinical biochemistry findings:
no effects observed
Urinalysis findings:
no effects observed
Behaviour (functional findings):
not examined
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:
not examined
Details on results:
CLINICAL SIGNS AND MORTALITY: 1 male receiving the highest dose (Animal No. 5072; 1000 mg/kg/day) exhibited a number of adverse signs (lethargy, emaciation, watery stool, yellow ano-genital staining, decreased fecal volume) during the first week of the study but recovered by the end of Week 2. No unusual post-mortem findings were seen in this particular animal at study termination. Although the occurrence of adverse signs were in a high-dose animal is suggestive of an effect of the test material, the rapid recovery and absence of subsequent signs make such a relationship questionable. These reversible signs in a single animal are not considered to be toxicologically questionable. No other remarkable signs suggestive of an effect of test material administration were seen subsequently. (A mass seen in low-dose female No. 2572 during the last week of treatment was confirmed at necropsy to be an abscess and was considered unrelated to treatment.
BODY WEIGHT AND WEIGHT GAIN: Mean body weights and body weights gains for the 1000 mg/kg/day male group were slightly lower than control values at Week 1, primarily as a result of a large weight loss in the animal which exhibited the signs noted above. The differences were not statistically significant. Values for the control and treated groups of males were comparable from Weeks 2 through 4. During the recovery period, weights for treated males were slightly higher than control weights, reflecting the selection of slightly heavier animals in this group for recovery. Body weight values for control and treated females were comparable throughout the treatment and recovery periods.
FOOD CONSUMPTION AND COMPOUND INTAKE: Mean food consumption in the male 1000 m/kg/day group was slightly lower during Week 1, reflecting a large decrease in food consumption in the animal (No. 5072) discussed above. Values in subsequent weeks were similar to, or higher than, control values. No effect on food consumption was seen in with any other dose either in the male or female groups. Food consumption values for these groups were comparable to concurrent control values or exhibited normal variability.
GROSS PATHOLOGY: None of the macroscopic findings in rats sacrificed at the end of the treatment and post-treatment recovery periods were considered to be treatment related.
HAEMATOLOGY: No effects evident after 4 weeks treatment or after 2-week treatment-free recovery period. Values for control and treated groups were comparable to concurrent control values or exhibited normal variability. A single statistically significant difference (slightly elevated platelet count in low dose females at study termination) was not considered to be an effect of test material administration.
CLINICAL CHEMISTRY: No effects of test material administration on clinical chemistry values were evident. Values for control and treated groups were generally comparable after 4 weeks of treatment and at termination of the recovery period. The single statistically significant difference (slight increase in the mean blood urea nitrogen value of females in the 150 mg/kg/day group at study termination) was slight and not dose-related and was not considered to be an effect of test material administration.
URINALYSIS: Urinalyses were unremarkable; there were no differences between values for control and treated groups which were considered indicative of an effect of the test material administration.
ORGAN WEIGHTS: Organ weight values for animals euthanized after 4 weeks of treatment and at termination of the recovery period were unremarkable; no effects of the test material administration were evident.
MACROSCOPIC PATHOLOGY: None of the macroscopic findings in rats sacrificed at the end of the treatment and post recovery periods were considered to be treatment related. They occurred sporadically in the treated and control groups and were considered to be incidental. Similar findings had been observed in untreated control rats of a similar strain and age used in other studies conducted in the same facility.
MICROSCOPIC PATHOLOGY: None of the microscopic findings in the 29 various organs and tissues examined at the end of the treatment and post-recovery periods were considered to be treatment related. They occurred sporadically in the treated and control groups and were considered to be incidental. Similar findings had been observed in untreated control rats of a similar strain and age used in other studies conducted in the same facility.

Effect levels

Dose descriptor:
NOEL
Effect level:
1 000 mg/kg bw/day (actual dose received)
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

Control 50 mg/kg/d 150 mg/kg/d  400 mg/kg/d 1000 mg/kg/d 
Body weight   A  A  A  A  B
Food consumption   A  A  A  A  B
General symptoms  A  A  A  A  B
Blood chemistry  A  A  A  A  A
Haematology  A  A  A  A  A
Urinalysis  A  A  A  A  A
Organ weights  A  A  A  A  A
Autopsy findings  A  A  A  A  A
Histology  A  A  A  A

A = within normal limits

B = transient weight loss, decreased food consumption and abnormal signs in a single (male) animal seen during the first week only, with recovery by the second week, and not considered toxicollogically significant.

Applicant's summary and conclusion

Conclusions:
Oral gavage administration of the test material to rats for 4 weeks at doses of 50, 150,400 and 1000 produced no significant toxicity at any dose level. Under the conditions of this study, the No Observed Effect Level (NOEL) was determined to be 1000 mg/kg/day.