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Administrative data

Endpoint:
sub-chronic toxicity: oral
Remarks:
Administration via diet for up to 8 weeks
Type of information:
experimental study
Remarks:
GLP, range-finder study; no specific guideline followed
Adequacy of study:
supporting study
Study period:
April - June 2005
Reliability:
2 (reliable with restrictions)
Rationale for reliability incl. deficiencies:
study well documented, meets generally accepted scientific principles, acceptable for assessment

Data source

Reference
Reference Type:
study report
Title:
Unnamed
Year:
2006
Report date:
2006

Materials and methods

Test guideline
Qualifier:
no guideline followed
GLP compliance:
yes
Limit test:
no

Test material

Constituent 1
Chemical structure
Reference substance name:
-
EC Number:
446-620-9
EC Name:
-
Cas Number:
120983-72-4
Molecular formula:
Hill formula: C5H6Cl2O CAS formula: C5H6Cl20
IUPAC Name:
2-Chloro-1-(1-chlorocyclopropyl)ethanone
Test material form:
liquid
Details on test material:
purity: 92.3 %

Test animals

Species:
rat
Strain:
Wistar
Details on species / strain selection:
Wistar Crl:(WI)WU BR
Sex:
male/female

Administration / exposure

Route of administration:
oral: feed
Vehicle:
unchanged (no vehicle)
Remarks:
test item blended with diet Provimi Kliba SA 25 G4
Analytical verification of doses or concentrations:
no
Remarks:
Details on analytical verification of doses or concentrations:
In parallel to this study analytical tests on stability and homogenity of the test substance in the diet were initated in 15 kg 1 and 500 ppm mixtures. These analyses revealed that JAU 6476 Chlormethylketon was homogenously distributed in these mixtures.
The stability tests done on day 0, 4 and 8 revealed that the test substance content decreased quickly [in a 1 ppm mixture to 0 % on day 8 (see Table 5-3) probably also earlier] in all mixtures probably caused by the high volatility (as noted by a very strong odor) of the test substance at room temperature.
Analytical investigations on mixtures fed to the animals were not performed.
Duration of treatment / exposure:
8 weeks; no recovery period
Frequency of treatment:
daily
Doses / concentrationsopen allclose all
Dose / conc.:
0 ppm
Dose / conc.:
10 ppm
Dose / conc.:
100 ppm
Dose / conc.:
500 ppm
Dose / conc.:
1 000 ppm
No. of animals per sex per dose:
6
Control animals:
yes, plain diet

Examinations

Observations and examinations performed and frequency:
The animals were regularly observed and weighed. Food intake was determined.
Investigations on blood samples were performed. Organs and tissues were subjected to gross and histopathological investigations in liver, spleen, kidney and stomach (done in 0, 10, 100 and 500 ppm rats only). Selected organs (liver, spleen, kidneys) were weighed.

Inspection of Animals: for morbidity and mortality Twice daily, once daily on weekends and public holidays
detailed clinical examinations Weekly
Determination of:
body weight(s): Weekly
food consumption: Weekly
Clinical Laboratory Investigations: All rats per group
hematology: Day 56/57
clinical chemistry: Day 56/57

Sacrifice and pathology:
GROSS PATHOLOGY: Yes
- Organ weights: liver, spleen, kidney stomach

HISTOPATHOLOGY: Yes
- Organs: liver, spleen, kidney stomach (done in 10, 100 and 500 ppm rats only)
Other examinations:
If animals became ill, they were marked (with cage labels) or set apart, observed more frequently and sacrificed prematurely, if death seemed imminent.
Statistics:
Statistical evaluations on body weight and organ weight data were done using the
Dunnett-test in connection with a variance analysis. Evaluating biochemical parameters an analysis of variance followed by a Dunnett test, an adjusted Welch test or a Kruskal-Wallis test was performed. For all these tests SAS® routines were used.

Results and discussion

Results of examinations

Clinical signs:
effects observed, treatment-related
Description (incidence and severity):
- Piloerection in 1000 ppm males and in females from 500 ppm onwards.
- Emaciation in 1000 ppm females.
- Sunken flunks in 1000 ppm males and females.
- Squatting position in one 1000 ppm female.
There was also an increased urine excretion in 1000 ppm rats and in two of six 100 ppm males noted at clinical observation.
Mortality:
no mortality observed
Description (incidence):
no effects observed
Body weight and weight changes:
effects observed, treatment-related
Description (incidence and severity):
There were no changes in body weights and body
weight gain at 10 and 100 ppm in males and females. At 500 ppm (only males) and
at 1000 ppm statistically significantly depressed body weights were observed.
Food consumption and compound intake (if feeding study):
effects observed, treatment-related
Description (incidence and severity):
No toxicologically relevant changes in food intake were observed up to 100 ppm.
At 500 and 1000 ppm reduced food intake was visible, if considered per week or cumulative, even when the different time of exposure is considered.
Food efficiency:
not examined
Haematological findings:
effects observed, treatment-related
Description (incidence and severity):
No toxicologically relevant chances in the red blood parameters up to 1000 ppm. There was an elevated mean for leucocytes in 1000 ppm males (p>0.05) and an increase in thrombocytes from 500 ppm onwards in both sexes. Other statistical significances are of no toxicological relevance, because the deviations to controls were very small.
Clinical biochemistry findings:
effects observed, treatment-related
Description (incidence and severity):
No toxicologically relevant changes occurred in AS AT and ALAT activity in the plasma up to 1000 ppm in both sexes. The alkaline phosphatase activity was reduced from 100 ppm onwards in both sexes.
The plasma substrate concentrations of glucose, cholesterol, triglyceride and protein were not altered up to 500 ppm in males and up to 100 ppm in females. At higher concentration levels each a decrease in these parameters was observed (mostly statistically significant).
The means for urea and bilirubin were unaffected up to 1000 ppm. The significantly higher creatinine mean of 1000 ppm males is not considered as toxicologically relevant, as the deviation to the control mean is too small.
Immunological findings:
not examined
Organ weight findings including organ / body weight ratios:
effects observed, treatment-related
Description (incidence and severity):
There was a statistically significant decrease in liver weights from 500 ppm (absolute) or 10 ppm (relative) in males. A statistically significant decrease in absolute liver weights of 1000 ppm females is considered to be secondary to reduced body weights, because corresponding relative liver weights were increased (p>0.05). At 1000 ppm reduced (p<0.05) absolute spleen and kidney weights were found in males most likely due to reduced body weights, because corresponding relative weights were increased statistically significantly. This is true also for the reduced (p>0.05) absolute spleen weights of 1000 ppm females. 1000 ppm females exhibited elevated absolute (p>0.05) and relative (p<0.01) kidney weights.
Gross pathological findings:
effects observed, treatment-related
Description (incidence and severity):
There were no macroscopical organ changes attributable to the test substance were detected up to 100 ppm. At 500 and 1000 ppm discoloration on the kidneys (both sexes) and at 1000 ppm pelvic dilatations/enlargement on/of the kidney (females) were noted more frequently.
Neuropathological findings:
not examined
Histopathological findings: non-neoplastic:
effects observed, treatment-related
Description (incidence and severity):
At histopathology (done in 10, 100 and 500 ppm rats only) the kidneys of all males and females dosed at 500 showed swelling/degeneration of cortical tubules. Affected tubules revealed a paler cytoplasmic staining. The epithelial cells were swollen and their nuclei were hypertrophic. The severity score of these lesions was considered to be slight in most of the rats of either sex. At 100 ppm, one male and female each showed borderline changes of that type (grade 1, minimal). In the liver of females dosed at 500 ppm, cytoplasmic change was observed in two out of six. In four females of that group Kupffer cells were activated. Both liver changes had minimal severity scores (grade 1). All other findings present in the organs investigated in this study are known to occur spontaneously in rats of that age and strain.
Histopathological findings: neoplastic:
no effects observed
Description (incidence and severity):
no effects observed
Other effects:
effects observed, treatment-related
Description (incidence and severity):
As at study begin surprisingly the 1000 ppm diet mixture was found to produce a very strong acid odor the feeding of 1000 ppm diet was postponed for one week to have one week time to check at first possible high toxicity in rats receiving 500 ppm.

Effect levels

Dose descriptor:
LOAEL
Effect level:
67 ppm
Based on:
test mat.
Sex:
male/female
Basis for effect level:
body weight and weight gain
clinical biochemistry
food consumption and compound intake
haematology
histopathology: non-neoplastic
organ weights and organ / body weight ratios

Target system / organ toxicity

open allclose all
Critical effects observed:
yes
Lowest effective dose / conc.:
500 ppm
System:
urinary
Organ:
kidney
Treatment related:
yes
Relevant for humans:
not specified
Critical effects observed:
yes
Lowest effective dose / conc.:
500 ppm
System:
hepatobiliary
Organ:
liver
Treatment related:
yes
Relevant for humans:
not specified

Applicant's summary and conclusion

Conclusions:
no reliable NOAEL or NOEL could be determined due to instability of the substance in the diet
Executive summary:

JAU 6476-Chlormethylketon was administered in the diet to 6 male and 6 female Wistar rats per dose group in concentrations of 0, 10, 100 and 500 ppm for a period of 8 weeks. The same number of rats received 1000 ppm starting one week later for 7 weeks.


 


This study was intended as an exploratory feeding study for the dose selection for a 1  generation study.


 


Investigations on blood samples were performed. Organs and tissues were subjected to gross and histopathological investigations in liver, spleen, kidney and stomach (done in 0, 10, 100 and 500 ppm rats only). Selected organs (liver, spleen, kidneys) were weighed. The test substance was homogeneously distributable in the diet. Based on analytically established instability of test substance in the diet averaged dose levels of maximally 67, 335 and 670 ppm were roughly estimated for the 100, 500 and 1000 ppm concentrations. Consequently the diet as administration route has turned out to be inadequate.


 


In summary there were no remarkable effects at 10 ppm except the decrease of liver weights in males, which is not considered to reflect an adverse effect as corresponding histopathologically or biochemical findings are lacking. From 100 ppm onwards effects on the kidneys were evident and APh plasma activities were decreased. At 500 ppm and above body weights, blood parameters and liver were affected and clinical findings indicated poor general condition. At 1000 ppm more prounouced toxicity was evident.


As the test substance had turned out to be strongly instable in the diet caused by its high volatility the outcome of this study is limited (especially for the 10 ppm group) and cannot be used for setting a doubtless NOEL or NOAEL. However, following rough estimations of the decrease of the test substance content during a feeding period of 7 days the concentration of about 67 ppm (instead of 100 ppm) corresponding to 35.0 (males) and 36.3 mg/kg body weight per day (females) could be expected to be the LOAEL under the conditions described.