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Toxicological information

Repeated dose toxicity: oral

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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:
other: GLP Guideline study
Cross-reference
Reason / purpose:
reference to other study

Data source

Reference
Reference Type:
study report
Title:
Unnamed
Year:
1998
Report Date:
1998

Materials and methods

Test guidelineopen allclose all
Qualifier:
according to
Guideline:
OECD Guideline 408 (Repeated Dose 90-Day Oral Toxicity in Rodents)
Deviations:
no
Qualifier:
according to
Guideline:
EU Method B.26 (Sub-Chronic Oral Toxicity Test: Repeated Dose 90-Day Oral Toxicity Study in Rodents)
Deviations:
no
GLP compliance:
yes
Limit test:
no

Test material

Reference
Name:
Unnamed
Type:
Constituent

Test animals

Species:
rat
Strain:
Wistar
Sex:
male/female
Details on test animals and environmental conditions:
Wistar rats (Chbb: THOM (SPF)) were obtained from Dr. Karl Thomae GmbH, Biberach/Riss, D

Administration / exposure

Route of administration:
oral: drinking water
Vehicle:
unchanged (no vehicle)
Analytical verification of doses or concentrations:
yes
Details on analytical verification of doses or concentrations:
Test solutions were analysed at the start and end of the study to assure that the concentrations were correct and the 4-day stability was assessed as 97%. The mixtures were prepared at no less than 4-day intervals.
Duration of treatment / exposure:
3 months
Frequency of treatment:
continuously
Doses / concentrations
Remarks:
Doses / Concentrations:
0, 600, 2400, 7200 and 15000 ppm (ca. 0, 37, 207, 586 and 1125 mg/kg/d)
Basis:
no data
No. of animals per sex per dose:
10
Control animals:
yes, concurrent vehicle
Details on study design:
Post-exposure period: none

Examinations

Observations and examinations performed and frequency:
Water consumption was determined once/week over a period of 4-days. Animals were weighed weekly and given a thorough physical examination at each weighing. Food consumption was determined weekly. Urine samples were taken on day 85, blood was sampled and analyzed on study day 88, the final bodyweight was recovered on day 91 and necropsies were conducted over days 92 to day 95. Food consumption, water consumption and body weight were determined each week. The animals’ state of health was checked each day. When the animals were weighed they were subjected to an additional comprehensive clinical examination. Clinincal chemistry parameters were: alanine aminotransferase, aspartate aminotransferase, alkaline phosphatase, serum-gammaglutamyltransferase. Blood chemistry parameters were: sodium, potassium, chloride, inorganic phosphate, calcium, urea, creatinine, glucose, total bilirubin, total protein, albumin, globulins, triglycerides, cholesterol, magnesium. In addition, complete hematology and urinalysis were conducted.
Sacrifice and pathology:
At necropsy, major organs were weighed and sections were fixed for histopathology. All animals were subjected to gross-pathological assessment, followed by histopathological examination using a complete tissue list.
Statistics:
Means and standard deviations for the variables food consumption, body weight, body weight change, water consumption and test substance intake (except control group) were calculated for the animals of each test group. They were printed out in the summary and individual value tables, with the exception that for test substance intake and body weight change only summary tables were prepared. For the parameters food consumption, water consumption, body weight and body weight change a parametric one-way analysis of variance was done via the F-test (ANOVA). If the resulting p-values were equal to or less than 0.05, a comparison of each dose group with the control group was carried out. These comparisons were performed simultaneously via Dunnett’s test for the hypothesis of equal means. If the results of this test were significant, labels (* for p < 0.05, ** for p < 0.01) were printed together with the group means in the tables. Both tests were performed two-sided. Statistical analysis of histopathology was conduced with a proprietary computer program.

Results and discussion

Results of examinations

Clinical signs:
no effects observed
Mortality:
no mortality observed
Body weight and weight changes:
effects observed, treatment-related
Food efficiency:
not specified
Water consumption and compound intake (if drinking water study):
effects observed, treatment-related
Haematological findings:
effects observed, treatment-related
Clinical biochemistry findings:
effects observed, treatment-related
Urinalysis findings:
effects observed, treatment-related
Behaviour (functional findings):
not specified
Organ weight findings including organ / body weight ratios:
effects observed, treatment-related
Gross pathological findings:
effects observed, treatment-related
Histopathological findings: non-neoplastic:
no effects observed
Histopathological findings: neoplastic:
no effects observed

Effect levels

Dose descriptor:
NOAEL
Effect level:
207 mg/kg bw/day (nominal)
Sex:
male/female
Basis for effect level:
other: Effects on kidneys

Target system / organ toxicity

Critical effects observed:
not specified

Any other information on results incl. tables

Mean test material consumption in mg/kg*day were:
- males: 33, 184, 529 and 1062 mg/kg
- females 42, 230, 643 and 1189 mglkg

MORTALITY, CLINICAL SIGNS and OPHTHALMOSCOPY
No animal died during the study and no adverse clinical signs or ophthalmoscopic findings were noted.

FOOD and WATER CONSUMPTION
Food consumption was impaired in the high dose treatment in males (-4 to -10%) and females (-5 to -12%) with statistical significance on several days. In addition, water consumption was significantly impaired in the high dose treatment in males (-5 to -19%) and females (-14 to -27%) with statistical significance on several days.

BODY WEIGHT and BODY WEIGHT GAIN
Body weight was impaired statistically significant in males and females of the 15000 ppm treatment and on few days also in females of the 7200 ppm treatment. On day 91, the values were 9% (15000 ppm males and females), and 6% (7200 ppm females) below controls, respectively.
Body weight change was impaired statistically significant in males and females of the 7200 and 15000 ppm treatment. On day 91, the values were 14% (15000 ppm males),
20% (7200 ppm females), 7% (7200 ppm males) and 16%
(7200 ppm males) below controls, respectively.

INTAKE OF TS
The approximate, mean daily intake of test substance in mg/kg body weight for the complete administration period is 33 -42 for the 600 ppm treatment, 184-230 for the 2400 ppm treatment, 529-643 for the 7200 ppm treatment and 1062-1189 for the 15000 ppm treatment.

CLINICAL CHEMISTRY, HEMATOLOGY and URINALYSES
Hematology: No substance-related changes were observed in the results of the hematology examinations of both sexes.
Clotting analyses: At the end of the study statistically significant prolonged prothrombin times were measured in the plasma of the high dose males. In the high dose females there was only a trend towards prolonged prothrombin times.
Enzymes: No substance-related changes were observed in the
results of the serum enzyme examinations of both sexes.
Blood chemistry: After 3 months of test substance administration statistically significant decreased total protein and globulin concentrations were found in the serum of the male and female animals of the high dose treatment.
Additionally, statistically significant decreased total protein levels were detected in the females of the 7200 ppm treatment. In the high dose males, triglyceride concentrations were statistically significant increased, whereas in the high dose females this finding was seen only as a trend. Moreover, statistically significant reduced creatinine levels were found in females of the two highest dosed treatments; in the males of the two highest dosed treatments only a trend towards decreased creatinine concentrations was observed.
Urinalyses: Increased specific gravity and reduced volume were found in the urine specimens of the male animals of the two highest dosed treatments. Furthermore, in most of the urine specimens of the males of the two highest dosed treatments a dark-yellow discoloration was noted. In the females no corresponding findings were observed.

OTHER DEVIATIONS
There are some further statistically significant inter-group differences in the results of the clinical pathology testing. These deviations are marginal, sporadic or inconsistent, when compared with the other sex, or lack dosage-relationship. Accordingly, these changes are considered to be of no toxicological significance.

PATHOLOGY
Relative kidney weights were increased in males and females of the 15000 ppm treatment and in males of the 7200 ppm treatment. In females of all treatments with the TS an altered cellular composition of the thymic cortex was seen.

CONCLUSION
The kidney appears to be a target organ at dose levels of 7200 ppm (about 586 mg/kg) in the drinking water and above. The NOAEL is 2400 ppm in drinking water or about 207 mg/kg*bw*day.
In all treated females an altered cellular composition of the thymic cortex was seen. This was assessed as being incidental, due to the following reasons:
1. there was no dose-response relationship
2. only females were affected
3. in randomly selected control groups of other subchronic studies the same finding was observed in several cases, also.

Applicant's summary and conclusion

Conclusions:
The test material is of low systemic toxicity if administered in repeated manner.
Executive summary:

2 -pyrrolidone was tested for its subchronic toxicity in a -month drinking water study. The study was conducted in compliance with the OECD Guideline 408 and the GLP.

The purpose of this study was an assessment of the systemic toxicity after repeated exposures to the test material. Wistar rats received 0, 37, 207, 586 and 1125 mg/kg/d 2-Pyrrolidone (corresponding to 600, 2400, 7200 and 15000 ppm) in the drinking water, respectively during 90 days.

All animals were subjected daily to clinical examinations. Food consumption, water consumption and body weight were determined each week. Urinalysis, clinicochemical and hematological examinations were carried out towards the end of the administration period. All animals were subjected to gross-pathological assessment, followed by histopathological examination.

No substance related effects were seen in the two lowest dose groups. In the mid-dose group all parameters checked were slightly adversed. In the high dose group, effects were dose-dependently more severe. The kidneys were the most affected organs.

Due to adverse effects on the renal system, the NOAEL is considered to be 207 mg/kg b.w. for male and female rats.