Registration Dossier
Registration Dossier
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EC number: 605-140-1 | CAS number: 158237-07-1
Repeated dose toxicity: oral
Administrative data
- Endpoint:
- chronic toxicity: oral
- Type of information:
- experimental study
- Adequacy of study:
- key study
- Study period:
- 6 Dec 1994 - 17 Dec 1996
- Reliability:
- 1 (reliable without restriction)
- Rationale for reliability incl. deficiencies:
- guideline study
Cross-reference
- Reason / purpose for cross-reference:
- reference to same study
Reference
- Endpoint:
- carcinogenicity: oral
- Type of information:
- experimental study
- Adequacy of study:
- key study
- Study period:
- 6 Dec 1994 - 17 Dec 1996
- Reliability:
- 1 (reliable without restriction)
- Rationale for reliability incl. deficiencies:
- guideline study
- Reason / purpose for cross-reference:
- reference to same study
- Qualifier:
- according to guideline
- Guideline:
- OECD Guideline 453 (Combined Chronic Toxicity / Carcinogenicity Studies)
- Version / remarks:
- adopted 12th May 1981
- Deviations:
- no
- GLP compliance:
- yes
- Species:
- rat
- Strain:
- Wistar
- Remarks:
- Hsd/WIN: WU
- Sex:
- male/female
- Details on test animals or test system and environmental conditions:
- TEST ANIMALS
- Source: Harlan Winkelmann
- Females nulliparous and non-pregnant: yes
- Age at study initiation: 5 - 6 weeks
- Weight at study initiation: 110 - 182 g (males), 102 - 158 g (females)
- Housing: 5 animals separated by sex in Type III Makrolon® and Ha cages, The cages containing the experimental animals were separated by groups and placed on shelves in order of ascending animal number. The position of the shelves was changed on a random basis every 4 weeks.
- Diet: Altromin® 1321 meal (Altromin GmbH, Lage) containing 1% peanut oil, ad libitum except during the urine collection period
- Water: tap water, ad libitum
- Acclimation period: 1 week
DETAILS OF FOOD AND WATER QUALITY:
The tap water complied with drinking water standards in accordance with the Deutsche Trinkwasserverordnung.
ENVIRONMENTAL CONDITIONS
- Temperature: 22 ± 2°C
- Humidity: 55% ± 5%
- Air changes (per hr): 15 - 20
- Photoperiod (hrs dark / hrs light): 12/12 - Route of administration:
- oral: feed
- Vehicle:
- unchanged (no vehicle)
- Details on exposure:
- - DIET PREPARATION
- Rate of preparation of diet (frequency): weekly
- Mixing appropriate amounts with: the test substance was blended (using a mixing granulator manufactured by Loedige, Paderborn) with Altromin® 1321 containing 1% peanut oil to minimize dust formation (including 0 ppm concentration). - Analytical verification of doses or concentrations:
- yes
- Details on analytical verification of doses or concentrations:
- Analytical investigations to demonstrate homogeneity and stability of the test compound in diet preparations were done prior to commencement of the study. The test substance content of diet mixtures fed to the animals was checked analytically at regular intervals throughout the study (start of study, randomly each 3 month period, end of study). Per dose one sample of the food mixes was taken on the day the mixture was prepared, and another was taken after being kept underanimal room conditions for the feeding period (7 days). All these samples were kept deep frozen (at temperatures of approx. -20°C) until analysis.
- Duration of treatment / exposure:
- 24 month
- Frequency of treatment:
- daily per feed, ad libitum
- Dose / conc.:
- 50 ppm
- Remarks:
- corresponding to
2.5 mg/kg bw/day (males)
3.4 mg/kg bw/day (females) - Dose / conc.:
- 200 ppm
- Remarks:
- corresponding to
10.3 mg/kg bw/day (males)
14.6 mg/kg bw/day (females) - Dose / conc.:
- 1 000 ppm
- Remarks:
- corresponding to
52.7 mg/kg bw/day (males)
75.4 mg/kg bw/day (females) - Dose / conc.:
- 3 000 ppm
- Remarks:
- corresponding to
170.4 mg/kg bw/day (males) - Dose / conc.:
- 4 000 ppm
- Remarks:
- corresponding to
326.7 mg/kg bw/day (females) - No. of animals per sex per dose:
- 60
- Control animals:
- yes, plain diet
- Details on study design:
- - Dose selection rationale: The dosages were selected on the basis of the results of a subchronic feeding study in Wistar rats in which the test substance had been administered to 10 males and 10 females at doses of 0, 20, 100, 400, 1600 and 6400 ppm for 13 weeks. In this study concentrations of up to 400 ppm were tolerated without adverse effects. At higher concentrations body weight depression, effects on blood parameters (essentially reduced hemoglobin and hematocrit values and elevated reticulocyte means), inhibition of the cholinesterase activity (erythrocytes) and signs of changed liver function such as altered biochemical parameters (enhanced cholesterol plasma levels, reduced contents of triglycerides and unesterified fatty acids, lower albumin means), enzyme induction, increased liver weights and occurrence of hepatocellular hypertrophy were evident. Rats receiving 6400 ppm exhibited diffuse hyperplasia of the bladder urothelium as well. In further electron-microscopical investigations on the urinary bladder cytotoxicity with mild regenerative hyperplasia was detected in 6400 ppm males (females were not investigated) with higher frequency than in controls.
- Rationale for animal assignment: The rats were weighed individually beforehand and the required number of animals were then grouped by weight (light, midde and heavy subgroups) and randomly distributed into large containers. - Positive control:
- Not applicable
- Observations and examinations performed and frequency:
- CAGE SIDE OBSERVATIONS: Yes
- Time schedule:
twice daily and daily at weekends and on bank holidays
DETAILED CLINICAL OBSERVATIONS: Yes
- Time schedule: weekly
Parameters: orifices, posture, general behavior, respiration and excretory products were carefully examined, with any significant findings were registered. Animals that became ill or had developed neoplasms that could lead to death were kept apart, observed more frequently, and killed prematurely if death was imminent.
BODY WEIGHT: Yes
- Time schedule for examinations:
weekly from start to Week 13, and every 2 weeks thereafter up to Week 105
FOOD CONSUMPTION AND COMPOUND INTAKE:
- Food consumption for each animal determined and mean daily diet consumption calculated as g food/kg body weight/day: Yes
- Compound intake calculated as time-weighted averages from the consumption and body weight gain data: Yes
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
WATER CONSUMPTION AND COMPOUND INTAKE: Yes
- Time schedule for examinations: weekly within the first 13 weeks of treatment and every 4 weeks thereafter up to Week 101
OPHTHALMOSCOPIC EXAMINATION: Yes
- Time schedule for examinations and dose groups that were examined:
during the first two weeks of the study in all rats. In Week 54 and prior to the final bleeding all living animals scheduled for the final necropsy from the control group and the 3000 or 4000 ppm dose groups were examined. At study termination 200 and 1000 ppm males were investigated, additionally.
HAEMATOLOGY: Yes
- Time schedule for collection of blood:
Weeks 26/27, 52, 78 and 105/106
- Anaesthetic used for blood collection: Yes, for glucose and deproteinized whole blood only
- Animals fasted: Yes
- How many animals:
10
- Parameters analysed: differential blood count, erythrocyt morphology, erythrocyte count, Heinz bodies, hemoglobin, hematocrit, leucocyte count, mean corpuscular hemoglobin, mean corpuscular hemoglobin concentration, mean corpuscular volume, platelet count, thromboplastin time, reticulocytes
CLINICAL CHEMISTRY: Yes
- Time schedule for collection of blood:
Weeks 26/27, 52, 78 and 105/106
- Animals fasted: Yes
- How many animals:
10
- Parameters analysed: alkaline phosphatase, alanine aminotransferase, aspartate aminotransferase, gamma-glutamyl transferase, glutamate dehydrogenase, glucose, bilirubin, albumin, cholesterol, creatinine, total protein, triglyceride, urea, calcium, sodium, potassium, chloride and inorganic phosphate
URINALYSIS: Yes
- Time schedule for collection of urine:
Weeks 26/27, 52, 79 and 98/99
- Metabolism cages used for collection of urine: Yes
- Animals fasted: Yes
- Parameters analysed: sediment, bilirubin, blood, glucose, ketone bodies, pH, protein, urobilinogen (semiquantitative), density, volume, total protein (quantitative)
NEUROBEHAVIOURAL EXAMINATION: No
IMMUNOLOGY: No
OTHER:
Cholinesterase activity in brain
- Time schedule: Weeks 52/53 and 104/105
Cholinesterase activities in plasma and erythrocytes
- Time schedule: Week 26/27, measurement were repeated for males in the following week due to some implausible values
Determination of crystals in urine sediment
- Time schedule: Week 103
- Dose groups that were examined: 8 animals, in controls and rats of the 3000 and 4000 ppm groups - Sacrifice and pathology:
- GROSS PATHOLOGY: Yes
- Organ weights: brain, heart, liver, spleen, kidneys, adrenals, testes, ovaries
HISTOPATHOLOGY: Yes, the following organs and tissues were fixed for analysis: Adrenals , Aorta , Brain (3 locations) [in some cases half brain] , Caecum, Colon, Duodenum, Epididymides , Esophagus, Eyes (with eyelids), Exorbital lacrimal glands, Femur (incl. bone marrow and knee joint), Harderian glands, Head-Nose-Pharynx area, Heart, lleum, Jejunum, Kidneys, Larynx, Liver, Lungs, Lymph nodes (mandibular and mesenteric), Mammary glands, Optic nerve, Ovaries (incl. oviduct), Pancreas, Pituitary, Prostate, Rectum, Residual Intestine, Salivary glands, Sciatic nerve, Seminal vesicles, Skeletal muscle, Skin, Spinal cord (3 regions), Spleen, Sternum (with bone marrow), Stomach (with forestomach), Tattooed ears, Testes, Thymus (if present), Thyroid (parathyroid), Tongue, Trachea, Ureters, Urethra, Urinary bladder, Uterus, Vagina, Zymbal glands and all tissues showing abnormalities - Statistics:
- Dunnett-Test in connection with a variance analysis (body and organ weight data)
Analysis of variance followed by Dunnett test (Erythrocytes, Hemoglobin, Hematocrit, Mean Corpuscular Hemoglobin, Mean Corpuscular Hemoglobin Concentration, Mean Corpuscular Volume Erythrocytes, Thrombocytes, Albumin, Creatinine, Chloride, Glucose, Protein, Triglycerides, Urea)
Kruskal-Wallis-Test with a Steel-Test (food and water intake data)
Kruskal-Wallis test followed by adjusted U test (Heinz Bodies, Hepato Quick, Leucocytes, Reticulocytes, Band NeutrophiIs, EosinophiIs, Lymphocytes, Monocytes, Segmented Neutrophils, Bilirubin total, Gamma-Gtutamyltransferase, Specific Gravity, Glucose, dipstick, pH, dipstick, Protein Quantitative, Volume, pH)
Adjusted Welsh test (Atanine aminotransferase, Alkaline phosphatase, Aspartate aminotransferase, Choiinesterase, Choiinesterase in Erythrocytes, Choiinesterase in Brain, Cholesterol, Calcium, Glutamate dehydrogenase, Potassium, Sodium, Inorganic Phosphate) - Clinical signs:
- no effects observed
- Description (incidence and severity):
- no effects observed
- Mortality:
- mortality observed, non-treatment-related
- Description (incidence):
- All dose groups: a higher mortality in females was evident from Week 52 onwords; dose correlation is lacking
For details please refer to Table 1 in "any other information on results incl. tables". - Body weight and weight changes:
- effects observed, treatment-related
- Description (incidence and severity):
- 3000/4000 ppm: body weight depression in males from Week 7 / in females from Week 13 onwords
For details please refer to Table 2 in "any other information on results incl. tables". - Food consumption and compound intake (if feeding study):
- effects observed, treatment-related
- Description (incidence and severity):
- 3000/4000 ppm: slightly elevated food intake per kg body weight in both sexes (12.3%/19.2%)
The test substance intake in the treatment groups roughly corresponds to the theoretical dose intervals.
For details please refer to Table 3 in "any other information on results incl. tables". - Food efficiency:
- effects observed, treatment-related
- Description (incidence and severity):
- 3000/4000 ppm: indication for slightly reduced food efficiency in both sexes
For details please refer to Table 3 in "any other information on results incl. tables". - Water consumption and compound intake (if drinking water study):
- no effects observed
- Description (incidence and severity):
- no effects observed
For details please refer to Table 4 in "any other information on results incl. tables". - Ophthalmological findings:
- effects observed, treatment-related
- Description (incidence and severity):
- 1000 and 3000 ppm: higher incidence of moderate or severe opacities in the (whole) lens cortex of males at termination
For details please refer to Table 5 in "any other information on results incl. tables". - Haematological findings:
- effects observed, treatment-related
- Description (incidence and severity):
- 3000/4000 ppm: decreased Hb and Ht in males in Week 26/27 only, decreased MCH and MCHC in females
For details please refer to Tables 6 and 7 in "any other information on results incl. tables". - Clinical biochemistry findings:
- effects observed, non-treatment-related
- Description (incidence and severity):
- 3000/4000 ppm: slightly increased gamma glutamyltransferase (GOT) activity in both sexes), but not at all dates and without relevant time-dependent increase
Slightly decreased ASAT and APh activities in animals dosed at 1000 ppm and higher are not considered to indicate an adverse effect, since the values were within the physiological range.
For details please refer to Table 8 in "any other information on results incl. tables". - Urinalysis findings:
- effects observed, non-treatment-related
- Description (incidence and severity):
- Quantitative urinalyses did not reveal a treatment-related effect on pH, sediment, proteinurea, urine volume or urine density. Significant deviations from control values in these parameters were not considered relevant, since dose correlations were lacking.
For details please refer to Tables 10 and 11 in "any other information on results incl. tables". - 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):
- Interim necropsy:
3000/4000 ppm: significantly increased liver weights (absolute and relative ) in both sexes
Terminal necropsy:
3000/4000 ppm: partly significantly increased liver weights (absolute and relative) in both sexes, slightly decreased spleen weights (absolute and relative) in both sexes, increased relative testis weights in males
For details please refer to Tables 12 and 13 in "any other information on results incl. tables". - Gross pathological findings:
- effects observed, non-treatment-related
- Description (incidence and severity):
- Interim necropsy:
no effects observed
Terminal necropsy:
3000/4000 ppm: 6 males and 5 females appeared skinny - Neuropathological findings:
- not examined
- Histopathological findings: non-neoplastic:
- effects observed, treatment-related
- Description (incidence and severity):
- Interim examinations:
200 ppm: diffuse hyperplasia in the transitional epithelium of the urinary bladder in males
1000 ppm: diffuse hyperplasia in the transitional epithelium of the urinary bladder in males, hepatocellular hypertrophy (centrilobular) in both sexes
3000/4000 ppm: diffuse hyperplasia in the transitional epithelium of the urinary bladder, hepatocellular hypertrophy (centrilobular) and slight increase in the incidence and severity of vacuolation of zona fasciculata cells in the adrenal cortex in both sexes
For details please refer to Table 14 in "any other information on results incl. tables".
Terminal examinations:
1000 ppm: centrilobular hepatocytes with cytoplasmic changes and/or hypertrophy more frequently and decreasing tendency for pituitary hyperplasia as well as for degenerative alterations in the adrenals, tongue, heart and kidney in females
3000/4000 ppm: eosinophilic foci, focal degenerative changes (periportal) and cytoplasmic changes of hepatocytes (periportal and centrilobular) in the liver of males, peliotic foci, single cell necrosis and cytoplasmic changes of hepatocytes (centrilobular) and/or hypertrophy in the liver of females;
increased number of animals with a focal or diffuse simple transitional hyperplasia in the urinary bladder, increases of Proliferating Cell Nuclear Antigen (PCNA) labelling index of the urinary bladder in the hyperplastic segment of the transitional epithelium of females;
increased incidence of follicular hyperplasia (males) and mineralization of the follicular colloid (both sexes) in thyroid glands;
increased number of males with increase in vacuolation of cells in the zona fasciculata of the adrenal glands;
higher frequency of atrophy and/or fibre degenerations of the skeletal muscle in females;
markedly higher incidence and degree of degenerative myelinopathy in the sciatic nerve in both sexes;
decreasing tendency for pituitary hyperplasias as well as for degenerative alterations in the adrenals, tongue, heart and kidney in females;
dilated glands in the stomach mucosa and increase in pigment storage in the spleen of females
For details please refer to Table 15 in "any other information on results incl. tables". - Histopathological findings: neoplastic:
- effects observed, treatment-related
- Description (incidence and severity):
- Interim examinations:
In animals scheduled for interim kill, a no dose-related occurrence of a tumour type was evident.
Terminal examinations:
1000 ppm: adenomas in the thyroid follicles of one male
3000/4000 ppm: one transitional cell carcinoma and two transitional cell papillomas in the urinary bladder of females and one transitional carcinoma in the urethra of a male;
adenomas/carcinoma in the thyroid follicles of males (incidence per increasing dose group: 0-0-0-1/0-2/1)
For details please refer to Tables 16 to 19 in "any other information on results incl. tables". - Other effects:
- effects observed, treatment-related
- Description (incidence and severity):
- ChE activity plasma
No relevant treatment-related effects observed.
ChE activity erythrocytes
1000 ppm: significantly reduced ChE activity in both sexes
3000/4000 ppm: significantly reduced activity in both sexes
ChE activity brain
1000 ppm: reduced ChE activity in females
3000/4000 ppm: reduced ChE activity in both sexes
For details please refer to Table 9 in "any other information on results incl. tables". - Details on results:
- Mortality:
The number of males which had died unscheduled was equally distributed among the groups up to 3000 ppm. In all female treatment groups a higher mortality is evident from week 52 onwards. Since a dose correlation is lacking in the wide dose range (50 and 4000 ppm) a treatment effect is not assumed. Neither for males nor for females did the global comparison of the survival curves (Wilcoxon Test) gave any indication of statistically relevant differences between the groups (p = 0.5782).
Body weight and weight changes:
There were no significant and dosedependent effects on body weights up to 1000 ppm in males. Males ingesting 3000 ppm gained less body weight from week 7 onwards with a maximum difference of 11.0% in week 105. The differences were significant in most cases. In female rats there is also no toxicologially relevant effect on body weights up to the dose of 1000 ppm. The few significantly lower means at 1000 ppm (between weeks 43 to 61) are not interpreted as an adverse effect, since deviations to control data are very small (maximal 6.3% in week 51). At 4000 ppm a significant (from week 13 onwards) body weight depression was observed with a maximum of 14.3% in week 85.
Food consumption and compound intake:
The food intake per animal at doses of up to 3000 ppm (males) or 4000 ppm (females) was comparable to that of untreated rats. The slightly elevated intake data per kg body weight in rats receiving 3000 (12.3%) or 4000 ppm (19.2%) could be interpreted as an indication of a slightly reduced feed efficiency.
Opthalmological findings:
Males of the 1000 and 3000 ppm groups showed a higher incidence of moderate or severe opacities in the (whole) lens cortex. The highest frequency of lenses showing slight (wedge shaped) waterclefts, being a prestage of cortical opacities, was noted in 0 ppm males. This incidence distribution indicates a shift of lens waterclefts towards cortical opacities in 1000 and 3000 ppm males. In 4000 ppm females no remarkable lens findings could be detected compared to controls. Concerning the remaining eye compartments no treatment effect is visible.
Haematological findings:
There were no toxicologically relevant changes in the erythrocyte parameters (count, MVC, MCHC and MCH), in the hemoglobin concentration or in the hematocrit in males or females up to 1000 ppm. Some significant differences in these groups and parameters are not considered as a sign of toxicity, since deviations to control values were very small (MCV, MCH and elevated MCHC means in treated males in week 26/27) and/or a dose correlation was lacking (erythrocyte counts). The hemoglogin concentration and hematocrit values were significantly reduced in 3000 males at week 26/27, but not thereafter. Other significant changes in this group were slight and without corroborating findings. In 4000 ppm females no significant changes were seen with regard to hemoglobin concentration, hematocrit and erythrocyte counts but the calculation of MCH and MCHC revealed consistently lower (partly p<0.01) values than in controls and remaining groups. Analysis of erythrocyte morphology did not reveal abnormalities that could be regarded as a toxic effect. No relevant changes in platelet counts were found in treated males and females. The very slight increase in the means of platelet count of 4000 ppm females in week 26/27 and 52 do not reflect an adverse effect, since individual values were within the reference range. The means of the thromboplastin time in the treatment groups were not remarkably different from those at 0 ppm. No changes of reticulocytes was observed up to highest dose level in males and females. No toxicologically relevant changes in leukocyte counts and differential blood counts were visible in the dose range investigated.
Clinical biochemistry findings:
Neither male nor female rats showed any remarkable changes in the plasma activity of aspartate aminotransferase (ASAT), alkaline phosphatase (APh) and glutamate dehydrogenase (GLDH) up to 200 ppm. At higher doses there was a tendency towards slightly (sometimes significantly) lower aspartate aminotransferase (ASAT) and alkaline phosphatase (APh) activities. These changes were not considered to indicate an adverse effect, since the values were within the physiological range. The activity of the alanine aminotransferase (ALAT) was unchanged in all treatment groups. A trend towards slightly elevated gamma glutamyltransferase (GOT) activity was at 3000 or 4000 ppm (p<0.01), but not at all dates and without relevant time dependent increase.
Urinalysis findings:
In week 98/99 quantitative pH measurements were performed by a microelectrode. Results of these determinations do not indicate any influence of the test substance on the urine pH in the dose range investigate. Scanning-electron-microscopy of urine sediment did not reveal any indication of a change in amount or morphology of urine crystals due to the treatment. Quantitative urinalyses did not reveal a treatment-related effect on proteinurea, urine volume or urine density. Significant deviations from control values in these parameters were not considered relevant, since dose correlations were lacking.
Organ weight findings including organ body weight ratios:
At interim necropsy all absolute and relative organ weights except the liver weights were comparable with those of controls. A few significant changes can be attributed to differences in body weight. The liver weights were not remarkably changed up to 1000 ppm. In 3000 ppm males they were significantly increased by 18.5% (absolute) and 17.6% (relative), whereas in 4000 ppm females the increase was 18.8% (absolute) and 41.7% (relative). At terminal necropsy there were no toxicologically relevant differences between the dose groups and the controls with respect to the weights of the brain, adrenals, heart, kidneys or ovaries. The few differences marked as significant in these organs were very small and attributable to varying body weights. The liver weights were inconspicuous up to 1000 ppm. At higher concentrations absolute and relative liver weights were increased (partly p<0.01). The relative means were 20.2% ( 3000 ppm males) or 32.9% (4000 ppm females) above the corresponding control value. Up to 1000 ppm spleen and testis weights were unremarkable. At the top concentration levels slightly lower (up to -23%) spleen weights (absolute and relative) were seen in both sexes and the relative testes weights were elevated (p<0.01) by about 17%. These deviations were partly significant.
Gross pathological findings:
No treatment related macroscopical findings were observed in rats scheduled for a treatment period of 12 months. Gross-pathological findings recorded at the necropsies performed during the in life phase in case of unscheduled deaths or at the end of the study revealed no evidence of dose-related organ lesions in any of the groups up to and including doses of 3000 or 4000 ppm. There were six 3000 ppm males and five 4000 ppm females that appeared skinny at necropsy.
Histpathological findings (non neoplastic):
In animals scheduled for a 12-months treatment revealed test substance-related lesions in the adrenals, liver and urinary bladder. In the adrenal cortex a slight increase in the incidence and severity of vacuolation of the zona fasciculata cells was recorded at 3000 or 4000 ppm. A minimal to moderate hepatocellular hypertrophy, predominantly centrolobular, was noted in rats at 1000 ppm and above.
In the urinary bladder a diffuse hyperplasia (minimal to slight) was observed in the transitional epithelium at 200 ppm and above (males) or in the group 4000 ppm (females). For the lesions in the urinary bladder, liver and adrenals a statistically significant trend was established. All other microscopical findings recorded were considered to be incidental. In animals scheduled for terminal kill, there were no treatment-related non-neoplastic lesions at concentrations of up to 1000 ppm in males and 200 ppm in females. In the liver of 3000 ppm males eosinophilic foci, focal degenerative changes (periportal) and cytoplasmic changes of hepatocytes (periportal and centrilobular) were noted with a markedly higher incidence than in the other groups. These findings were associated with a reduced number of clear cell foci at 3000 ppm. In males of the groups 0, 50, 200 and 1000 ppm and in all treated females the frequency of all these findings did not exceed the reference range of rats of this age. In females centrilobular hepatocytes with cytoplasmic changes and/or hypertrophy were detected more frequently from 1000 ppm onwards. Additionally, in 4000 ppm females peliotic foci and single cell necroses were noted with increased incidences. Most of the liver lesions mentioned were found to show a statistically significant positive trend. The number of animals with a focal or diffuse simple transitional hyperplasia in the urinary bladder was increased (trend mostly p<0.01) in high dose males and females. In 4000 ppm females atrophy (p<0.01) and/or fibre degenerations (p<0.05) of the skeletal muscle were found with a higher frequency than in the other groups. In 3000 ppm males and 4000 ppm females a degenerative myelinopathy was detected in the sciatic nerve with a markedly higher incidence (trend p<0.01) and a higher degree than in the other groups, where the frequency of this lesion did not exceed the range normally found in old rats. In the thyroid glands the incidence of follicular hyperplasia (males) and mineralization of the follicular colloid (males and females) was significantly increased in 3000/4000 ppm rats. At 3000 ppm there were more (trend p<0.01) males exhibiting an increase in vacuolation of cells in the zona fasciculata of the adrenal glands. Statistically significant increases of proliferating cell nuclear antigen (PCNA) labeling index of the urinary bladder were only achieved in the hyperplastic segment of the transitional epithelium of 4000 ppm females. In the bladder of 1000 ppm females and in the non-hyperplastic bladder segment of the 4000 ppm rats the calculated values were similar to those of control females.The remaining observed findings were considered to be secondary possibly due to retarded aging, which is often seen in toxicological studies, if body weight is reduced as in the present study. These findings included a decreasing tendency for pituitary hyperplasias as well as for degenerative alterations in the adrenals, tongue, heart and kidney essentially in 1000 and 4000 ppm females. The elevated (p<0.05) number of 4000 ppm females showing dilated glands in the stomach mucosa was considered unspecific and not treatment-related. Also the increase in pigment storage in the spleen (p<0.01) was not considered treatment-related.
Histpathological findings (neoplastic):
In animals scheduled for interin kill, a no dose-related occurrence of a tumor type was evident. At the terminal kill, one transitional cell carcinoma and two transitional cell papillomas (trend p<0.01) were detected in the urinary bladder of 4000 ppm females, whereas no bladder tumor occurred in the remaining female groups or in males. Additionally, one transitional carcinoma was seen in the urethra of a 3000 ppm male. In thyroid follicles of males adenomas were observed with a frequency of 0-0-0-1-2 (trend p<0.05) and carcinomas with an incidence of 0-0-0-0-1. Uterine adenocarcinomas were noted in 4000 ppm females more often (trend p<0.01) than in the other groups. Additionally, there was one squamous cell carcinoma (p<0.05) in this group. However, the incidences of both tumor types do not exceed the historical control range and are therefore, considered to be not compound-induced. Decreased tumor incidences were noted for the mammary fibroadenomas (15-10-7-6-5) and pituitary adenomas (24-18-24-14-11) in the female treatment groups. The incidences of the remaining tumors were not distributed in a treatment-related manner and comparable with those known from own historical control collectives.The number of cases, in which a tumor type occurred in one group only was comparable in the 3000/4000 ppm and 0 ppm group (Please refer to table 16 in the "any other information on results incl. tables " section). The overall tumor incidence and the total number of benign and malignant neoplasms per group were not increased in treated males and females at all concentrations (Please refer to table 17 in the "any other information on results incl. tables " section). The number of malignant tumors was even reduced in males receiving 200 ppm and above. No remarkable differences were observed deaths in the number of tumor bearing rats or animals with a benign, malignant or benign and malignant tumor between the control and treatment groups for both, intercurrent or scheduled deaths (Please refer to table 18 in the "any other information on results incl. tables " section). As regards the time of occurrence of the tumors, there were no significant differences between treated rats and controls (Please refer to table 19 in the "any other information on results incl. tables " section).
Other findings: Cholinesterase activities (CHE)
No significant CHE inhibition occurred in the plasma. The 24% lower plasma CHE activity of 3000 ppm males (week 78) is not regarded as toxicologically relevant, because this was an isolated event, where in the control group relatively high activities were measured compared to those of the other time points. In addition, plasma CHE activity was even not affected when e.g. CHE activity in erythrocytes was inhibited by 90%. No remarkable CHE inhibition in erythrocytes were detected in males and females up to 200 ppm. At higher concentrations a dose-dependently and significantly lower CHE activity (inhibited up to 82% in males and 90% in females) was measured compared to control means. The significantly lower activity in 200 ppm females in week 52/53 is considered to be incidental, because activity at this dose level was not remarkably inhibited at the other three time points, where stronger effects were noted in the two highest dose groups. Also in week 52/53, a relatively high activity in controls may have contributed. Up to 3000 ppm (males) and 200 ppm (females) no significant or biologically relevant effect was noted on the brain CHE activity. In females there was a significantly reduced activity at 1000 and 4000 ppm (up to 41%). The inhibition values of 13 and 17% in 1000 ppm females and those of 14% calculated for 3000 ppm males at termination might be of questionable toxicological relevance. - Relevance of carcinogenic effects / potential:
- Mortality:
The number of males which had died unscheduled was equally distributed among the groups up to 3000 ppm. In all female treatment groups a higher mortality is evident from Week 52 onwards. Since a dose correlation is lacking in the wide dose range (50 and 4000 ppm) a treatment effect is not assumed. Neither for males nor for females did the global comparison of the survival curves (Wilcoxon Test) gave any indication of statistically relevant differences between the groups (p = 0.5782).
Body weight and weight changes:
There were no significant and dose-dependent effects on body weights up to 1000 ppm in males. Males ingesting 3000 ppm gained less body weight from week 7 onwards with a maximum difference of 11.0% in Week 105. The differences were significant in most cases. In female rats there is also no toxicologically relevant effect on body weights up to the dose of 1000 ppm. The few significantly lower means at 1000 ppm (between Weeks 43 to 61) are not interpreted as an adverse effect, since deviations to control data are very small (maximal 6.3% in Week 51). At 4000 ppm a significant (from Week 13 onwards) body weight depression was observed with a maximum of 14.3% in Week 85.
Food consumption and compound intake:
The food intake per animal at doses of up to 3000 ppm (males) or 4000 ppm (females) was comparable to that of untreated rats. The slightly elevated intake data per kg body weight in rats receiving 3000 (12.3%) or 4000 ppm (19.2%) could be interpreted as an indication of a slightly reduced feed efficiency.
Opthalmological findings:
Males of the 1000 and 3000 ppm groups showed a higher incidence of moderate or severe opacities in the (whole) lens cortex. The highest frequency of lenses showing slight (wedge shaped) waterclefts, being a pre-stage of cortical opacities, was noted in control males. This incidence distribution indicates a shift of lens waterclefts towards cortical opacities in 1000 and 3000 ppm males. In 4000 ppm females no remarkable lens findings could be detected compared to controls. Concerning the remaining eye compartments no treatment effect is visible.
Haematological findings:
There were no toxicologically relevant changes in the erythrocyte parameters (count, MVC, MCHC and MCH), in the haemoglobin concentration or in the haematocrit in males or females up to 1000 ppm. Some significant differences in these groups and parameters are not considered as a sign of toxicity, since deviations to control values were very small (MCV, MCH and elevated MCHC means in treated males in Week 26/27) and/or a dose correlation was lacking (erythrocyte counts). The haemoglobin concentration and haematocrit values were significantly reduced in 3000 males at Week 26/27, but not thereafter. Other significant changes in this group were slight and without corroborating findings. In 4000 ppm females no significant changes were seen with regard to haemoglobin concentration, haematocrit and erythrocyte counts but the calculation of MCH and MCHC revealed consistently lower (partly p<0.01) values than in controls and remaining groups. Analysis of erythrocyte morphology did not reveal abnormalities that could be regarded as a toxic effect. No relevant changes in platelet counts were found in treated males and females. The very slight increase in the means of platelet count of 4000 ppm females in Weeks 26/27 and 52 do not reflect an adverse effect, since individual values were within the reference range. The means of the thromboplastin time in the treatment groups were not remarkably different from those at 0 ppm. No changes of reticulocytes were observed up to the highest dose level in both sexes. No toxicologically relevant changes in leukocyte counts and differential blood counts were visible in the dose range investigated.
Clinical biochemistry findings:
Neither male nor female rats showed any remarkable changes in the plasma activity of aspartate aminotransferase (ASAT), alkaline phosphatase (APh) and glutamate dehydrogenase (GLDH) up to 200 ppm. At higher doses there was a tendency towards slightly (sometimes significantly) lower aspartate aminotransferase (ASAT) and alkaline phosphatase (APh) activities. These changes were not considered to indicate an adverse effect, since the values were within the physiological range. The activity of the alanine aminotransferase (ALAT) was unchanged in all treatment groups. A trend towards slightly elevated gamma glutamyltransferase (GOT) activity was at 3000 or 4000 ppm (p<0.01), but not at all dates and without relevant time dependent increase.
Urinalysis findings:
In Week 98/99 quantitative pH measurements were performed by a microelectrode. Results of these determinations do not indicate any influence of the test substance on the urine pH in the dose range investigated. Scanning-electron-microscopy of urine sediment did not reveal any indication of a change in amount or morphology of urine crystals due to the treatment. Quantitative urinalyses did not reveal a treatment-related effect on proteinurea, urine volume or urine density. Significant deviations from control values in these parameters were not considered relevant, since dose correlations were lacking.
Organ weight findings including organ body weight ratios:
At interim necropsy all absolute and relative organ weights except the liver weights were comparable with those of controls. A few significant changes can be attributed to differences in body weight. The liver weights were not remarkably changed up to 1000 ppm. In 3000 ppm males they were significantly increased by 18.5% (absolute) and 17.6% (relative), whereas in 4000 ppm females the increase was 18.8% (absolute) and 41.7% (relative). At terminal necropsy there were no toxicologically relevant differences between the dose groups and the controls with respect to the weights of the brain, adrenals, heart, kidneys or ovaries. The few differences marked as significant in these organs were very small and attributable to varying body weights. The liver weights were inconspicuous up to 1000 ppm. At higher concentrations absolute and relative liver weights were increased (partly p<0.01). The relative means were 20.2% ( 3000 ppm males) or 32.9% (4000 ppm females) above the corresponding control value. Up to 1000 ppm spleen and testis weights were unremarkable. At the top concentration levels slightly lower (up to -23%) spleen weights (absolute and relative) were seen in both sexes and the relative testes weights were elevated (p<0.01) by about 17%. These deviations were partly significant.
Gross pathological findings:
No treatment-related macroscopical findings were observed in rats scheduled for a treatment period of 12 months. Gross-pathological findings recorded at the necropsies performed during the in life phase in case of unscheduled deaths or at the end of the study revealed no evidence of dose-related organ lesions in any of the groups up to and including doses of 3000 or 4000 ppm. There were six 3000 ppm males and five 4000 ppm females that appeared skinny at necropsy.
Histopathological findings (non-neoplastic):
In animals scheduled for a 12-months treatment revealed test substance-related lesions in the adrenals, liver and urinary bladder. In the adrenal cortex a slight increase in the incidence and severity of vacuolation of the zona fasciculata cells was recorded at 3000 or 4000 ppm. A minimal to moderate hepatocellular hypertrophy, predominantly centrolobular, was noted in rats at 1000 ppm and above.
In the urinary bladder a diffuse hyperplasia (minimal to slight) was observed in the transitional epithelium at 200 ppm and above (males) or in the group 4000 ppm (females). For the lesions in the urinary bladder, liver and adrenals a statistically significant trend was established. All other microscopical findings recorded were considered to be incidental. In animals scheduled for terminal kill, there were no treatment-related non-neoplastic lesions at concentrations of up to 1000 ppm in males and 200 ppm in females. In the liver of 3000 ppm males eosinophilic foci, focal degenerative changes (periportal) and cytoplasmic changes of hepatocytes (periportal and centrilobular) were noted with a markedly higher incidence than in the other groups. These findings were associated with a reduced number of clear cell foci at 3000 ppm. In males of the groups 0, 50, 200 and 1000 ppm and in all treated females the frequency of all these findings did not exceed the reference range of rats of this age. In females centrilobular hepatocytes with cytoplasmic changes and/or hypertrophy were detected more frequently from 1000 ppm onwards. Additionally, in 4000 ppm females peliotic foci and single cell necrosis were noted with increased incidences. Most of the liver lesions mentioned were found to show a statistically significant positive trend. The number of animals with a focal or diffuse simple transitional hyperplasia in the urinary bladder was increased (trend mostly p<0.01) in high dose males and females. In 4000 ppm females atrophy (p<0.01) and/or fibre degenerations (p<0.05) of the skeletal muscle were found with a higher frequency than in the other groups. In 3000 ppm males and 4000 ppm females a degenerative myelinopathy was detected in the sciatic nerve with a markedly higher incidence (trend p<0.01) and a higher degree than in the other groups, where the frequency of this lesion did not exceed the range normally found in old rats. In the thyroid glands the incidence of follicular hyperplasia (males) and mineralization of the follicular colloid (males and females) was significantly increased in 3000/4000 ppm rats. At 3000 ppm there were more (trend p<0.01) males exhibiting an increase in vacuolation of cells in the zona fasciculata of the adrenal glands. Statistically significant increases of proliferating cell nuclear antigen (PCNA) labeling index of the urinary bladder were only achieved in the hyperplastic segment of the transitional epithelium of 4000 ppm females. In the bladder of 1000 ppm females and in the non-hyperplastic bladder segment of the 4000 ppm rats the calculated values were similar to those of control females.The remaining observed findings were considered to be secondary possibly due to retarded aging, which is often seen in toxicological studies, if body weight is reduced as in the present study. These findings included a decreasing tendency for pituitary hyperplasia as well as for degenerative alterations in the adrenals, tongue, heart and kidney essentially in 1000 and 4000 ppm females. The elevated (p<0.05) number of 4000 ppm females showing dilated glands in the stomach mucosa was considered unspecific and not treatment-related. Also the increase in pigment storage in the spleen (p<0.01) was not considered treatment-related.
Histpathological findings (neoplastic):
In animals scheduled for interim kill, a no dose-related occurrence of a tumour type was evident. At the terminal kill, one transitional cell carcinoma and two transitional cell papillomas (trend p<0.01) were detected in the urinary bladder of 4000 ppm females, whereas no bladder tumour occurred in the remaining female groups or in males. Additionally, one transitional carcinoma was seen in the urethra of a 3000 ppm male. In thyroid follicles of males adenomas were observed with a frequency of 0-0-0-1-2 (trend p<0.05) and carcinomas with an incidence of 0-0-0-0-1. Uterine adenocarcinomas were noted in 4000 ppm females more often (trend p<0.01) than in the other groups. Additionally, there was one squamous cell carcinoma (p<0.05) in this group. However, the incidences of both tumour types do not exceed the historical control range and are therefore, considered to be not compound-induced. Decreased tumour incidences were noted for the mammary fibroadenomas(15-10-7-6-5) and pituitary adenomas (24-18-24-14-11) in the female treatment groups. The incidences of the remaining tumours were not distributed in a treatment-related manner and comparable with those known from own historical control collectives. The number of cases, in which a tumour type occurred in one group only was comparable in the 3000/4000 ppm and 0 ppm group (for details please refer to Tables 16 in "any other information on results incl. tables".). The overall tumour incidence and the total number of benign and malignant neoplasms per group were not increased in treated males and females at all concentrations (for details please refer to Table 17 in "any other information on results incl. tables"). The number of malignant tumours was even reduced in males receiving 200 ppm and above. No remarkable differences were observed deaths in the number of tumour bearing rats or animals with a benign, malignant or benign and malignant tumour between the control and treatment groups for both, intercurrent or scheduled deaths (for details please refer to Table 18 in "any other information on results incl. tables"). As regards the time of occurrence of the tumours, there were no significant differences between treated rats and controls (for details please refer to Table 19 in "any other information on results incl. tables").
Other findings: Cholinesterase activities (ChE)
No significant ChE inhibition occurred in the plasma. The 24% lower plasma ChE activity of 3000 ppm males (Week 78) is not regarded as toxicologically relevant, because this was an isolated event, where in the control group relatively high activities were measured compared to those of the other time points. In addition, plasma ChE activity was even not affected when e.g. ChE activity in erythrocytes was inhibited by 90%. No remarkable ChE inhibition in erythrocytes were detected in males and females up to 200 ppm. At higher concentrations a dose-dependently and significantly lower ChE activity (inhibited up to 82% in males and 90% in females) was measured compared to control means. The significantly lower activity in 200 ppm females in Week 52/53 is considered to be incidental, because activity at this dose level was not remarkably inhibited at the other three time points, where stronger effects were noted in the two highest dose groups. Also in Week 52/53, a relatively high activity in controls may have contributed. Up to 3000 ppm (males) and 200 ppm (females) no significant or biologically relevant effect was noted on the brain ChE activity. In females there was a significantly reduced activity at 1000 and 4000 ppm (up to 41%). The inhibition values of 13 and 17% in 1000 ppm females and those of 14% calculated for 3000 ppm males at termination might be of questionable toxicological relevance. - Key result
- Dose descriptor:
- NOAEL
- Remarks:
- neoplastic
- Effect level:
- 1 000 ppm
- Based on:
- test mat.
- Sex:
- male/female
- Basis for effect level:
- other: no carcinogenic effects observed up to this dose level
- Remarks on result:
- other: corresponding to: 52.7 mg/kg bw/day (males) / 75.4 mg/kg bw/day (females)
- Key result
- Dose descriptor:
- LOAEL
- Remarks:
- neoplastic
- Effect level:
- 3 000 ppm
- Based on:
- test mat.
- Sex:
- male
- Basis for effect level:
- histopathology: neoplastic
- Remarks on result:
- other: corresponding to: 170.4 mg/kg bw/day
- Key result
- Dose descriptor:
- LOAEL
- Remarks:
- neoplastic
- Effect level:
- 4 000 ppm
- Based on:
- test mat.
- Sex:
- female
- Basis for effect level:
- histopathology: neoplastic
- Remarks on result:
- other: corresponding to: 326.7 mg/kg bw/day
- Key result
- Critical effects observed:
- yes
- Lowest effective dose / conc.:
- 1 000 ppm
- System:
- nervous system
- Organ:
- other: cholinesterase activity inhibited in erythrocytes (1000 ppm and above) and brain (4000 ppm) (statistically significant inhibition by 20% or more in erythrocytes or brain is considered a clear toxicological effect, please refer to chapter 7.5.1 1997f)
- Treatment related:
- yes
- Dose response relationship:
- yes
- Relevant for humans:
- not specified
Table 1: Cumulative Mortality data (number of dead animals up to week indicated)
Dose (ppm) |
0 |
50 |
200 |
1000 |
3000 |
sex (male) |
|||||
Weeks 0-52 (n = 60) |
|||||
Week 26 |
0 |
1 |
0 |
0 |
0 |
Week 53 |
0 |
1 |
0 |
2 |
1 |
Weeks 53-104 (n = 50) |
|||||
Week 79 |
5 |
4 |
1 |
5 |
1 |
Week 104 |
19 |
17 |
21 |
17 |
13 |
Dose (ppm) |
0 |
50 |
200 |
1000 |
4000 |
sex (female) |
|||||
Weeks 0-52 (n = 60) |
|||||
Week 26 |
0 |
0 |
0 |
0 |
0 |
Week 53 |
0 |
2 |
1 |
0 |
2 |
Weeks 53-104 (n = 50) |
|||||
Week 79 |
3 |
7 |
8 |
8 |
9 |
Week 104 |
13 |
19 |
22 |
20 |
21 |
Table 2: Body weights [g]
sex |
m |
m |
m |
m |
m |
f |
f |
f |
f |
f |
Dose (ppm) |
0 |
50 |
200 |
1000 |
3000 |
0 |
50 |
200 |
1000 |
4000 |
0 1 2 3 4 5 6 7 8 9 10 11 12 13 15 17 19 21 23 25 27 29 31 33 35 37 39 41 43 45 47 49 51 53 55 57 59 61 63 65 67 69 71 73 75 77 79 81 83 85 87 89 91 93 95 97 99 101 103 105 |
155 203 239 272 299 322 336 362 378 382 393 403 412 427 435 456 463 471 472 477 482 492 492 502 500 510 515 519 509 517 519 523 526 531 532 535 535 542 542 542 543 553 552 553 558 550 551 558 555 559 554 553 550 549 550 553 551 544 542 543 |
156 203 243 276 304 325 344 362 378 394 406 417 428 436 448 465 468 479 480 486 494 498 503 509 507 515 518 521 519 523 521 528 532 530 531 534 534 536 540 539 541 548 549 548 550 552 543 551 549 548 542 537 529 530 529 534 530 520 508 + 511 |
155 203 245 278 306 329 348 364 378 395 406 417 423 430 444 459 463 470 480 487 493 495 503 507 509 508 514 516 515 518 520 524 529 533 534 538 539 539 541 541 542 547 545 541 544 539 536 536 538 532 520 + 522 513 + 527 528 522 519 510 + 514 529 |
154 200 241 275 301 326 341 356 368 385 401 400 417 420 438 448 455 464 472 480 483 488 496 495 502 509 507 510 512 512 513 519 522 525 527 530 534 533 532 538 542 549 547 547 550 550 552 556 555 553 549 547 533 542 535 531 528 523 518 515 |
152 189++ 238 264 289 312 330 344++ 360++ 372 382 391 401 401++ 421 427++ 441+ 449+ 454 463 466 468+ 480 479+ 485 483++ 486++ 489++ 492 490++ 490++ 496++ 500++ 502++ 505+ 506++ 513 511++ 512++ 515+ 516+ 521++ 519++ 516++ 518++ 512 ++ 523 + 524 ++ 527 + 526 + 518 + 513 ++ 493 ++ 504 ++ 498 ++ 497 ++ 492 ++ 495 ++ 491 ++ 483 ++ |
130 149 163 176 186 196 205 212 217 222 225 229 234 242 242 247 249 255 258 260 261 267 271 277 276 280 284 286 289 290 295 296 301 302 304 309 311 312 314 315 319 323 322 325 329 333 328 333 333 336 327 327 329 327 326 328 323 323 318 313 |
127 145 161 173 183 191 200 208 212 216 222 226 230 234 240 243 247 251 254 257 257 261 266 271 273 275 279 281 284 285 289 288 291 291 294 298 300 303 307 307 309 316 314 318 321 323 321 329 330 331 326 325 324 324 325 327 325 324 315 325 |
130 150 166 179 190 199 207 211 217 223 228 232 238 239 244 249 252 258 260 263 264 266 272 276 280 281 285 288 289 289 293 293 297 302 304 307 309 308 312 315 316 322 322 325 327 328 327 332 333 337 332 330 331 327 329 326 325 320 312 341 |
128 147 164 178 186 196 203 207 213 219 221 228 231 234 237 242 246 249 252 254 257 257 261 266 270 270 274 276 276+ 275+ 280+ 279++ 282++ 284+ 287+ 291+ 292+ 294+ 300 301 303 308 307 311 313 314 310 320 320 320 316 314 317 317 313 311 311 312 307 320 |
130 147 167 179 189 196 202 205 209 215 217 222 225 226++ 230+ 234++ 234++ 238++ 242++ 245++ 246++ 248++ 252++ 253++ 257++ 258++ 262++ 264++ 264++ 263++ 266++ 266++ 268++ 270++ 271++ 274++ 275++ 279++ 278++ 280++ 281++ 285++ 283++ 283++ 286++ 284 ++ 284 ++ 289 ++ 291 ++ 288 ++ 286 ++ 288 ++ 287 ++ 289 ++ 289 ++ 286 ++ 283 ++ 284 ++ 281 ++ 295 |
+ difference against control for p<0.05
++ difference against control for p<0.01
Table 3: Mean daily and cumulative intake of food and test compound
Intake food |
||||||
|
|
|
g/animal |
g/kg bw |
||
Dose (ppm) |
Sex |
Days |
Total |
per Day |
Total |
per Day |
0 |
m |
708 |
14715 |
20.8 |
35813 |
50.6 |
50 |
m |
708 |
14923 |
21.1 |
35653 |
50.4 |
200 |
m |
708 |
15169 |
21.4 |
36331 |
51.3 |
1000 |
m |
708 |
15391 |
21.7 |
37307 |
52.7 |
3000 |
m |
708 |
15807 |
22.3 |
40218 |
56.8 |
0 |
f |
708 |
12072 |
17.1 |
48515 |
68.5 |
50 |
f |
708 |
12522 |
17.7 |
51527 |
72.8 |
200 |
f |
708 |
12729 |
18.0 |
51524 |
72.8 |
1000 |
f |
708 |
12712 |
18.0 |
53372 |
75.4 |
4000 |
f |
708 |
13177 |
18.6 |
57821 |
81.7 |
Intake of test compound |
||||||
|
|
|
mg/animal |
mg/kg bw |
||
Dose (ppm) |
Sex |
Days |
Total |
per Day |
Total |
per Day |
50 |
m |
708 |
746 |
1.1 |
1783 |
2.5 |
200 |
m |
708 |
3034 |
4.3 |
7266 |
10.3 |
1000 |
m |
708 |
15391 |
21.7 |
37307 |
52.7 |
3000 |
m |
708 |
47421 |
67.0 |
120654 |
170.4 |
50 |
f |
708 |
626 |
0.9 |
2576 |
3.6 |
200 |
f |
708 |
2546 |
3.6 |
10305 |
14.6 |
1000 |
f |
708 |
12712 |
18.0 |
53372 |
75.4 |
4000 |
f |
708 |
52707 |
74.4 |
231285 |
326.7 |
Table 4: Mean daily cumulative water intake
Intake of water |
||||||
|
|
|
g/animal |
g/kg bw |
||
Dose (ppm) |
Sex |
Days |
Total |
per Day |
Total |
per Day |
0 |
m |
708 |
18267 |
25.9 |
40827 |
57.8 |
50 |
m |
708 |
18971 |
26.9 |
41396 |
58.6 |
200 |
m |
708 |
19254 |
27.3 |
42187 |
59.8 |
1000 |
m |
708 |
19034 |
27.0 |
42158 |
59.7 |
3000 |
m |
708 |
19391 |
27.5 |
44915 |
63.6 |
0 |
f |
708 |
18478 |
26.2 |
67633 |
96.8 |
50 |
f |
708 |
18745 |
26.6 |
70078 |
99.3 |
200 |
f |
708 |
18848 |
26.7 |
68707 |
97.3 |
1000 |
f |
708 |
18539 |
26.3 |
70478 |
99.8 |
4000 |
f |
708 |
17344 |
24.6 |
69761 |
98.8 |
Table 5: Ophthalmological findings at termination (Incidences in %)
sex |
m |
m |
m |
m |
f |
f |
Dose (ppm) |
0 |
200 |
1000 |
3000 |
0 |
4000 |
No. of eyes examined |
61 |
71 |
61 |
73 |
76 |
58 |
Turbid cornea, including vascularisation of cornea |
1.6 |
7.0 |
1.6 |
8.2 |
0 |
3.4 |
Turbid cornea, including vascularisation of cornea Snow ball like turbidities between lens and vitreous body (= posterior lenticular rupture) |
11.4 |
4.2 |
8.2 |
13.7 |
7.9 |
0 |
Diffuse retrolenticular opacity (all degrees) |
24.6 |
9.8 |
18.0 |
17.8 |
10.5 |
8.6 |
Wedge shaped waterclefts in the cortex, slight |
18.0 |
7.0 |
8.2 |
8.2 |
3.9 |
5.1 |
Wedge shaped waterclefts in the cortex, moderate |
0 |
0 |
3.3 |
1.4 |
0 |
1.7 |
Lens opacity in the whole cortex, slight |
1.6 |
1.4 |
0 |
2.7 |
2.6 |
1.7 |
Lens opacity in the whole cortex, moderate |
1.6 |
1.4 |
1.6 |
5.5 |
3.9 |
0 |
Lens opacity in the whole cortex, severe |
1.6 |
0 |
6.5 |
5.5 |
6.6 |
3.4 |
Table 6: Hematology
Dose |
|
|
LEU |
ERY |
HB |
HCT |
MCV |
MCH |
MCHC |
RETI |
HEINZ |
THRO |
HQUICK |
|
ppm |
Sex |
Week |
109/L |
109/L |
g/L |
L/L |
fl |
pg |
g/LERY |
0/00 |
0/00 |
109/L |
sec |
|
0 |
m |
26/27 |
10.3 |
9.04 |
147 |
0.459 |
50.8 |
16.3 |
320 |
18 |
0 |
1040 |
25.4 |
|
50 |
m |
26/27 |
8.7 |
8.91 |
147 |
0.453 |
50.9 |
16.5 |
324 |
19 |
0 |
998 |
25.2 |
|
200 |
m |
26/27 |
8.3 |
8.51 |
141 |
0.439 |
51.6 |
16.5 |
320 |
18 |
0 |
1109 |
24.7 |
|
1000 |
m |
26/27 |
8.3 |
9.06 |
141 |
0.434 |
48.0++ |
15.6 |
325 |
17 |
0 |
1053 |
26.2 |
|
3000 |
m |
26/27 |
8.0 |
8.58 |
138+ |
0.425++ |
49.6 |
16.1 |
325 |
17 |
1 |
982 |
25.4 |
|
0 |
m |
52 |
9.5 |
9.42 |
153 |
0.487 |
51.8 |
16.2 |
313 |
16 |
0 |
1082 |
26.3 |
|
50 |
m |
52 |
8.5 |
9.17 |
149 |
0.479 |
52.3 |
16.3 |
312 |
17 |
0 |
1059 |
26.0 |
|
200 |
m |
52 |
8.4 |
8.87+ |
148 |
0.471 |
53.1 |
16.7 |
314 |
18 |
0 |
1172 |
26.1 |
|
1000 |
m |
52 |
7.4 |
9.37 |
146 |
0.462 |
49.4 |
15.6 |
315 |
20 |
0 |
1082 |
25.4 |
|
3000 |
m |
52 |
8.2 |
9.29 |
146 |
0.472 |
50.8 |
15.7 |
309 |
20 |
0 |
1026 |
26.3 |
|
0 |
m |
78 |
8.0 |
9.04 |
151 |
0.476 |
52.7 |
16.8 |
318 |
17 |
0 |
1091 |
24.7 |
|
50 |
m |
78 |
7.3 |
8.99 |
149 |
0.472 |
52.5 |
16.6 |
316 |
14 |
0 |
1073 |
24.6 |
|
200 |
m |
78 |
6.7 |
8.20+ |
141 |
0.441 |
53.7 |
17.2 |
319 |
16 |
0 |
1277 |
24.0 |
|
1000 |
m |
78 |
6.4 |
8.93 |
141 |
0.441 |
49.5+ |
15.9+ |
321 |
17 |
0 |
1041 |
24.1 |
|
3000 |
m |
78 |
7.5 |
8.77 |
139 |
0.440 |
50.2 |
15.9+ |
316 |
15 |
0 |
998 |
24.1 |
|
0 |
m |
103/104 |
9.2 |
9.26 |
160 |
0.512 |
55.4 |
17.3 |
312 |
20 |
0 |
1220 |
25.9 |
|
50 |
m |
103/104 |
7.9 |
8.63 |
148 |
0.457+ |
52.9 |
17.1 |
322++ |
27 |
0 |
1158 |
25.0 |
|
200 |
m |
103/104 |
7.9 |
8.85 |
154 |
0.473 |
53.5 |
17.5 |
326++ |
16 |
0 |
1156 |
24.2 |
|
1000 |
m |
103/104 |
7.3 |
8.73 |
144 |
0.443++ |
50.9++ |
16.5 |
324++ |
23 |
0 |
1186 |
24.8 |
|
3000 |
m |
103/104 |
6.9 |
9.07 |
150 |
0.466 |
51.5++ |
16.5 |
321+ |
29 |
0 |
1081 |
25.8 |
|
0 |
f |
26/27 |
6.0 |
8.21 |
143 |
0.439 |
53.5 |
17.4 |
325 |
15 |
0 |
915 |
24.5 |
|
50 |
f |
26/27 |
6.2 |
7.94 |
136 |
0.416 |
52.4 |
17.1 |
328 |
15 |
0 |
978 |
23.7 |
|
200 |
f |
26/27 |
5.9 |
8.10 |
139 |
0.422 |
52.2 |
17.2 |
330 |
14 |
0 |
979 |
23.8 |
|
1000 |
f |
26/27 |
6.2 |
8.01 |
137 |
0.424 |
52.8 |
17.1 |
325 |
17 |
0 |
955 |
23.8 |
|
4000 |
f |
26/27 |
5.8 |
8.57 |
139 |
0.434 |
50.6++ |
16.2++ |
320 |
17 |
0 |
1046++ |
22.4 |
|
0 |
f |
52 |
4.6 |
8.03 |
140 |
0.440 |
54.8 |
17.4 |
318 |
20 |
0 |
809 |
24.7 |
|
50 |
f |
52 |
5.8 |
8.15 |
138 |
0.437 |
53.6 |
17.0 |
317 |
18 |
0 |
921 |
25.4 |
|
200 |
f |
52 |
5.7 |
8.26 |
142 |
0.440 |
53.4 |
17.2 |
322 |
17 |
0 |
956 |
25.2 |
|
1000 |
f |
52 |
5.7 |
8.25 |
140 |
0.447 |
54.2 |
17.0 |
314 |
19 |
0 |
897 |
24.1 |
|
4000 |
f |
52 |
5.8 |
8.36 |
136 |
0.440 |
53.1 |
16.4+ |
309++ |
20 |
0 |
1017+ |
23.8 |
|
0 |
f |
78 |
4.1 |
8.33 |
146 |
0.454 |
54.6 |
17.5 |
321 |
25 |
0 |
858 |
23.2 |
|
50 |
f |
78 |
4.6 |
7.79 |
136 |
0.429 |
56.6 |
17.8 |
316 |
44 |
0 |
795 |
23.5 |
|
200 |
f |
78 |
5.6 |
8.24 |
142 |
0.443 |
53.8 |
17.3 |
321 |
31 |
0 |
840 |
23.4 |
|
1000 |
f |
78 |
4.3 |
8.16 |
143 |
0.451 |
55.4 |
17.6 |
317 |
31 |
0 |
848 |
26.2++ |
|
4000 |
f |
78 |
4.7 |
8.06 |
133 |
0.431 |
53.6 |
16.6 |
309++ |
36 |
0 |
930 |
24.8+ |
|
0 |
f |
103/104 |
5.6 |
8.29 |
151 |
0.453 |
54.9 |
18.3 |
333 |
31 |
0 |
1003 |
24.0 |
|
50 |
f |
103/104 |
4.7 |
8.34 |
151 |
0.454 |
54.5 |
18.1 |
333 |
28 |
0 |
984 |
23.7 |
|
200 |
f |
103/104 |
6.8 |
8.22 |
149 |
0.450 |
54.7 |
18.1 |
330 |
26 |
0 |
994 |
24.6 |
|
1000 |
f |
103/104 |
4.3 |
8.33 |
151 |
0.463 |
55.7 |
18.1 |
325 |
29 |
0 |
948 |
24.8 |
|
4000 |
f |
103/104 |
5.1 |
8.22 |
143 |
0.443 |
54.2 |
17.4 |
322+ |
31 |
0 |
1066 |
23.9 |
|
+ difference against control for p<0.05
++ difference against control for p<0.01
Table 7: Differential blood count
Dose |
Week |
Sex |
LYM |
SEGM |
EOS |
MONO |
BAND |
ppm |
|
|
% |
% |
% |
% |
% |
0 |
26/27 |
m |
87.5 |
8.0 |
0.2 |
4.3 |
0.0 |
50 |
26/27 |
m |
89.8 |
6.5 |
0.3 |
3.4 |
0.0 |
200 |
26/27 |
m |
86.0 |
10.1 |
0.6 |
3.3 |
0.0 |
1000 |
26/27 |
m |
87.9 |
8.1 |
0.7 |
3.3 |
0.0 |
3000 |
26/27 |
m |
89.4 |
7.6 |
0.6 |
2.5 |
0.0 |
0 |
52 |
m |
79.7 |
12.7 |
1.0 |
6.6 |
0.0 |
50 |
52 |
m |
78.8 |
11.7 |
1.8 |
7.7 |
0.0 |
200 |
52 |
m |
79.1 |
12.7 |
0.9 |
7.3 |
0.0 |
1000 |
52 |
m |
77.5 |
13.6 |
1.4 |
7.6 |
0.0 |
3000 |
52 |
m |
78.0 |
13.7 |
0.6 |
7.8 |
0.0 |
0 |
78 |
m |
78.4 |
13.9 |
1.5 |
6.2 |
0.0 |
50 |
78 |
m |
75.9 |
16.2 |
2.2 |
5.6 |
0.0 |
200 |
78 |
m |
74.9 |
17.4 |
1.7 |
6.2 |
0.0 |
1000 |
78 |
m |
74.7 |
18.9 |
1.1 |
5.3 |
0.0 |
3000 |
78 |
m |
77.6 |
17.0 |
0.8 |
4.6 |
0.0 |
0 |
103/104 |
m |
67.8 |
23.5 |
1.2 |
7.4 |
0.0 |
50 |
103/104 |
m |
62.7 |
29.7 |
1.7 |
6.0 |
0.0 |
200 |
103/104 |
m |
70.0 |
23.6 |
1.5 |
4.8 |
0.1 |
1000 |
103/104 |
m |
66.6 |
25.4 |
2.6 |
5.5 |
0.0 |
3000 |
103/104 |
m |
71.8 |
23.0 |
1.2 |
4.2 |
0.0 |
0 |
26/27 |
f |
88.4 |
8.4 |
0.8 |
2.5 |
0.0 |
50 |
26/27 |
f |
86.0 |
11.3 |
0.8 |
1.9 |
0.0 |
200 |
26/27 |
f |
89.5 |
7.8 |
0.8 |
1.9 |
0.0 |
1000 |
26/27 |
f |
90.7 |
8.2 |
0.4 |
0.7+ |
0.0 |
4000 |
26/27 |
f |
88.7 |
8.5 |
0.8 |
2.0 |
0.0 |
0 |
52 |
f |
79.9 |
14.0 |
1.4 |
4.7 |
0.0 |
50 |
52 |
f |
76.5 |
16.5 |
1.0 |
6.0 |
0.0 |
200 |
52 |
f |
80.7 |
13.3 |
1.2 |
4.8 |
0.0 |
1000 |
52 |
f |
80.5 |
15.0 |
0.6 |
3.9 |
0.0 |
4000 |
52 |
f |
86.4 |
9.7 |
0.5 |
3.4 |
0.0 |
0 |
78 |
f |
71.8 |
23.7 |
1.2 |
3.3 |
0.0 |
50 |
78 |
f |
75.7 |
16.2 |
1.7 |
6.4 |
0.1 |
200 |
78 |
f |
65.9 |
26.9 |
1.0 |
6.3 |
0.0 |
1000 |
78 |
f |
73.5 |
19.8 |
1.2 |
5.6 |
0.0 |
4000 |
78 |
f |
78.4 |
16.0 |
1.1 |
4.5 |
0.0 |
0 |
103/104 |
f |
70.4 |
27.4 |
1.2 |
1.1 |
0.0 |
50 |
103/104 |
f |
69.3 |
28.8 |
0.8 |
1.1 |
0.0 |
200 |
103/104 |
f |
60.4 |
37.3 |
1.3 |
1.0 |
0.0 |
1000 |
103/104 |
f |
75.8 |
22.0 |
1.0 |
1.2 |
0.0 |
4000 |
103/104 |
f |
73.1 |
23.8 |
1.6 |
1.6 |
0.0 |
+ difference against control for p<0.05
++ difference against control for p<0.01
Table 8: Clinical Chemistry
Dose |
Week |
Sex |
ASAT (GOT) |
ALAT (GPT) |
APh |
GGT |
GLDH |
ppm |
|
|
U/L |
U/L |
U/L |
U/L |
U/L |
0 |
26/27 |
m |
41.3 |
38.9 |
190 |
1 |
5.7 |
50 |
26/27 |
m |
38.8 |
40.1 |
186 |
1 |
5.4 |
200 |
26/27 |
m |
44.4 |
40.3 |
171 |
2 |
7.9 |
1000 |
26/27 |
m |
38.5 |
40.4 |
157+ |
2 |
5.0 |
3000 |
26/27 |
m |
35.1 |
37.7 |
150+ |
3++ |
3.8 |
0 |
52 |
m |
39.9 |
42.2 |
184 |
1 |
16.2 |
50 |
52 |
m |
40.5 |
44.l |
193 |
1 |
22.8 |
200 |
52 |
m |
36.0 |
37.9 |
162 |
2 |
12.0 |
1000 |
52 |
m |
31.2 |
37.4 |
154+ |
1 |
12.6 |
3000 |
52 |
m |
34.6 |
41.0 |
156+ |
2 |
12.8 |
0 |
78 |
m |
38.8 |
36.5 |
157 |
1 |
16.0 |
50 |
78 |
m |
42.5 |
43.8 |
178 |
2 |
22.0 |
200 |
78 |
m |
35.1 |
32.1 |
142 |
1 |
14.0 |
1000 |
78 |
m |
33.2 |
36.0 |
141 |
1 |
17.9 |
3000 |
78 |
m |
32.7 |
35.7 |
144 |
2 |
20.0 |
0 |
103/104 |
m |
40.2 |
37.5 |
164 |
3 |
21.4 |
50 |
103/104 |
m |
42.2 |
33.1 |
175 |
2 |
21.6 |
200 |
103/104 |
m |
37.4 |
32.3 |
134 |
2 |
10.8 |
1000 |
103/104 |
m |
37.4 |
29.8 |
137 |
3 |
14.8 |
3000 |
103/104 |
m |
36.9 |
35.3 |
140 |
3 |
19.0 |
0 |
26/27 |
f |
40.7 |
43.4 |
121 |
0 |
11.5 |
50 |
26/27 |
f |
48.0 |
43.6 |
133 |
0 |
18.7 |
200 |
26/27 |
f |
41.0 |
44.9 |
122 |
0 |
11.3 |
1000 |
26/27 |
f |
32.3+ |
41.9 |
103 |
0 |
6.8 |
4000 |
26/27 |
f |
30.9++ |
42.6 |
105 |
1 |
1.0 |
0 |
52 |
f |
44.7 |
42.6 |
120 |
0 |
17.2 |
50 |
52 |
f |
46.8 |
44.1 |
134 |
0 |
22.9 |
200 |
52 |
f |
42.5 |
43.8 |
111 |
0 |
20.4 |
1000 |
52 |
f |
55.4 |
55.2 |
99 |
0 |
63.3 |
4000 |
52 |
f |
33.3 |
45.0 |
118 |
0 |
9.2 |
0 |
78 |
f |
59.4 |
45.7 |
100 |
1 |
59.2 |
50 |
78 |
f |
89.4 |
62.2 |
113 |
0 |
104.8 |
200 |
78 |
f |
73.9 |
54.4 |
91 |
0 |
87.7 |
1000 |
78 |
f |
40.3 |
36.3 |
81+ |
1 |
29.1 |
4000 |
78 |
f |
41.2 |
37.5 |
90 |
2+ |
18.4 |
0 |
103/104 |
f |
61.3 |
45.9 |
114 |
0 |
35.5 |
50 |
103/104 |
f |
61.3 |
49.0 |
114 |
1 |
53.7 |
200 |
103/104 |
f |
76.6 |
53.8 |
102 |
0 |
70.9 |
1000 |
103/104 |
f |
45.7 |
41.2 |
92 |
0 |
26.4 |
4000 |
103/104 |
f |
43.4 |
43.6 |
102 |
2++ |
41.5 |
+ difference against control for p<0.05
++ difference against control for p<0.01
Table 9: Activities and inhibition of cholinesterases (CHE)
Dose |
Week |
Sex |
CHE/Brain |
CHE/Plasma |
CHE/Erythrocytes |
|||
ppm |
|
|
U/g |
% Inhibition |
kU/L |
% Inhibition |
kU/L |
% Inhibition |
0 |
27 |
m |
- |
|
0.50 |
|
1.06 |
|
50 |
27 |
m |
- |
|
0.52 |
|
1.06 |
|
200 |
27 |
m |
- |
|
0.52 |
|
0.88 |
17 |
1000 |
27 |
m |
- |
|
0.50 |
|
0.65+ |
39 |
3000 |
27 |
m |
- |
|
0.45 |
10 |
0.32++ |
70 |
0 |
52/53 |
m |
10.64 |
|
0.62 |
|
1.14 |
|
50 |
52/53 |
m |
10.46 |
2 |
0.68 |
|
1.15 |
|
200 |
52/53 |
m |
10.56 |
|
0.63 |
|
1.01 |
11 |
1000 |
52/53 |
m |
10.26 |
4 |
0.61 |
2 |
1.00 |
12 |
3000 |
52/53 |
m |
9.54 |
10 |
0.56 |
10 |
0.57++ |
50 |
0 |
78 |
m |
- |
|
0.72 |
|
1.24 |
|
50 |
78 |
m |
- |
|
0.71 |
|
1.35 |
|
200 |
78 |
m |
- |
|
0.71 |
1 |
1.16 |
6 |
1000 |
78 |
m |
- |
|
0.63 |
13 |
0.84+ |
32 |
3000 |
78 |
m |
|
|
0.55 |
24 |
0.55++ |
55 |
0 |
103/104/105 |
m |
9.88 |
|
0.66 |
|
0.83 |
|
50 |
103/104/105 |
m |
9.87 |
|
0.95 |
|
0.76 |
8 |
200 |
103/104/105 |
m |
9.86 |
|
0.86 |
|
0.70 |
16 |
1000 |
103/104/105 |
m |
9.47 |
4 |
0.65 |
2 |
0.50++ |
40 |
3000 |
103/104/105 |
m |
8.50 |
14 |
0.55 |
17 |
0.15++ |
82 |
0 |
26/27 |
f |
- |
|
2.47 |
|
0.77 |
|
50 |
26/27 |
f |
- |
|
2.24 |
9 |
0.83 |
|
200 |
26/27 |
f |
- |
|
2.15 |
13 |
0.70 |
9 |
1000 |
26/27 |
f |
- |
|
2.57 |
|
0.38++ |
51 |
4000 |
26/27 |
f |
- |
|
2.30 |
7 |
0.08++ |
90 |
0 |
52/53 |
f |
11.28 |
|
2.39 |
|
1.24 |
|
50 |
52/53 |
f |
10.82+ |
4 |
2.21 |
8 |
1.17 |
6 |
200 |
52/53 |
f |
10.80 |
4 |
2.39 |
|
0.96+ |
23 |
1000 |
52/53 |
f |
9.41++ |
17 |
2.73 |
|
0.58++ |
53 |
4000 |
52/53 |
f |
7.07++ |
37 |
2.49 |
|
0.34++ |
73 |
0 |
78 |
f |
- |
|
2.18 |
|
1.19 |
|
50 |
78 |
f |
- |
|
2.03 |
7 |
1.18 |
1 |
200 |
78 |
f |
- |
|
2.12 |
3 |
1.01 |
15 |
1000 |
78 |
f |
- |
|
2.45 |
|
0.45++ |
62 |
4000 |
78 |
f |
- |
|
2.22 |
|
0.13++ |
89 |
0 |
103/104/105 |
f |
10.22 |
|
2.25 |
|
0.74 |
|
50 |
103/104/105 |
f |
10.02 |
2 |
2.31 |
|
0.66 |
11 |
200 |
103/104/105 |
f |
9.86 |
4 |
1.95 |
13 |
0.73 |
1 |
1000 |
103/104/105 |
f |
8.91++ |
13 |
2.11 |
6 |
0.28++ |
62 |
4000 |
103/104/105 |
f |
6.02++ |
41 |
2.08 |
8 |
0.08++ |
89 |
+ difference against control for p<0.05
++ difference against control for p<0.01
Table 10: Determination of urine pH week 98/99
|
pH mean value |
|
Sex |
male |
female |
Dose (ppm) |
||
0 |
7.55 |
7.20 |
50 |
7.57 |
7.47 |
200 |
7.32 |
7.12 |
1000 |
7.45 |
7.64 |
3000 |
7.34 |
- |
4000 |
- |
7.67 |
Table 11: Urinalyses
Dose (ppm) |
Week |
Sex |
VOL (ml) |
Density (g/L) |
PROT*Vol (mg) |
PROT (g/L) |
0 |
26 |
m |
13.9 |
1023 |
20.3 |
1.78 |
50 |
26 |
m |
14.8 |
1021 |
30.1 |
2.88 |
200 |
26 |
m |
12.2 |
1025 |
56.5 |
5.23 |
1000 |
26 |
m |
10.8 |
1032 |
26.7 |
3.29 |
3000 |
26 |
m |
15.6 |
1020 |
31.1 |
3.07 |
0 |
52 |
m |
6.3 |
1036 |
33.5 |
5.73 |
50 |
52 |
m |
8.2 |
1035 |
57.9 |
7.95 |
200 |
52 |
m |
7.0 |
1039 |
101.2 |
13.71 |
1000 |
52 |
m |
11.2 |
1032 |
61.4 |
9.03 |
3000 |
52 |
m |
9.3 |
1034 |
58.2 |
7.84 |
0 |
79 |
m |
7.1 |
1037 |
35.2 |
5.35 |
50 |
79 |
m |
7.8 |
1030 |
42.9 |
5.49 |
200 |
79 |
m |
12.3 |
1023 |
76.1 |
6.53 |
1000 |
79 |
m |
7.5 |
1025 |
36.4 |
5.99 |
3000 |
79 |
m |
8.4 |
1028 |
43.5 |
6.37 |
0 |
103 |
m |
11.6 |
1024 |
75.9 |
7.70 |
50 |
103 |
m |
8.6 |
1031 |
77.2 |
8.98 |
200 |
103 |
m |
10.9 |
1024 |
84.1 |
8.65 |
1000 |
103 |
m |
8.8 |
1030 |
91.4 |
9.67 |
3000 |
103 |
m |
11.8 |
1024 |
75.7 |
6.64 |
0 |
26 |
f |
8.5 |
1026 |
5.5 |
0.68 |
50 |
26 |
f |
14.0 |
1016 |
2.8 |
0.25 |
200 |
26 |
f |
12.5 |
1022 |
2.4 |
0.24+ |
1000 |
26 |
f |
6.9 |
1028 |
1.8 |
0.31 |
4000 |
26 |
f |
7.3 |
1026 |
1.9 |
0.34 |
0 |
52 |
f |
8.4 |
1029 |
13.3 |
1.25 |
50 |
52 |
f |
9.0 |
1021 |
3.2 |
0.37 |
200 |
52 |
f |
10.7 |
1022 |
4.5 |
0.45 |
1000 |
52 |
f |
12.5 |
1015+ |
2.5 |
0.32+ |
4000 |
52 |
f |
8.4 |
1030 |
1.8 |
0.29+ |
0 |
79 |
f |
5.8 |
1026 |
10.1 |
1.85 |
50 |
79 |
f |
8.0 |
1020+ |
6.3 |
0.91 |
200 |
79 |
f |
11.2+ |
1019+ |
10.9 |
1.25 |
1000 |
79 |
f |
10.8+ |
1015+ |
6.0 |
0.66 |
4000 |
79 |
f |
10.6 |
1018+ |
3.3 |
0.42 |
0 |
103 |
f |
8.4 |
1024 |
46.0 |
4.94 |
50 |
103 |
f |
10.0 |
1021 |
42.5 |
4.80 |
200 |
103 |
f |
7.9 |
1024 |
23.6 |
2.85 |
1000 |
103 |
f |
9.8 |
1022 |
19.7 |
1.16+ |
4000 |
103 |
f |
7.9 |
1029 |
7.5 |
1.14+ |
+ difference against control for p<0.05
++ difference against control for p<0.01
Table 12: Absolute and relative organ weights (mg/100 g bw) – interim kill
Dose |
Sex |
Body weight |
Brain |
Adrenals |
Heart |
Liver |
Spleen |
Kidneys |
Testes |
Ovaries |
Absolute organ weights |
g |
mg |
mg |
mg |
mg |
mg |
mg |
mg |
mg |
|
0 |
m |
517 |
2052 |
54 |
1402 |
17938 |
853 |
2910 |
3844 |
- |
50 |
m |
557 |
2159+ |
53 |
1534 |
19013 |
944 |
3220 |
4062 |
- |
200 |
m |
528 |
2064 |
54 |
1467 |
17702 |
881 |
3085 |
3815 |
- |
1000 |
m |
531 |
2128 |
52 |
1560 |
19351 |
801 |
3054 |
3652 |
- |
3000 |
m |
519 |
2105 |
56 |
1439 |
21255+ |
715 |
2998 |
4051 |
- |
0 |
f |
312 |
1912 |
84 |
1023 |
10016 |
535 |
1994 |
- |
172 |
50 |
f |
302 |
1890 |
83 |
1031 |
9690 |
561 |
2057 |
- |
173 |
200 |
f |
296 |
1920 |
83 |
1059 |
9442 |
476 |
2061 |
- |
197 |
1000 |
f |
285 |
1879 |
80 |
988 |
10299 |
684 |
1983 |
- |
204 |
4000 |
f |
261++ |
1827 |
69 |
974 |
11901++ |
488 |
1856 |
- |
162 |
Relative organ weights |
Body weight |
Brain |
Adrenals |
Heart |
Liver |
Spleen |
Kidneys |
Testes |
Ovaries |
|
g |
mg |
mg |
mg |
mg |
mg |
mg |
mg |
mg |
||
0 |
m |
517 |
400 |
10 |
272 |
3480 |
166 |
561 |
749 |
- |
50 |
m |
557 |
390 |
10 |
276 |
3424 |
170 |
580 |
731 |
- |
200 |
m |
528 |
393 |
10 |
279 |
3347 |
167 |
585 |
723 |
- |
1000 |
m |
531 |
403 |
10 |
296 |
3649 |
151 |
579 |
697 |
- |
3000 |
m |
519 |
406 |
11 |
278 |
4094+ |
137 |
578 |
782 |
- |
0 |
f |
312 |
620 |
27 |
331 |
3235 |
173 |
642 |
- |
55 |
50 |
f |
302 |
628 |
28 |
342 |
3205 |
186 |
681 |
- |
57 |
200 |
f |
296 |
652 |
28 |
357 |
3196 |
195 |
696 |
- |
68 |
1000 |
f |
285 |
660 |
28 |
347 |
3615+ |
235 |
696 |
- |
71 |
4000 |
f |
261++ |
704++ |
27 |
375+ |
4584++ |
186 |
711+ |
- |
63 |
+ difference against control for p<0.05
++ difference against control for p<0.01
Table 13: Absolute and relative organ weights (mg/100 g bw) – terminal kill
Dose |
Sex |
Body weight |
Brain |
Adrenals |
Heart |
Liver |
Spleen |
Kidneys |
Testes |
Ovaries |
Absolute organ weights |
g |
mg |
mg |
mg |
mg |
mg |
mg |
mg |
mg |
|
0 |
m |
534 |
2176 |
64 |
1866 |
18013 |
1217 |
3491 |
3785 |
- |
50 |
m |
507 |
2191 |
68 |
1960 |
17363 |
1098 |
3429 |
3929 |
- |
200 |
m |
508 |
2172 |
58 |
1830 |
18083 |
1157 |
3611 |
3847 |
- |
1000 |
m |
513 |
2165 |
71 |
18368 |
18368 |
1035+ |
3407 |
4048 |
- |
3000 |
m |
483++ |
2118 |
78 |
19494 |
19494 |
937++ |
3369 |
4029 |
- |
0 |
f |
318 |
1931 |
84 |
11395 |
11395 |
721 |
2295 |
- |
216 |
50 |
f |
318 |
1957 |
77 |
11784 |
11784 |
714 |
2380 |
- |
173 |
200 |
f |
312 |
1947 |
82 |
11347 |
11347 |
738 |
2328 |
- |
181 |
1000 |
f |
308 |
1938 |
69 |
11579 |
11579 |
671 |
2197 |
- |
193 |
4000 |
f |
282++ |
1914 |
71 |
13476++ |
13476++ |
585++ |
2190 |
- |
194 |
Relative organ weights |
Body weight |
Brain |
Adrenals |
Heart |
Liver |
Spleen |
Kidneys |
Testes |
Ovaries |
|
g |
mg |
mg |
mg |
mg |
mg |
mg |
mg |
mg |
||
0 |
m |
534 |
410 |
12 |
351 |
3369 |
229 |
655 |
712 |
- |
50 |
m |
507 |
435 |
14 |
387 |
3416 |
229 |
680 |
776 |
- |
200 |
m |
508 |
434 |
12 |
367 |
3608 |
230 |
721 |
756 |
- |
1000 |
m |
513 |
426 |
14 |
364 |
3608 |
203 |
669 |
793 |
- |
3000 |
m |
483++ |
483++ |
17 |
400+ |
4050++ |
196+ |
706 |
835++ |
- |
0 |
f |
318 |
617 |
27 |
448 |
3608 |
229 |
728 |
- |
68 |
50 |
f |
318 |
625 |
25 |
444 |
3758 |
229 |
754 |
- |
54 |
200 |
f |
312 |
633 |
27 |
440 |
3674 |
239 |
754 |
- |
58 |
1000 |
f |
308 |
638 |
23 |
437 |
3773 |
219 |
716 |
- |
62 |
4000 |
f |
282++ |
682++ |
25 |
470 |
4795++ |
208 |
781 |
- |
69 |
+ difference against control for p<0.05
++ difference against control for p<0.01
Table 14: Number of rats with remarkable non-neoplastic lesions – interim kill
Sex |
|
m |
m |
m |
m |
|
f |
f |
f |
f |
f |
Dose |
(ppm) |
0 |
50 |
200 |
1000 |
3000 |
0 |
50 |
200 |
1000 |
4000 |
Organ/Finding |
Number Animals |
10 |
10 |
10 |
10 |
10 |
10 |
10 |
10 |
10 |
10 |
Adrenal Glands - increased vacuolation in zona fascicu.* |
§4 |
6 |
4 |
6 |
10 |
§2 |
0 |
0 |
0 |
9 |
|
Liver - hepatocellular hypertrophy (centilobul.) |
§0 |
0 |
0 |
6 |
10 |
§0 |
0 |
0 |
3 |
10 |
|
Urinary Bladder - transitional hyperplasia |
#0 |
0 |
2 |
2 |
3 |
§0 |
0 |
0 |
0 |
3 |
|
*= increased cytoplasmic vacuolisation in zona fasciculata
# trend (p < 0.05)
§ trend (p < 0.01)
Table 15:Number of rats with remarkable non-neoplastic lesions – terminal kill
Sex |
|
|
m |
m |
m |
m |
|
|
f |
f |
f |
f |
f |
Dose |
(ppm) |
|
0 |
50 |
200 |
1000 |
3000 |
|
0 |
50 |
200 |
1000 |
4000 |
Organ/Finding |
Number Animals |
|
50 |
50 |
50 |
50 |
50 |
|
50 |
50 |
50 |
50 |
50 |
Liver - eosinophilic foci |
# |
3 |
3 |
1 |
2 |
10 |
# |
|
|
3 |
3 |
3 |
|
- focal degenerative changes (periportal) |
§ |
|
|
|
2 |
13 |
|
|
|
|
|
1 |
|
- clear cell foci |
|
36 |
34 |
31 |
35 |
28 |
|
6 |
9 |
6 |
9 |
1 |
|
- cytoplasmic changes (periportal) |
§ |
|
|
|
2 |
22 |
|
|
|
|
|
|
|
- cytoplasmic changes (centrilobular) |
§ |
|
|
1 |
|
14 |
§ |
1 |
3 |
3 |
8 |
38 |
|
- hepatocellular hypertrophy (centilobul.) |
|
1 |
3 |
|
1 |
4 |
# |
|
|
1 |
3 |
40 |
|
- peliotic foci |
|
|
|
|
|
|
§ |
|
|
|
|
3 |
|
- single cell necrosis |
|
|
|
|
|
|
§ |
4 |
7 |
5 |
5 |
16 |
|
Urinary Bladder - transitional hyperplasia (focal) |
§ |
|
|
|
|
4 |
|
1 |
|
|
|
3 |
|
- simple transitional hyperplasia (diffuse) |
§ |
|
|
3 |
|
11 |
§ |
|
|
|
|
20 |
|
Skeletal Muscle - atrophy |
# |
7 |
2 |
9 |
10 |
11 |
§ |
|
|
1 |
2 |
9 |
|
- fibre degeneration (thigh) |
|
4 |
5 |
6 |
3 |
6 |
# |
2 |
|
1 |
|
7 |
|
Sciatic Nerve - degenerative myelinopathy |
§ |
21 |
25 |
27 |
32 |
43 |
§ |
17 |
20 |
13 |
23 |
38 |
|
Thyroid Gland - follicular hyperplasia |
§ |
1 |
1 |
2 |
3 |
7 |
|
2 |
|
|
2 |
1 |
|
- mineralization of follicular colloid |
# |
33 |
28 |
31 |
35 |
39 |
§ |
13 |
13 |
17 |
17 |
43 |
|
Adrenal Glands - increased vacuolation in zona fascicu.* |
§ |
5 |
7 |
7 |
3 |
20 |
|
1 |
2 |
2 |
3 |
4 |
|
- degenerative changes in the cortex |
|
5 |
3 |
3 |
3 |
3 |
|
19 |
14 |
11 |
8 |
4 |
|
Pituitary Gland (pars distalis) - hyperplasia (focal) |
|
10 |
6 |
4 |
13 |
10 |
|
18 |
13 |
9 |
11 |
9 |
|
Stomach (glandular mucosa) - dilated glands |
|
6 |
4 |
8 |
7 |
6 |
# |
7 |
7 |
6 |
8 |
16 |
|
Tongue - necrotizing arteritis |
|
7 |
12 |
7 |
11 |
9 |
|
7 |
7 |
4 |
2 |
0 |
|
Heart - cardiomyopathy |
|
43 |
45 |
48 |
44 |
48 |
|
37 |
37 |
33 |
28 |
22 |
|
Kidneys - chronic progressive nephropathy |
|
47 |
47 |
48 |
44 |
48 |
|
41 |
32 |
36 |
26 |
21 |
|
- diffuse urothelial hyperplasia |
|
10 |
18 |
16 |
8 |
15 |
|
35 |
35 |
30 |
21 |
19 |
|
Spleen - pigment storage |
|
19 |
24 |
31 |
20 |
27 |
§ |
9 |
9 |
11 |
15 |
23 |
|
# trend (p < 0.05)
§ trend (p < 0.01)
Table 16: Tumor incidences – Number with neoplastic lesions – terminal kill
Sex |
|
Males |
Females |
||||||||
Dose |
(ppm) |
0 |
50 |
200 |
1000 |
3000 |
0 |
50 |
200 |
1000 |
4000 |
Organ/Findings |
No.animals |
50 |
50 |
50 |
50 |
50 |
50 |
50 |
50 |
50 |
50 |
Liver |
No. exam. |
50 |
49 |
50 |
50 |
50 |
50 |
49 |
50 |
50 |
50 |
- Hepatocellular Adenoma |
|
1 |
|
1 |
|
|
|
1 |
|
|
1 |
- Hcpatocellular Adcnocarcinoma |
|
1 |
|
|
|
|
|
|
|
|
|
-Cholangioma |
|
|
|
|
|
|
|
|
|
1 |
|
- Cholangiocarcinoma |
|
|
1 |
|
|
|
|
|
|
|
|
Heart |
No. exam. |
50 |
49 |
50 |
50 |
50 |
50 |
49 |
50 |
50 |
50 |
- Endocardial Tumor (mal.) |
|
2 |
1 |
|
|
|
|
1 |
|
|
|
Kidneys |
No. exam. |
50 |
49 |
50 |
50 |
50 |
50 |
49 |
50 |
50 |
50 |
-Adenoma |
|
|
|
|
1 |
|
|
|
|
|
|
- Lipoma |
|
|
|
|
|
1 |
|
|
|
|
|
- Liposarcoma |
|
|
|
1 |
|
|
|
|
|
|
|
Urinary Bladder |
No. exam. |
50 |
48 |
50 |
50 |
50 |
50 |
49 |
48 |
49 |
49 |
- Transitional Cell Papilloma |
|
|
|
|
|
|
§ |
|
|
|
2 |
- Transitional Cell Carcinoma |
|
|
|
|
|
|
|
|
|
|
1 |
Urethra (residual)&) |
No. exam. |
|
|
|
|
1 |
|
|
|
|
|
- Transitional Cell Carcinoma |
|
|
|
|
|
1 |
|
|
|
|
|
LungsNo. Exam. |
No. exam. |
50 |
49 |
50 |
50 |
50 |
50 |
49 |
50 |
50 |
50 |
- AlvcolarffironchiolarAdenoma
|
|
1 |
|
|
|
|
|
|
|
|
|
- Alveolar/UronchiolarCarcinoma
|
|
|
1 |
1 |
|
|
|
|
|
|
|
Metastasis primary siteunknown |
|
|
1 |
|
|
|
|
|
1 |
|
|
Stomach No. Exam. |
No. exam. |
50 |
49 |
50 |
50 |
50 |
50 |
49 |
50 |
50 |
50 |
- Forestomach Papilloma |
|
|
|
|
|
|
|
|
|
|
1 |
Colon |
No. exam. |
50 |
48 |
50 |
50 |
50 |
50 |
49 |
49 |
49 |
49 |
- Fibrosarcoma |
|
|
|
|
|
|
|
1 |
|
|
|
Intestine (residual)&) |
No. exam. |
|
|
|
|
|
|
|
1 |
2 |
2 |
-Fibroma |
|
|
|
|
|
|
|
|
|
1 |
|
Pancreas |
No. exam. |
48 |
49 |
50 |
50 |
50 |
50 |
49 |
50 |
50 |
50 |
- Islet Cell Adenoma |
|
1 |
1 |
2 |
3 |
|
1 |
|
|
|
|
- Acinar Cell Adenoma |
|
|
|
|
|
1 |
|
|
|
1 |
|
Thymus |
No. exam. |
49 |
49 |
50 |
50 |
50 |
48 |
49 |
50 |
50 |
48 |
- Thymoma (benigne) |
|
1 |
1 |
|
1 |
|
|
I |
2 |
|
1 |
- Thymoma (maligne) |
|
1 |
|
|
|
|
|
|
|
|
|
Mesent. Lymph Node Mesent. Lymph NodeNo. Exam. |
No. exam. |
49 449 |
49 5049 |
50 50 |
50 50 |
50 50 |
50 50 |
49 49 |
50 50 |
49 49 |
50 50 |
- Hemangioma |
|
1 |
1 |
1 |
|
1 |
|
|
|
|
|
Spleen |
No. exam. |
50 |
49 |
50 |
50 |
50 |
50 |
49 |
50 |
49 |
50 |
- Hemangiosarcoma |
|
1 |
|
|
|
|
|
|
|
|
|
Skin/Other&) |
No. exam. |
9 |
3 |
5 |
5 |
4 |
6 |
2 |
2 |
2 |
2 |
- Schwannoma (mal.) |
|
|
1 |
|
|
|
|
|
|
|
|
- Fibroma |
|
|
|
|
1 |
|
|
|
|
|
|
- Fibrosarcoma |
|
1 |
|
|
|
|
|
1 |
|
|
|
- Squamous Cell Carcinoma |
|
2 |
1 |
|
1 |
|
|
|
|
|
|
- Keratoacanthoma |
|
3 |
|
2 |
|
1 |
|
|
|
|
|
- Papilloma |
|
|
|
|
|
1 |
|
|
|
|
|
- Hair Follicle Tumor (ben.) |
|
|
|
1 |
|
|
|
|
|
|
|
-Lipoma |
|
|
|
|
|
|
1 |
|
|
|
|
- Basal Cell Carcinoma |
|
|
|
|
|
|
|
|
1 |
|
|
Mammary Gland/Region |
No. exam. |
- |
- |
- |
- |
- |
50 |
49 |
50 |
50 |
50 |
-Adenoma |
|
|
|
|
|
|
|
1 |
|
|
|
- Adenocarcinoma |
|
|
|
|
|
|
3 |
1 |
6 |
|
2 |
- Adenocarcinoma arising in a fibroadenoma |
|
|
|
|
|
|
1 |
|
|
|
1 |
- Fihroadenoma |
|
|
|
|
|
|
15 |
10 |
7 |
6 |
5 |
- Tumor Mixed (mal.) |
|
|
|
|
|
|
1 |
|
|
1 |
1 |
Brain |
No. exam. |
50 |
49 |
50 |
50 |
50 |
50 |
49 |
50 |
50 |
50 |
- Astrocytoma (ben.) |
|
1 |
|
|
|
|
|
|
|
|
|
- Granular Cell Tumor (ben.) |
|
|
|
|
|
1 |
|
1 |
|
|
|
Testes |
No. exam. |
50 |
49 |
50 |
50 |
50 |
- |
- |
- |
- |
- |
- Leydig Cell Adenoma |
|
1 |
4 |
|
1 |
2 |
|
|
|
|
|
Prostate |
No. exam. |
50 |
49 |
50 |
50 |
50 |
- |
- |
- |
- |
- |
- Fibrosarcoma |
|
|
|
|
1 |
|
|
|
|
|
|
Ovaries |
No. exam. |
- |
- |
- |
- |
- |
50 |
49 |
50 |
50 |
50 |
- Granulosa Cell Tumor (ben.) |
|
|
|
|
|
|
1 |
|
|
|
|
- Granulosa Cell Tumor (mal.) |
|
|
|
|
|
|
|
|
|
|
1 |
- Tumor/Sex Cord (ben.) |
|
|
|
|
|
|
|
1 |
1 |
1 |
|
Uterus Uterus |
No. exam. No. Exam. |
|
|
|
|
|
50 |
49 550 |
50 49 |
50 50 |
50 50 |
-Adenoma |
|
|
|
|
|
|
|
|
|
1 |
|
- Adenocarcinoma |
|
|
|
|
|
|
§ 2 |
2 |
3 |
3 |
8 |
- Adenosquamous Carcinoma |
|
|
|
|
|
|
1 |
1 |
1 |
|
|
- Squamous Cell Carcinoma |
|
|
|
|
|
|
|
|
|
|
1 |
- Schwannoma (mal.) |
|
|
|
|
|
|
1 |
|
1 |
|
|
- Stromal Sarcoma |
|
|
|
|
|
|
|
1 |
|
1 |
1 |
- Stromal Polyp |
|
|
|
|
|
|
7 |
7 |
4 |
5 |
6 |
- Glandular Polyp |
|
|
|
|
|
|
1 |
|
3 |
|
|
Vagina |
No. exam. |
- |
- |
- |
- |
- |
50 |
49 |
50 |
50 |
50 |
- Granular Cell Tumor (ben.) |
|
|
|
|
|
|
|
|
|
|
1 |
Adrenal Glands |
No. exam. |
50 |
49 |
50 |
50 |
50 |
50 |
49 |
50 |
50 |
50 |
- Cortical Adenoma |
|
|
|
1 |
|
1 |
|
1 |
|
|
|
- Cortical Adenocarcinoma |
|
|
|
|
|
|
1 |
|
|
|
|
- Medullary Tumor (ben.) |
|
10 |
9 |
5 |
8 |
10 |
1 |
3 |
1 |
1 |
|
- Medullary Tumor (mal.) |
|
2 |
1 |
1 |
1 |
|
|
|
1 |
|
|
Pituitary Gland |
No. exam. |
49 |
49 |
50 |
50 |
50 |
50 |
49 |
50 |
50 |
50 |
- Adenoma (Pars Distalis) |
|
5 |
6 |
11 |
5 |
8 |
24 |
18 |
24 |
14 |
11 |
- Adenoma (Pars Intermedia) |
|
|
|
|
|
|
1 |
|
|
|
1 |
- Adenocarcinoma (Pars Distalis) |
|
|
|
|
|
|
1 |
|
1 |
|
|
Thyroid Gland |
No. exam. |
50 |
49 |
50 |
50 |
50 |
50 |
49 |
50 |
50 |
50 |
- C-Cell Adenoma |
|
3 |
7 |
2 |
2 |
3 |
3 |
2 |
3 |
4 |
3 |
- C-Cell Adenocarcinoma |
|
1 |
|
|
|
|
|
|
|
|
|
- Follicular Adenoma |
|
# |
|
|
1 |
2 |
1 |
1 |
1 |
1 |
1 |
- Follicular Adenocarcinoma |
|
|
|
|
|
1 |
|
|
|
|
|
Bone/NOS&) |
|
|
|
|
|
|
|
|
1 |
1 |
|
- Osteosarcoma |
|
|
|
|
|
|
|
|
1 |
|
|
- Schwannoma (maligne) |
|
|
|
|
|
|
|
|
|
1 |
|
Systemic Tumors |
No. exam. |
50 |
49 |
50 |
50 |
50 |
50 |
49 |
50 |
50 |
50 |
- Lymphoma (maligne) |
|
2 |
1 |
|
2 |
|
|
|
1 |
1 |
|
- Myeloid Leukemia |
|
|
1 |
|
1 |
|
|
|
|
|
|
- Histiocytic Sarcoma |
|
2 |
1 |
|
1 |
1 |
|
|
|
|
|
* in case of two tumors in a paired organ one tumor was taken into consideration
&) only investigated in case of macro lesions
# positive trend p<0.05
§ positive trend p<0.01
NOS = not otherwise specified
no. = number
exam. = examinations
Table 17: Number of Animals with neoplastic lesions – terminal kill
Sex |
|
male |
female |
||||||||
Dose ppm |
|
0 |
50 |
200 |
1000 |
3000 |
0 |
50 |
200 |
1000 |
4000 |
Organ/Finding |
No. animals |
50 |
50 |
50 |
50 |
50 |
50 |
50 |
50 |
50 |
50 |
Body cavities &) |
No. exam. |
5 |
3 |
6 |
6 |
3 |
3 |
3 |
3 |
4 |
6 |
- Mesothelioma (benign) |
|
1 |
|
|
|
|
|
|
|
|
|
- Lipoma |
|
|
|
1 |
|
|
1 |
|
|
1 |
|
- Fibroma |
|
|
|
|
|
|
1 |
|
|
|
|
Clitorial glands &) |
No. exam. |
|
|
|
|
|
|
1 |
1 |
|
|
- Squamous cell carcinoma |
|
|
|
|
|
|
|
|
1 |
|
|
Number of benign tumors |
|
29 |
29 |
27 |
23 |
32 |
58 |
47 |
47 |
36 |
33 |
Malignant tumors |
|
15 |
10 |
3 |
7 |
3 |
11 |
8 |
18 |
7 |
16 |
Total tumors |
|
44 |
39 |
30 |
30 |
35 |
69 |
55 |
65 |
43 |
49 |
* in case of two tumors in a paired organ one tumor was taken into consideration
&) only investigated in case of macro lesions
# positive trend p<0.05
§ positive trend p<0.01
NOS = not otherwise specified
no. = number
exam. = examinations
Table 18: Animals with tumors – terminal kill
Sex |
Males |
Females |
||||||||
Dose ppm |
0 |
50 |
200 |
1000 |
3000 |
0 |
50 |
200 |
1000 |
4000 |
|
lntercurrent Deaths |
|||||||||
No. of animals examined |
21 |
19 |
21 |
17 |
13 |
12 |
20 |
24 |
20 |
22 |
Animals with tumors |
12 |
11 |
13 |
7 |
5 |
10 |
13 |
19 |
12 |
15 |
Animals with benign tumors only |
5 |
7 |
10 |
3 |
4 |
8 |
7 |
9 |
7 |
5 |
Animals with malignant tumors only |
5 |
5 |
3 |
2 |
|
1 |
4 |
5 |
2 |
3 |
Animals with benign and malignant tumors |
2 |
|
|
2 |
1 |
1 |
2 |
5 |
3 |
7 |
|
Terminal Kill |
|||||||||
No. of animals examined |
29 |
31 |
29 |
33 |
37 |
38 |
30 |
26 |
30 |
28 |
Animals with tumors |
17 |
16 |
13 |
16 |
21 |
31 |
20 |
22 |
16 |
15 |
Animals with benign tumors only |
10 |
11 |
13 |
13 |
19 |
25 |
19 |
15 |
14 |
10 |
Animals with malignant tumors only |
2 |
2 |
|
2 |
1 |
1 |
|
|
|
1 |
Animals with benign and malignant tumors |
5 |
3 |
|
1 |
1 |
5 |
1 |
7 |
2 |
4 |
|
All Animals |
|||||||||
No. of animals examined |
50 |
50 |
50 |
50 |
50 |
50 |
50 |
50 |
50 |
50 |
Animals with tumors |
29 |
27 |
26 |
23 |
26 |
41 |
33 |
41 |
28 |
30 |
Animals with benign tumors only |
15 |
18 |
23 |
16 |
23 |
33 |
26 |
24 |
21 |
15 |
Animals with malignant tumors only |
7 |
7 |
3 |
4 |
1 |
2 |
4 |
5 |
2 |
4 |
Animals with benign and malignant tumors |
7 |
3 |
|
3 |
2 |
6 |
3 |
12 |
5 |
11 |
Table 19: Occurrence of tumor bearing animals in time
Sex |
|
Males |
Females |
|||||||||
Dose ppm |
|
0 |
50 |
200 |
1000 |
3000 |
0 |
50 |
200 |
1000 |
4000 |
|
Number of Animals with Neoplasms |
|
|
|
|
|
|
|
|
|
|
|
|
Dying Prematurely |
|
12 |
11 |
13 |
7 |
5 |
10 |
13 |
19 |
12 |
15 |
|
Neoplasms |
benign |
11 |
9 |
13 |
7 |
7 |
13 |
11 |
17 |
11 |
14 |
|
|
maligne |
7 |
5 |
3 |
4 |
1 |
3 |
7 |
9 |
5 |
11 |
|
|
total |
18 |
14 |
16 |
11 |
8 |
16 |
18 |
26 |
16 |
25 |
|
Number of Animals with Tumors |
||||||||||||
Week |
0-13 |
|
|
1 |
|
|
|
|
|
|
|
|
|
13-26 |
|
|
|
|
|
|
|
|
|
|
|
|
27-39 |
|
|
|
|
|
|
|
|
|
|
|
|
40-52 |
|
|
|
|
1 |
|
|
1 |
1 |
|
|
|
53-65 |
|
1 |
2 |
|
|
|
|
|
|
2 |
1 |
|
66-78 |
|
1 |
|
1 |
1 |
|
1 |
2 |
4 |
1 |
2 |
|
79-91 |
|
4 |
2 |
6 |
2 |
2 |
3 |
3 |
8 |
4 |
7 |
|
92-Necropsy |
|
6 |
7 |
6 |
3 |
3 |
6 |
7 |
6 |
5 |
5 |
Data source
Reference
- Reference Type:
- study report
- Title:
- Unnamed
- Year:
- 1 997
- Report date:
- 1997
Materials and methods
Test guideline
- Qualifier:
- according to guideline
- Guideline:
- OECD Guideline 453 (Combined Chronic Toxicity / Carcinogenicity Studies)
- Version / remarks:
- adopted 12th May 1981
- Deviations:
- no
- GLP compliance:
- yes
- Limit test:
- no
Test material
- Reference substance name:
- 4-(2-chlorophenyl)-N-cyclohexyl-N-ethyl-5-oxo-4,5-dihydro-1H-1,2,3,4-tetrazole-1-carboxamide
- EC Number:
- 605-140-1
- Cas Number:
- 158237-07-1
- Molecular formula:
- C16H20ClN5O2
- IUPAC Name:
- 4-(2-chlorophenyl)-N-cyclohexyl-N-ethyl-5-oxo-4,5-dihydro-1H-1,2,3,4-tetrazole-1-carboxamide
Constituent 1
Test animals
- Species:
- rat
- Strain:
- Wistar
- Remarks:
- Hsd/WIN: WU
- Sex:
- male/female
- Details on test animals or test system and environmental conditions:
- TEST ANIMALS
- Source: Harlan Winkelmann
- Females nulliparous and non-pregnant: yes
- Age at study initiation: 5 - 6 weeks
- Weight at study initiation: 110 - 182 g (males), 102 - 158 g (females)
- Housing: 5 animals separated by sex in Type III Makrolon® and Ha cages. The cages containing the experimental animals were separated by groups and placed on shelves in order of ascending animal number. The position of the shelves was changed on a random basis every 4 weeks.
- Diet: Altromin® 1321 meal (Altromin GmbH, Lage) containing 1% peanut oil, ad libitum except during the urine collection period
- Water: tap water, ad libitum
- Acclimation period: 1 week
DETAILS OF FOOD AND WATER QUALITY:
The tap water complied with drinking water standards in accordance with the Deutsche Trinkwasserverordnung.
ENVIRONMENTAL CONDITIONS
- Temperature (°C): 22 ± 2°C
- Humidity (%): 55% ± 5%
- Air changes (per hr): 15 - 20
- Photoperiod (hrs dark / hrs light): 12/12
Administration / exposure
- Route of administration:
- oral: feed
- Vehicle:
- unchanged (no vehicle)
- Details on oral exposure:
- - DIET PREPARATION
- Rate of preparation of diet (frequency): weekly
- The test substance was blended (using a mixing granulator manufactured by Loedige, Paderborn) with Altromin® 1321 containing 1% peanut oil to minimize dust formation (including 0 ppm concentration). - Analytical verification of doses or concentrations:
- yes
- Details on analytical verification of doses or concentrations:
- Analytical investigations to demonstrate homogeneity and stability of the test compound in diet preparations were done prior to commencement of the study. The test substance content of diet mixtures fed to the animals was checked analytically at regular intervals throughout the study (start of study, randomly each 3 month period, end of study). Per dose one sample of the food mixtures was taken on the day the mixture was prepared, and another was taken after being kept underanimal room conditions for the feeding period (7 days). All these samples were kept deep frozen (at temperatures of approx. -20°C) until analysis.
- Duration of treatment / exposure:
- 24 months
- Frequency of treatment:
- daily per feed, ad libitum
Doses / concentrationsopen allclose all
- Dose / conc.:
- 50 ppm
- Remarks:
- corresponding to
2.5 mg/kg bw/day (males)
3.4 mg/kg bw/day (females)
- Dose / conc.:
- 200 ppm
- Remarks:
- corresponding to
10.3 mg/kg bw/day (males)
14.6 mg/kg bw/day (females)
- Dose / conc.:
- 1 000 ppm
- Remarks:
- corresponding to
52.7 mg/kg bw/day (males)
75.4 mg/kg bw/day (females)
- Dose / conc.:
- 3 000 ppm
- Remarks:
- corresponding to
170.4 mg/kg bw/day (males)
- Dose / conc.:
- 4 000 ppm
- Remarks:
- corresponding to
326.7 mg/kg bw/day (females)
- No. of animals per sex per dose:
- 60
- Control animals:
- yes, plain diet
- Details on study design:
- - Dose selection rationale: The dosages were selected on the basis of the results of a subchronic feeding study in Wistar rats in which the test substance had been administered to 10 males and 10 females at doses of 0, 20, 100, 400, 1600 and 6400 ppm for 13 weeks. In this study concentrations of up to 400 ppm were tolerated without adverse effects. At higher concentrations body weight depression, effects on blood parameters (essentially reduced hemoglobin and hematocrit values and elevated reticulocyte means), inhibition of the cholinesterase activity (erythrocytes) and signs of changed liver function such as altered biochemical parameters (enhanced cholesterol plasma levels, reduced contents of triglycerides and unesterified fatty acids, lower albumin means), enzyme induction, increased liver weights and occurrence of hepatocellular hypertrophy were evident. Rats receiving 6400 ppm exhibited diffuse hyperplasia of the bladder urothelium as well. In further electron-microscopical investigations on the urinary bladder cytotoxicity with mild regenerative hyperplasia was detected in 6400 ppm males (females were not investigated) with higher frequency than in controls
- Rationale for animal assignment: The rats were weighed individually beforehand and the required number of animals were then grouped by weight (light, midde and heavy subgroups) and randomly distributed into large containers. - Positive control:
- Not applicable
Examinations
- Observations and examinations performed and frequency:
- CAGE SIDE OBSERVATIONS: Yes
- Time schedule: twice daily and daily at weekends and on bank holidays
DETAILED CLINICAL OBSERVATIONS: Yes
- Time schedule: weekly
Parameters: orifices, posture, general behavior, respiration and excretory products were carefully examined, with any significant findings were registered. Animals that became ill or had developed neoplasms that could lead to death were kept apart, observed more frequently, and killed prematurely if death was imminent.
BODY WEIGHT: Yes
- Time schedule for examinations: weekly from start to Week 13, and every 2 weeks thereafter up to Week 105
FOOD CONSUMPTION AND COMPOUND INTAKE:
- Food consumption for each animal determined and mean daily diet consumption calculated as g food/kg body weight/day: Yes
- Compound intake calculated as time-weighted averages from the consumption and body weight gain data: Yes
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
WATER CONSUMPTION AND COMPOUND INTAKE: Yes
- Time schedule for examinations: weekly within the first 13 weeks of treatment and every 4 weeks thereafter up to Week 101
OPHTHALMOSCOPIC EXAMINATION: Yes
- Time schedule for examinations and dose groups that were examined: during the first two weeks of the study in all rats. In Week 54 and prior to the final bleeding all living animals scheduled for the final necropsy from the control group and the 3000 or 4000 ppm dose groups were examined. At study termination 200 and 1000 ppm males were investigated, additionally.
HAEMATOLOGY: Yes
- Time schedule for collection of blood: Weeks 26/27, 52, 78 and 105/106
- Anaesthetic used for blood collection: Yes, for glucose and deproteinized whole blood only
- Animals fasted: Yes
- How many animals: 10
- Parameters analysed: differential blood count, erythrocyt morphology, erythrocyte count, Heinz bodies, hemoglobin, hematocrit, leucocyte count, mean corpuscular hemoglobin, mean corpuscular hemoglobin concentration, mean corpuscular volume, platelet count, thromboplastin time, reticulocytes
CLINICAL CHEMISTRY: Yes
- Time schedule for collection of blood: Weeks 26/27, 52, 78 and 105/106
- Animals fasted: Yes
- How many animals: 10
- Parameters analysed: alkaline phosphatase, alanine aminotransferase, aspartate aminotransferase, gamma-glutamyl transferase, glutamate dehydrogenase, glucose, bilirubin, albumin, cholesterol, creatinine, total protein, triglyceride, urea, calcium, sodium, potassium, chloride and inorganic phosphate
URINALYSIS: Yes
- Time schedule for collection of urine: Weeks 26/27, 52, 79 and 98/99
- Metabolism cages used for collection of urine: Yes
- Animals fasted: Yes
- Parameters analysed: sediment, bilirubin, blood, glucose, ketone bodies, pH, protein, urobilinogen (semiquantitative), density, volume, total protein (quantitative)
NEUROBEHAVIOURAL EXAMINATION: No
IMMUNOLOGY: No
OTHER:
Cholinesterase activity in brain
- Time schedule: Weeks 52/53 and 104/105
Cholinesterase activities in plasma and erythrocytes
- Time schedule: Week 26/27, measurement were repeated for males in the following week due to some implausible values
Determination of crystals in urine sediment
- Time schedule: Week 103
- Dose groups that were examined: 8 animals, in controls and rats of the 3000 and 4000 ppm groups - Sacrifice and pathology:
- GROSS PATHOLOGY: Yes
- Organ weights: brain, heart, liver, spleen, kidneys, adrenals, testes, ovaries
HISTOPATHOLOGY: Yes, the following organs and tissues were fixed for analysis: Adrenals , Aorta , Brain (3 locations) [in some cases half brain] , Caecum, Colon, Duodenum, Epididymides , Esophagus, Eyes (with eyelids), Exorbital lacrimal glands, Femur (incl. bone marrow and knee joint), Harderian glands, Head-Nose-Pharynx area, Heart, lleum, Jejunum, Kidneys, Larynx, Liver, Lungs, Lymph nodes (mandibular and mesenteric), Mammary glands, Optic nerve, Ovaries (incl. oviduct), Pancreas, Pituitary, Prostate, Rectum, Residual Intestine, Salivary glands, Sciatic nerve, Seminal vesicles, Skeletal muscle, Skin, Spinal cord (3 regions), Spleen, Sternum (with bone marrow), Stomach (with forestomach), Tattooed ears, Testes, Thymus (if present), Thyroid (parathyroid), Tongue, Trachea, Ureters, Urethra, Urinary bladder, Uterus, Vagina, Zymbal glands and all tissues showing abnormalities - Statistics:
- Dunnett-Test in connection with a variance analysis (body and organ weight data)
Analysis of variance followed by Dunnett test (Erythrocytes, Hemoglobin, Hematocrit, Mean Corpuscular Hemoglobin, Mean Corpuscular Hemoglobin Concentration, Mean Corpuscular Volume Erythrocytes, Thrombocytes, Albumin, Creatinine, Chloride, Glucose, Protein, Triglycerides, Urea)
Kruskal-Wallis-Test with a Steel-Test (food and water intake data)
Kruskal-Wallis test followed by adjusted U test (Heinz Bodies, Hepato Quick, Leucocytes, Reticulocytes, Band NeutrophiIs, EosinophiIs, Lymphocytes, Monocytes, Segmented Neutrophils, Bilirubin total, Gamma-Gtutamyltransferase, Specific Gravity, Glucose, dipstick, pH, dipstick, Protein Quantitative, Volume, pH)
Adjusted Welsh test (Atanine aminotransferase, Alkaline phosphatase, Aspartate aminotransferase, Choiinesterase, Choiinesterase in Erythrocytes, Choiinesterase in Brain, Cholesterol, Calcium, Glutamate dehydrogenase, Potassium, Sodium, Inorganic Phosphate)
Results and discussion
Results of examinations
- Clinical signs:
- no effects observed
- Description (incidence and severity):
- no effects observed
- Mortality:
- mortality observed, non-treatment-related
- Description (incidence):
- All dose groups: a higher mortality in females was evident from Week 52 onwords; dose correlation is lacking
For details please refer to Table 1 in "any other information on results incl. tables". - Body weight and weight changes:
- effects observed, treatment-related
- Description (incidence and severity):
- 3000/4000 ppm: body weight depression in males from Week 7 / in females from Week 13 onwords
For details please refer to Table 2 in "any other information on results incl. tables". - Food consumption and compound intake (if feeding study):
- effects observed, treatment-related
- Description (incidence and severity):
- 3000/4000 ppm: slightly elevated food intake per kg body weight in both sexes (12.3%/19.2%)
The test substance intake in the treatment groups roughly corresponds to the theoretical dose intervals.
For details please refer to Table 3 in "any other information on results incl. tables". - Food efficiency:
- effects observed, treatment-related
- Description (incidence and severity):
- 3000/4000 ppm: indication for slightly reduced food efficiency in both sexes
For details please refer to Table 3 in "any other information on results incl. tables". - Water consumption and compound intake (if drinking water study):
- no effects observed
- Description (incidence and severity):
- no effects observed
For details please refer to Table 4 in "any other information on results incl. tables". - Ophthalmological findings:
- effects observed, treatment-related
- Description (incidence and severity):
- 1000 and 3000 ppm: higher incidence of moderate or severe opacities in the (whole) lens cortex of males at termination
For details please refer to Table 5 in "any other information on results incl. tables". - Haematological findings:
- effects observed, treatment-related
- Description (incidence and severity):
- 3000/4000 ppm: decreased Hb and Ht in males in Week 26/27 only, decreased MCH and MCHC in females
For details please refer to Tables 6 and 7 in "any other information on results incl. tables". - Clinical biochemistry findings:
- effects observed, non-treatment-related
- Description (incidence and severity):
- 3000/4000 ppm: slightly increased gamma glutamyltransferase (GOT) activity in both sexes), but not at all dates and without relevant time-dependent increase
Slightly decreased ASAT and APh activities in animals dosed at 1000 ppm and higher are not considered to indicate an adverse effect, since the values were within the physiological range.
For details please refer to Table 8 in "any other information on results incl. tables". - Urinalysis findings:
- effects observed, non-treatment-related
- Description (incidence and severity):
- Quantitative urinalyses did not reveal a treatment-related effect on pH, sediment, proteinurea, urine volume or urine density. Significant deviations from control values in these parameters were not considered relevant, since dose correlations were lacking.
For details please refer to Tables 10 and 11 in "any other information on results incl. tables". - 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):
- Interim necropsy:
3000/4000 ppm: significantly increased liver weights (absolute and relative ) in both sexes
Terminal necropsy:
3000/4000 ppm: partly significantly increased liver weights (absolute and relative) in both sexes, slightly decreased spleen weights (absolute and relative) in both sexes, increased relative testis weights in males
For details please refer to Tables 12 and 13 in "any other information on results incl. tables". - Gross pathological findings:
- effects observed, non-treatment-related
- Description (incidence and severity):
- Interim necropsy:
no effects observed
Terminal necropsy:
3000/4000 ppm: 6 males and 5 females appeared skinny - Neuropathological findings:
- not examined
- Histopathological findings: non-neoplastic:
- effects observed, treatment-related
- Description (incidence and severity):
- Interim examinations:
200 ppm: diffuse hyperplasia in the transitional epithelium of the urinary bladder in males
1000 ppm: diffuse hyperplasia in the transitional epithelium of the urinary bladder in males, hepatocellular hypertrophy (centrilobular) in both sexes
3000/4000 ppm: diffuse hyperplasia in the transitional epithelium of the urinary bladder, hepatocellular hypertrophy (centrilobular) and slight increase in the incidence and severity of vacuolation of zona fasciculata cells in the adrenal cortex in both sexes
For details please refer to Table 14 in "any other information on results incl. tables".
Terminal examinations:
1000 ppm: centrilobular hepatocytes with cytoplasmic changes and/or hypertrophy more frequently and decreasing tendency for pituitary hyperplasia as well as for degenerative alterations in the adrenals, tongue, heart and kidney in females
3000/4000 ppm: eosinophilic foci, focal degenerative changes (periportal) and cytoplasmic changes of hepatocytes (periportal and centrilobular) in the liver of males, peliotic foci, single cell necrosis and cytoplasmic changes of hepatocytes (centrilobular) and/or hypertrophy in the liver of females;
increased number of animals with a focal or diffuse simple transitional hyperplasia in the urinary bladder, increases of Proliferating Cell Nuclear Antigen (PCNA) labelling index of the urinary bladder in the hyperplastic segment of the transitional epithelium of females;
increased incidence of follicular hyperplasia (males) and mineralization of the follicular colloid (both sexes) in thyroid glands;
increased number of males with increase in vacuolation of cells in the zona fasciculata of the adrenal glands;
higher frequency of atrophy and/or fibre degenerations of the skeletal muscle in females;
markedly higher incidence and degree of degenerative myelinopathy in the sciatic nerve in both sexes;
decreasing tendency for pituitary hyperplasias as well as for degenerative alterations in the adrenals, tongue, heart and kidney in females;
dilated glands in the stomach mucosa and increase in pigment storage in the spleen of females
For details please refer to Table 15 in "any other information on results incl. tables". - Histopathological findings: neoplastic:
- effects observed, treatment-related
- Description (incidence and severity):
- Interim examinations:
In animals scheduled for interim kill, a no dose-related occurrence of a tumour type was evident.
Terminal examinations:
1000 ppm: adenomas in the thyroid follicles of one male
3000/4000 ppm: one transitional cell carcinoma and two transitional cell papillomas in the urinary bladder of females and one transitional carcinoma in the urethra of a male;
adenomas/carcinoma in the thyroid follicles of males (incidence per increasing dose group: 0-0-0-1/0-2/1)
For details please refer to Tables 16 to 19 in "any other information on results incl. tables". - Other effects:
- effects observed, treatment-related
- Description (incidence and severity):
- ChE activity plasma
No relevant treatment-related effects observed.
ChE activity erythrocytes
1000 ppm: significantly reduced ChE activity in both sexes
3000/4000 ppm: significantly reduced activity in both sexes
ChE activity brain
1000 ppm: reduced ChE activity in females
3000/4000 ppm: reduced ChE activity in both sexes
For details please refer to Table 9 in "any other information on results incl. tables". - Details on results:
- Mortality:
The number of males which had died unscheduled was equally distributed among the groups up to 3000 ppm. In all female treatment groups a higher mortality is evident from Week 52 onwards. Since a dose correlation is lacking in the wide dose range (50 and 4000 ppm) a treatment effect is not assumed. Neither for males nor for females did the global comparison of the survival curves (Wilcoxon Test) gave any indication of statistically relevant differences between the groups (p = 0.5782).
Body weight and weight changes:
There were no significant and dose-dependent effects on body weights up to 1000 ppm in males. Males ingesting 3000 ppm gained less body weight from week 7 onwards with a maximum difference of 11.0% in Week 105. The differences were significant in most cases. In female rats there is also no toxicologically relevant effect on body weights up to the dose of 1000 ppm. The few significantly lower means at 1000 ppm (between Weeks 43 to 61) are not interpreted as an adverse effect, since deviations to control data are very small (maximal 6.3% in Week 51). At 4000 ppm a significant (from Week 13 onwards) body weight depression was observed with a maximum of 14.3% in Week 85.
Food consumption and compound intake:
The food intake per animal at doses of up to 3000 ppm (males) or 4000 ppm (females) was comparable to that of untreated rats. The slightly elevated intake data per kg body weight in rats receiving 3000 (12.3%) or 4000 ppm (19.2%) could be interpreted as an indication of a slightly reduced feed efficiency.
Opthalmological findings:
Males of the 1000 and 3000 ppm groups showed a higher incidence of moderate or severe opacities in the (whole) lens cortex. The highest frequency of lenses showing slight (wedge shaped) waterclefts, being a pre-stage of cortical opacities, was noted in control males. This incidence distribution indicates a shift of lens waterclefts towards cortical opacities in 1000 and 3000 ppm males. In 4000 ppm females no remarkable lens findings could be detected compared to controls. Concerning the remaining eye compartments no treatment effect is visible.
Haematological findings:
There were no toxicologically relevant changes in the erythrocyte parameters (count, MVC, MCHC and MCH), in the haemoglobin concentration or in the haematocrit in males or females up to 1000 ppm. Some significant differences in these groups and parameters are not considered as a sign of toxicity, since deviations to control values were very small (MCV, MCH and elevated MCHC means in treated males in Week 26/27) and/or a dose correlation was lacking (erythrocyte counts). The haemoglobin concentration and haematocrit values were significantly reduced in 3000 males at Week 26/27, but not thereafter. Other significant changes in this group were slight and without corroborating findings. In 4000 ppm females no significant changes were seen with regard to haemoglobin concentration, haematocrit and erythrocyte counts but the calculation of MCH and MCHC revealed consistently lower (partly p<0.01) values than in controls and remaining groups. Analysis of erythrocyte morphology did not reveal abnormalities that could be regarded as a toxic effect. No relevant changes in platelet counts were found in treated males and females. The very slight increase in the means of platelet count of 4000 ppm females in Weeks 26/27 and 52 do not reflect an adverse effect, since individual values were within the reference range. The means of the thromboplastin time in the treatment groups were not remarkably different from those at 0 ppm. No changes of reticulocytes were observed up to the highest dose level in both sexes. No toxicologically relevant changes in leukocyte counts and differential blood counts were visible in the dose range investigated.
Clinical biochemistry findings:
Neither male nor female rats showed any remarkable changes in the plasma activity of aspartate aminotransferase (ASAT), alkaline phosphatase (APh) and glutamate dehydrogenase (GLDH) up to 200 ppm. At higher doses there was a tendency towards slightly (sometimes significantly) lower aspartate aminotransferase (ASAT) and alkaline phosphatase (APh) activities. These changes were not considered to indicate an adverse effect, since the values were within the physiological range. The activity of the alanine aminotransferase (ALAT) was unchanged in all treatment groups. A trend towards slightly elevated gamma glutamyltransferase (GOT) activity was at 3000 or 4000 ppm (p<0.01), but not at all dates and without relevant time dependent increase.
Urinalysis findings:
In Week 98/99 quantitative pH measurements were performed by a microelectrode. Results of these determinations do not indicate any influence of the test substance on the urine pH in the dose range investigated. Scanning-electron-microscopy of urine sediment did not reveal any indication of a change in amount or morphology of urine crystals due to the treatment. Quantitative urinalyses did not reveal a treatment-related effect on proteinurea, urine volume or urine density. Significant deviations from control values in these parameters were not considered relevant, since dose correlations were lacking.
Organ weight findings including organ body weight ratios:
At interim necropsy all absolute and relative organ weights except the liver weights were comparable with those of controls. A few significant changes can be attributed to differences in body weight. The liver weights were not remarkably changed up to 1000 ppm. In 3000 ppm males they were significantly increased by 18.5% (absolute) and 17.6% (relative), whereas in 4000 ppm females the increase was 18.8% (absolute) and 41.7% (relative). At terminal necropsy there were no toxicologically relevant differences between the dose groups and the controls with respect to the weights of the brain, adrenals, heart, kidneys or ovaries. The few differences marked as significant in these organs were very small and attributable to varying body weights. The liver weights were inconspicuous up to 1000 ppm. At higher concentrations absolute and relative liver weights were increased (partly p<0.01). The relative means were 20.2% ( 3000 ppm males) or 32.9% (4000 ppm females) above the corresponding control value. Up to 1000 ppm spleen and testis weights were unremarkable. At the top concentration levels slightly lower (up to -23%) spleen weights (absolute and relative) were seen in both sexes and the relative testes weights were elevated (p<0.01) by about 17%. These deviations were partly significant.
Gross pathological findings:
No treatment-related macroscopical findings were observed in rats scheduled for a treatment period of 12 months. Gross-pathological findings recorded at the necropsies performed during the in life phase in case of unscheduled deaths or at the end of the study revealed no evidence of dose-related organ lesions in any of the groups up to and including doses of 3000 or 4000 ppm. There were six 3000 ppm males and five 4000 ppm females that appeared skinny at necropsy.
Histopathological findings (non-neoplastic):
In animals scheduled for a 12-months treatment revealed test substance-related lesions in the adrenals, liver and urinary bladder. In the adrenal cortex a slight increase in the incidence and severity of vacuolation of the zona fasciculata cells was recorded at 3000 or 4000 ppm. A minimal to moderate hepatocellular hypertrophy, predominantly centrolobular, was noted in rats at 1000 ppm and above.
In the urinary bladder a diffuse hyperplasia (minimal to slight) was observed in the transitional epithelium at 200 ppm and above (males) or in the group 4000 ppm (females). For the lesions in the urinary bladder, liver and adrenals a statistically significant trend was established. All other microscopical findings recorded were considered to be incidental. In animals scheduled for terminal kill, there were no treatment-related non-neoplastic lesions at concentrations of up to 1000 ppm in males and 200 ppm in females. In the liver of 3000 ppm males eosinophilic foci, focal degenerative changes (periportal) and cytoplasmic changes of hepatocytes (periportal and centrilobular) were noted with a markedly higher incidence than in the other groups. These findings were associated with a reduced number of clear cell foci at 3000 ppm. In males of the groups 0, 50, 200 and 1000 ppm and in all treated females the frequency of all these findings did not exceed the reference range of rats of this age. In females centrilobular hepatocytes with cytoplasmic changes and/or hypertrophy were detected more frequently from 1000 ppm onwards. Additionally, in 4000 ppm females peliotic foci and single cell necrosis were noted with increased incidences. Most of the liver lesions mentioned were found to show a statistically significant positive trend. The number of animals with a focal or diffuse simple transitional hyperplasia in the urinary bladder was increased (trend mostly p<0.01) in high dose males and females. In 4000 ppm females atrophy (p<0.01) and/or fibre degenerations (p<0.05) of the skeletal muscle were found with a higher frequency than in the other groups. In 3000 ppm males and 4000 ppm females a degenerative myelinopathy was detected in the sciatic nerve with a markedly higher incidence (trend p<0.01) and a higher degree than in the other groups, where the frequency of this lesion did not exceed the range normally found in old rats. In the thyroid glands the incidence of follicular hyperplasia (males) and mineralization of the follicular colloid (males and females) was significantly increased in 3000/4000 ppm rats. At 3000 ppm there were more (trend p<0.01) males exhibiting an increase in vacuolation of cells in the zona fasciculata of the adrenal glands. Statistically significant increases of proliferating cell nuclear antigen (PCNA) labeling index of the urinary bladder were only achieved in the hyperplastic segment of the transitional epithelium of 4000 ppm females. In the bladder of 1000 ppm females and in the non-hyperplastic bladder segment of the 4000 ppm rats the calculated values were similar to those of control females.The remaining observed findings were considered to be secondary possibly due to retarded aging, which is often seen in toxicological studies, if body weight is reduced as in the present study. These findings included a decreasing tendency for pituitary hyperplasia as well as for degenerative alterations in the adrenals, tongue, heart and kidney essentially in 1000 and 4000 ppm females. The elevated (p<0.05) number of 4000 ppm females showing dilated glands in the stomach mucosa was considered unspecific and not treatment-related. Also the increase in pigment storage in the spleen (p<0.01) was not considered treatment-related.
Histpathological findings (neoplastic):
In animals scheduled for interim kill, a no dose-related occurrence of a tumour type was evident. At the terminal kill, one transitional cell carcinoma and two transitional cell papillomas (trend p<0.01) were detected in the urinary bladder of 4000 ppm females, whereas no bladder tumour occurred in the remaining female groups or in males. Additionally, one transitional carcinoma was seen in the urethra of a 3000 ppm male. In thyroid follicles of males adenomas were observed with a frequency of 0-0-0-1-2 (trend p<0.05) and carcinomas with an incidence of 0-0-0-0-1. Uterine adenocarcinomas were noted in 4000 ppm females more often (trend p<0.01) than in the other groups. Additionally, there was one squamous cell carcinoma (p<0.05) in this group. However, the incidences of both tumour types do not exceed the historical control range and are therefore, considered to be not compound-induced. Decreased tumour incidences were noted for the mammary fibroadenomas(15-10-7-6-5) and pituitary adenomas (24-18-24-14-11) in the female treatment groups. The incidences of the remaining tumours were not distributed in a treatment-related manner and comparable with those known from own historical control collectives. The number of cases, in which a tumour type occurred in one group only was comparable in the 3000/4000 ppm and 0 ppm group (for details please refer to Tables 16 in "any other information on results incl. tables".). The overall tumour incidence and the total number of benign and malignant neoplasms per group were not increased in treated males and females at all concentrations (for details please refer to Table 17 in "any other information on results incl. tables"). The number of malignant tumours was even reduced in males receiving 200 ppm and above. No remarkable differences were observed deaths in the number of tumour bearing rats or animals with a benign, malignant or benign and malignant tumour between the control and treatment groups for both, intercurrent or scheduled deaths (for details please refer to Table 18 in "any other information on results incl. tables"). As regards the time of occurrence of the tumours, there were no significant differences between treated rats and controls (for details please refer to Table 19 in "any other information on results incl. tables").
Other findings: Cholinesterase activities (ChE)
No significant ChE inhibition occurred in the plasma. The 24% lower plasma ChE activity of 3000 ppm males (Week 78) is not regarded as toxicologically relevant, because this was an isolated event, where in the control group relatively high activities were measured compared to those of the other time points. In addition, plasma ChE activity was even not affected when e.g. ChE activity in erythrocytes was inhibited by 90%. No remarkable ChE inhibition in erythrocytes were detected in males and females up to 200 ppm. At higher concentrations a dose-dependently and significantly lower ChE activity (inhibited up to 82% in males and 90% in females) was measured compared to control means. The significantly lower activity in 200 ppm females in Week 52/53 is considered to be incidental, because activity at this dose level was not remarkably inhibited at the other three time points, where stronger effects were noted in the two highest dose groups. Also in Week 52/53, a relatively high activity in controls may have contributed. Up to 3000 ppm (males) and 200 ppm (females) no significant or biologically relevant effect was noted on the brain ChE activity. In females there was a significantly reduced activity at 1000 and 4000 ppm (up to 41%). The inhibition values of 13 and 17% in 1000 ppm females and those of 14% calculated for 3000 ppm males at termination might be of questionable toxicological relevance.
Effect levels
open allclose all
- Key result
- Dose descriptor:
- NOAEL
- Effect level:
- 200 ppm
- Based on:
- test mat.
- Sex:
- male/female
- Basis for effect level:
- other: no adverse effects observed up to this dose level
- Remarks on result:
- other: corresponding to: 10.3 mg/kg bw/day (males) / 14.6 mg/kg bw/day (females)
- Key result
- Dose descriptor:
- LOAEL
- Effect level:
- 1 000 ppm
- Based on:
- test mat.
- Sex:
- male/female
- Basis for effect level:
- histopathology: non-neoplastic
- other: Cholinesterase activity in erythrocytes and brain
- Remarks on result:
- other: corresponding to: 52.7 mg/kg bw/day (males) / 75.4 mg/kg bw/day (females)
Target system / organ toxicity
open allclose all
- Key result
- Critical effects observed:
- yes
- Lowest effective dose / conc.:
- 1 000 ppm
- System:
- hepatobiliary
- Organ:
- liver
- Treatment related:
- yes
- Dose response relationship:
- yes
- Relevant for humans:
- not specified
- Key result
- Critical effects observed:
- yes
- Lowest effective dose / conc.:
- 1 000 ppm
- System:
- nervous system
- Organ:
- other: cholinesterase activity inhibited in erythrocytes (1000 ppm and above) and brain (4000 ppm) (statistically significant inhibition by 20% or more in erythrocytes or brain is considered a clear toxicological effect)
- Treatment related:
- yes
- Dose response relationship:
- yes
Any other information on results incl. tables
Table 1: Cumulative Mortality data (number of dead animals up to week indicated)
Dose (ppm) |
0 |
50 |
200 |
1000 |
3000 |
sex (male) |
|||||
Weeks 0-52 (n = 60) |
|||||
Week 26 |
0 |
1 |
0 |
0 |
0 |
Week 53 |
0 |
1 |
0 |
2 |
1 |
Weeks 53-104 (n = 50) |
|||||
Week 79 |
5 |
4 |
1 |
5 |
1 |
Week 104 |
19 |
17 |
21 |
17 |
13 |
Dose (ppm) |
0 |
50 |
200 |
1000 |
4000 |
sex (female) |
|||||
Weeks 0-52 (n = 60) |
|||||
Week 26 |
0 |
0 |
0 |
0 |
0 |
Week 53 |
0 |
2 |
1 |
0 |
2 |
Weeks 53-104 (n = 50) |
|||||
Week 79 |
3 |
7 |
8 |
8 |
9 |
Week 104 |
13 |
19 |
22 |
20 |
21 |
Table 2: Body weights [g]
sex |
m |
m |
m |
m |
m |
f |
f |
f |
f |
f |
Dose (ppm) |
0 |
50 |
200 |
1000 |
3000 |
0 |
50 |
200 |
1000 |
4000 |
0 1 2 3 4 5 6 7 8 9 10 11 12 13 15 17 19 21 23 25 27 29 31 33 35 37 39 41 43 45 47 49 51 53 55 57 59 61 63 65 67 69 71 73 75 77 79 81 83 85 87 89 91 93 95 97 99 101 103 105 |
155 203 239 272 299 322 336 362 378 382 393 403 412 427 435 456 463 471 472 477 482 492 492 502 500 510 515 519 509 517 519 523 526 531 532 535 535 542 542 542 543 553 552 553 558 550 551 558 555 559 554 553 550 549 550 553 551 544 542 543 |
156 203 243 276 304 325 344 362 378 394 406 417 428 436 448 465 468 479 480 486 494 498 503 509 507 515 518 521 519 523 521 528 532 530 531 534 534 536 540 539 541 548 549 548 550 552 543 551 549 548 542 537 529 530 529 534 530 520 508 + 511 |
155 203 245 278 306 329 348 364 378 395 406 417 423 430 444 459 463 470 480 487 493 495 503 507 509 508 514 516 515 518 520 524 529 533 534 538 539 539 541 541 542 547 545 541 544 539 536 536 538 532 520 + 522 513 + 527 528 522 519 510 + 514 529 |
154 200 241 275 301 326 341 356 368 385 401 400 417 420 438 448 455 464 472 480 483 488 496 495 502 509 507 510 512 512 513 519 522 525 527 530 534 533 532 538 542 549 547 547 550 550 552 556 555 553 549 547 533 542 535 531 528 523 518 515 |
152 189++ 238 264 289 312 330 344++ 360++ 372 382 391 401 401++ 421 427++ 441+ 449+ 454 463 466 468+ 480 479+ 485 483++ 486++ 489++ 492 490++ 490++ 496++ 500++ 502++ 505+ 506++ 513 511++ 512++ 515+ 516+ 521++ 519++ 516++ 518++ 512 ++ 523 + 524 ++ 527 + 526 + 518 + 513 ++ 493 ++ 504 ++ 498 ++ 497 ++ 492 ++ 495 ++ 491 ++ 483 ++ |
130 149 163 176 186 196 205 212 217 222 225 229 234 242 242 247 249 255 258 260 261 267 271 277 276 280 284 286 289 290 295 296 301 302 304 309 311 312 314 315 319 323 322 325 329 333 328 333 333 336 327 327 329 327 326 328 323 323 318 313 |
127 145 161 173 183 191 200 208 212 216 222 226 230 234 240 243 247 251 254 257 257 261 266 271 273 275 279 281 284 285 289 288 291 291 294 298 300 303 307 307 309 316 314 318 321 323 321 329 330 331 326 325 324 324 325 327 325 324 315 325 |
130 150 166 179 190 199 207 211 217 223 228 232 238 239 244 249 252 258 260 263 264 266 272 276 280 281 285 288 289 289 293 293 297 302 304 307 309 308 312 315 316 322 322 325 327 328 327 332 333 337 332 330 331 327 329 326 325 320 312 341 |
128 147 164 178 186 196 203 207 213 219 221 228 231 234 237 242 246 249 252 254 257 257 261 266 270 270 274 276 276+ 275+ 280+ 279++ 282++ 284+ 287+ 291+ 292+ 294+ 300 301 303 308 307 311 313 314 310 320 320 320 316 314 317 317 313 311 311 312 307 320 |
130 147 167 179 189 196 202 205 209 215 217 222 225 226++ 230+ 234++ 234++ 238++ 242++ 245++ 246++ 248++ 252++ 253++ 257++ 258++ 262++ 264++ 264++ 263++ 266++ 266++ 268++ 270++ 271++ 274++ 275++ 279++ 278++ 280++ 281++ 285++ 283++ 283++ 286++ 284 ++ 284 ++ 289 ++ 291 ++ 288 ++ 286 ++ 288 ++ 287 ++ 289 ++ 289 ++ 286 ++ 283 ++ 284 ++ 281 ++ 295 |
+ difference against control for p<0.05
++ difference against control for p<0.01
Table 3: Mean daily and cumulative intake of food and test compound
Intake food |
||||||
|
|
|
g/animal |
g/kg bw |
||
Dose (ppm) |
Sex |
Days |
Total |
per Day |
Total |
per Day |
0 |
m |
708 |
14715 |
20.8 |
35813 |
50.6 |
50 |
m |
708 |
14923 |
21.1 |
35653 |
50.4 |
200 |
m |
708 |
15169 |
21.4 |
36331 |
51.3 |
1000 |
m |
708 |
15391 |
21.7 |
37307 |
52.7 |
3000 |
m |
708 |
15807 |
22.3 |
40218 |
56.8 |
0 |
f |
708 |
12072 |
17.1 |
48515 |
68.5 |
50 |
f |
708 |
12522 |
17.7 |
51527 |
72.8 |
200 |
f |
708 |
12729 |
18.0 |
51524 |
72.8 |
1000 |
f |
708 |
12712 |
18.0 |
53372 |
75.4 |
4000 |
f |
708 |
13177 |
18.6 |
57821 |
81.7 |
Intake of test compound |
||||||
|
|
|
mg/animal |
mg/kg bw |
||
Dose (ppm) |
Sex |
Days |
Total |
per Day |
Total |
per Day |
50 |
m |
708 |
746 |
1.1 |
1783 |
2.5 |
200 |
m |
708 |
3034 |
4.3 |
7266 |
10.3 |
1000 |
m |
708 |
15391 |
21.7 |
37307 |
52.7 |
3000 |
m |
708 |
47421 |
67.0 |
120654 |
170.4 |
50 |
f |
708 |
626 |
0.9 |
2576 |
3.6 |
200 |
f |
708 |
2546 |
3.6 |
10305 |
14.6 |
1000 |
f |
708 |
12712 |
18.0 |
53372 |
75.4 |
4000 |
f |
708 |
52707 |
74.4 |
231285 |
326.7 |
Table 4: Mean daily cumulative water intake
Intake of water |
||||||
|
|
|
g/animal |
g/kg bw |
||
Dose (ppm) |
Sex |
Days |
Total |
per Day |
Total |
per Day |
0 |
m |
708 |
18267 |
25.9 |
40827 |
57.8 |
50 |
m |
708 |
18971 |
26.9 |
41396 |
58.6 |
200 |
m |
708 |
19254 |
27.3 |
42187 |
59.8 |
1000 |
m |
708 |
19034 |
27.0 |
42158 |
59.7 |
3000 |
m |
708 |
19391 |
27.5 |
44915 |
63.6 |
0 |
f |
708 |
18478 |
26.2 |
67633 |
96.8 |
50 |
f |
708 |
18745 |
26.6 |
70078 |
99.3 |
200 |
f |
708 |
18848 |
26.7 |
68707 |
97.3 |
1000 |
f |
708 |
18539 |
26.3 |
70478 |
99.8 |
4000 |
f |
708 |
17344 |
24.6 |
69761 |
98.8 |
Table 5: Ophthalmological findings at termination (Incidences in %)
sex |
m |
m |
m |
m |
f |
f |
Dose (ppm) |
0 |
200 |
1000 |
3000 |
0 |
4000 |
No. of eyes examined |
61 |
71 |
61 |
73 |
76 |
58 |
Turbid cornea, including vascularisation of cornea |
1.6 |
7.0 |
1.6 |
8.2 |
0 |
3.4 |
Turbid cornea, including vascularisation of cornea Snow ball like turbidities between lens and vitreous body (= posterior lenticular rupture) |
11.4 |
4.2 |
8.2 |
13.7 |
7.9 |
0 |
Diffuse retrolenticular opacity (all degrees) |
24.6 |
9.8 |
18.0 |
17.8 |
10.5 |
8.6 |
Wedge shaped waterclefts in the cortex, slight |
18.0 |
7.0 |
8.2 |
8.2 |
3.9 |
5.1 |
Wedge shaped waterclefts in the cortex, moderate |
0 |
0 |
3.3 |
1.4 |
0 |
1.7 |
Lens opacity in the whole cortex, slight |
1.6 |
1.4 |
0 |
2.7 |
2.6 |
1.7 |
Lens opacity in the whole cortex, moderate |
1.6 |
1.4 |
1.6 |
5.5 |
3.9 |
0 |
Lens opacity in the whole cortex, severe |
1.6 |
0 |
6.5 |
5.5 |
6.6 |
3.4 |
Table 6: Hematology
Dose |
|
|
LEU |
ERY |
HB |
HCT |
MCV |
MCH |
MCHC |
RETI |
HEINZ |
THRO |
HQUICK |
|
ppm |
Sex |
Week |
109/L |
109/L |
g/L |
L/L |
fl |
pg |
g/LERY |
0/00 |
0/00 |
109/L |
sec |
|
0 |
m |
26/27 |
10.3 |
9.04 |
147 |
0.459 |
50.8 |
16.3 |
320 |
18 |
0 |
1040 |
25.4 |
|
50 |
m |
26/27 |
8.7 |
8.91 |
147 |
0.453 |
50.9 |
16.5 |
324 |
19 |
0 |
998 |
25.2 |
|
200 |
m |
26/27 |
8.3 |
8.51 |
141 |
0.439 |
51.6 |
16.5 |
320 |
18 |
0 |
1109 |
24.7 |
|
1000 |
m |
26/27 |
8.3 |
9.06 |
141 |
0.434 |
48.0++ |
15.6 |
325 |
17 |
0 |
1053 |
26.2 |
|
3000 |
m |
26/27 |
8.0 |
8.58 |
138+ |
0.425++ |
49.6 |
16.1 |
325 |
17 |
1 |
982 |
25.4 |
|
0 |
m |
52 |
9.5 |
9.42 |
153 |
0.487 |
51.8 |
16.2 |
313 |
16 |
0 |
1082 |
26.3 |
|
50 |
m |
52 |
8.5 |
9.17 |
149 |
0.479 |
52.3 |
16.3 |
312 |
17 |
0 |
1059 |
26.0 |
|
200 |
m |
52 |
8.4 |
8.87+ |
148 |
0.471 |
53.1 |
16.7 |
314 |
18 |
0 |
1172 |
26.1 |
|
1000 |
m |
52 |
7.4 |
9.37 |
146 |
0.462 |
49.4 |
15.6 |
315 |
20 |
0 |
1082 |
25.4 |
|
3000 |
m |
52 |
8.2 |
9.29 |
146 |
0.472 |
50.8 |
15.7 |
309 |
20 |
0 |
1026 |
26.3 |
|
0 |
m |
78 |
8.0 |
9.04 |
151 |
0.476 |
52.7 |
16.8 |
318 |
17 |
0 |
1091 |
24.7 |
|
50 |
m |
78 |
7.3 |
8.99 |
149 |
0.472 |
52.5 |
16.6 |
316 |
14 |
0 |
1073 |
24.6 |
|
200 |
m |
78 |
6.7 |
8.20+ |
141 |
0.441 |
53.7 |
17.2 |
319 |
16 |
0 |
1277 |
24.0 |
|
1000 |
m |
78 |
6.4 |
8.93 |
141 |
0.441 |
49.5+ |
15.9+ |
321 |
17 |
0 |
1041 |
24.1 |
|
3000 |
m |
78 |
7.5 |
8.77 |
139 |
0.440 |
50.2 |
15.9+ |
316 |
15 |
0 |
998 |
24.1 |
|
0 |
m |
103/104 |
9.2 |
9.26 |
160 |
0.512 |
55.4 |
17.3 |
312 |
20 |
0 |
1220 |
25.9 |
|
50 |
m |
103/104 |
7.9 |
8.63 |
148 |
0.457+ |
52.9 |
17.1 |
322++ |
27 |
0 |
1158 |
25.0 |
|
200 |
m |
103/104 |
7.9 |
8.85 |
154 |
0.473 |
53.5 |
17.5 |
326++ |
16 |
0 |
1156 |
24.2 |
|
1000 |
m |
103/104 |
7.3 |
8.73 |
144 |
0.443++ |
50.9++ |
16.5 |
324++ |
23 |
0 |
1186 |
24.8 |
|
3000 |
m |
103/104 |
6.9 |
9.07 |
150 |
0.466 |
51.5++ |
16.5 |
321+ |
29 |
0 |
1081 |
25.8 |
|
0 |
f |
26/27 |
6.0 |
8.21 |
143 |
0.439 |
53.5 |
17.4 |
325 |
15 |
0 |
915 |
24.5 |
|
50 |
f |
26/27 |
6.2 |
7.94 |
136 |
0.416 |
52.4 |
17.1 |
328 |
15 |
0 |
978 |
23.7 |
|
200 |
f |
26/27 |
5.9 |
8.10 |
139 |
0.422 |
52.2 |
17.2 |
330 |
14 |
0 |
979 |
23.8 |
|
1000 |
f |
26/27 |
6.2 |
8.01 |
137 |
0.424 |
52.8 |
17.1 |
325 |
17 |
0 |
955 |
23.8 |
|
4000 |
f |
26/27 |
5.8 |
8.57 |
139 |
0.434 |
50.6++ |
16.2++ |
320 |
17 |
0 |
1046++ |
22.4 |
|
0 |
f |
52 |
4.6 |
8.03 |
140 |
0.440 |
54.8 |
17.4 |
318 |
20 |
0 |
809 |
24.7 |
|
50 |
f |
52 |
5.8 |
8.15 |
138 |
0.437 |
53.6 |
17.0 |
317 |
18 |
0 |
921 |
25.4 |
|
200 |
f |
52 |
5.7 |
8.26 |
142 |
0.440 |
53.4 |
17.2 |
322 |
17 |
0 |
956 |
25.2 |
|
1000 |
f |
52 |
5.7 |
8.25 |
140 |
0.447 |
54.2 |
17.0 |
314 |
19 |
0 |
897 |
24.1 |
|
4000 |
f |
52 |
5.8 |
8.36 |
136 |
0.440 |
53.1 |
16.4+ |
309++ |
20 |
0 |
1017+ |
23.8 |
|
0 |
f |
78 |
4.1 |
8.33 |
146 |
0.454 |
54.6 |
17.5 |
321 |
25 |
0 |
858 |
23.2 |
|
50 |
f |
78 |
4.6 |
7.79 |
136 |
0.429 |
56.6 |
17.8 |
316 |
44 |
0 |
795 |
23.5 |
|
200 |
f |
78 |
5.6 |
8.24 |
142 |
0.443 |
53.8 |
17.3 |
321 |
31 |
0 |
840 |
23.4 |
|
1000 |
f |
78 |
4.3 |
8.16 |
143 |
0.451 |
55.4 |
17.6 |
317 |
31 |
0 |
848 |
26.2++ |
|
4000 |
f |
78 |
4.7 |
8.06 |
133 |
0.431 |
53.6 |
16.6 |
309++ |
36 |
0 |
930 |
24.8+ |
|
0 |
f |
103/104 |
5.6 |
8.29 |
151 |
0.453 |
54.9 |
18.3 |
333 |
31 |
0 |
1003 |
24.0 |
|
50 |
f |
103/104 |
4.7 |
8.34 |
151 |
0.454 |
54.5 |
18.1 |
333 |
28 |
0 |
984 |
23.7 |
|
200 |
f |
103/104 |
6.8 |
8.22 |
149 |
0.450 |
54.7 |
18.1 |
330 |
26 |
0 |
994 |
24.6 |
|
1000 |
f |
103/104 |
4.3 |
8.33 |
151 |
0.463 |
55.7 |
18.1 |
325 |
29 |
0 |
948 |
24.8 |
|
4000 |
f |
103/104 |
5.1 |
8.22 |
143 |
0.443 |
54.2 |
17.4 |
322+ |
31 |
0 |
1066 |
23.9 |
|
+ difference against control for p<0.05
++ difference against control for p<0.01
Table 7: Differential blood count
Dose |
Week |
Sex |
LYM |
SEGM |
EOS |
MONO |
BAND |
ppm |
|
|
% |
% |
% |
% |
% |
0 |
26/27 |
m |
87.5 |
8.0 |
0.2 |
4.3 |
0.0 |
50 |
26/27 |
m |
89.8 |
6.5 |
0.3 |
3.4 |
0.0 |
200 |
26/27 |
m |
86.0 |
10.1 |
0.6 |
3.3 |
0.0 |
1000 |
26/27 |
m |
87.9 |
8.1 |
0.7 |
3.3 |
0.0 |
3000 |
26/27 |
m |
89.4 |
7.6 |
0.6 |
2.5 |
0.0 |
0 |
52 |
m |
79.7 |
12.7 |
1.0 |
6.6 |
0.0 |
50 |
52 |
m |
78.8 |
11.7 |
1.8 |
7.7 |
0.0 |
200 |
52 |
m |
79.1 |
12.7 |
0.9 |
7.3 |
0.0 |
1000 |
52 |
m |
77.5 |
13.6 |
1.4 |
7.6 |
0.0 |
3000 |
52 |
m |
78.0 |
13.7 |
0.6 |
7.8 |
0.0 |
0 |
78 |
m |
78.4 |
13.9 |
1.5 |
6.2 |
0.0 |
50 |
78 |
m |
75.9 |
16.2 |
2.2 |
5.6 |
0.0 |
200 |
78 |
m |
74.9 |
17.4 |
1.7 |
6.2 |
0.0 |
1000 |
78 |
m |
74.7 |
18.9 |
1.1 |
5.3 |
0.0 |
3000 |
78 |
m |
77.6 |
17.0 |
0.8 |
4.6 |
0.0 |
0 |
103/104 |
m |
67.8 |
23.5 |
1.2 |
7.4 |
0.0 |
50 |
103/104 |
m |
62.7 |
29.7 |
1.7 |
6.0 |
0.0 |
200 |
103/104 |
m |
70.0 |
23.6 |
1.5 |
4.8 |
0.1 |
1000 |
103/104 |
m |
66.6 |
25.4 |
2.6 |
5.5 |
0.0 |
3000 |
103/104 |
m |
71.8 |
23.0 |
1.2 |
4.2 |
0.0 |
0 |
26/27 |
f |
88.4 |
8.4 |
0.8 |
2.5 |
0.0 |
50 |
26/27 |
f |
86.0 |
11.3 |
0.8 |
1.9 |
0.0 |
200 |
26/27 |
f |
89.5 |
7.8 |
0.8 |
1.9 |
0.0 |
1000 |
26/27 |
f |
90.7 |
8.2 |
0.4 |
0.7+ |
0.0 |
4000 |
26/27 |
f |
88.7 |
8.5 |
0.8 |
2.0 |
0.0 |
0 |
52 |
f |
79.9 |
14.0 |
1.4 |
4.7 |
0.0 |
50 |
52 |
f |
76.5 |
16.5 |
1.0 |
6.0 |
0.0 |
200 |
52 |
f |
80.7 |
13.3 |
1.2 |
4.8 |
0.0 |
1000 |
52 |
f |
80.5 |
15.0 |
0.6 |
3.9 |
0.0 |
4000 |
52 |
f |
86.4 |
9.7 |
0.5 |
3.4 |
0.0 |
0 |
78 |
f |
71.8 |
23.7 |
1.2 |
3.3 |
0.0 |
50 |
78 |
f |
75.7 |
16.2 |
1.7 |
6.4 |
0.1 |
200 |
78 |
f |
65.9 |
26.9 |
1.0 |
6.3 |
0.0 |
1000 |
78 |
f |
73.5 |
19.8 |
1.2 |
5.6 |
0.0 |
4000 |
78 |
f |
78.4 |
16.0 |
1.1 |
4.5 |
0.0 |
0 |
103/104 |
f |
70.4 |
27.4 |
1.2 |
1.1 |
0.0 |
50 |
103/104 |
f |
69.3 |
28.8 |
0.8 |
1.1 |
0.0 |
200 |
103/104 |
f |
60.4 |
37.3 |
1.3 |
1.0 |
0.0 |
1000 |
103/104 |
f |
75.8 |
22.0 |
1.0 |
1.2 |
0.0 |
4000 |
103/104 |
f |
73.1 |
23.8 |
1.6 |
1.6 |
0.0 |
+ difference against control for p<0.05
++ difference against control for p<0.01
Table 8: Clinical Chemistry
Dose |
Week |
Sex |
ASAT (GOT) |
ALAT (GPT) |
APh |
GGT |
GLDH |
ppm |
|
|
U/L |
U/L |
U/L |
U/L |
U/L |
0 |
26/27 |
m |
41.3 |
38.9 |
190 |
1 |
5.7 |
50 |
26/27 |
m |
38.8 |
40.1 |
186 |
1 |
5.4 |
200 |
26/27 |
m |
44.4 |
40.3 |
171 |
2 |
7.9 |
1000 |
26/27 |
m |
38.5 |
40.4 |
157+ |
2 |
5.0 |
3000 |
26/27 |
m |
35.1 |
37.7 |
150+ |
3++ |
3.8 |
0 |
52 |
m |
39.9 |
42.2 |
184 |
1 |
16.2 |
50 |
52 |
m |
40.5 |
44.l |
193 |
1 |
22.8 |
200 |
52 |
m |
36.0 |
37.9 |
162 |
2 |
12.0 |
1000 |
52 |
m |
31.2 |
37.4 |
154+ |
1 |
12.6 |
3000 |
52 |
m |
34.6 |
41.0 |
156+ |
2 |
12.8 |
0 |
78 |
m |
38.8 |
36.5 |
157 |
1 |
16.0 |
50 |
78 |
m |
42.5 |
43.8 |
178 |
2 |
22.0 |
200 |
78 |
m |
35.1 |
32.1 |
142 |
1 |
14.0 |
1000 |
78 |
m |
33.2 |
36.0 |
141 |
1 |
17.9 |
3000 |
78 |
m |
32.7 |
35.7 |
144 |
2 |
20.0 |
0 |
103/104 |
m |
40.2 |
37.5 |
164 |
3 |
21.4 |
50 |
103/104 |
m |
42.2 |
33.1 |
175 |
2 |
21.6 |
200 |
103/104 |
m |
37.4 |
32.3 |
134 |
2 |
10.8 |
1000 |
103/104 |
m |
37.4 |
29.8 |
137 |
3 |
14.8 |
3000 |
103/104 |
m |
36.9 |
35.3 |
140 |
3 |
19.0 |
0 |
26/27 |
f |
40.7 |
43.4 |
121 |
0 |
11.5 |
50 |
26/27 |
f |
48.0 |
43.6 |
133 |
0 |
18.7 |
200 |
26/27 |
f |
41.0 |
44.9 |
122 |
0 |
11.3 |
1000 |
26/27 |
f |
32.3+ |
41.9 |
103 |
0 |
6.8 |
4000 |
26/27 |
f |
30.9++ |
42.6 |
105 |
1 |
1.0 |
0 |
52 |
f |
44.7 |
42.6 |
120 |
0 |
17.2 |
50 |
52 |
f |
46.8 |
44.1 |
134 |
0 |
22.9 |
200 |
52 |
f |
42.5 |
43.8 |
111 |
0 |
20.4 |
1000 |
52 |
f |
55.4 |
55.2 |
99 |
0 |
63.3 |
4000 |
52 |
f |
33.3 |
45.0 |
118 |
0 |
9.2 |
0 |
78 |
f |
59.4 |
45.7 |
100 |
1 |
59.2 |
50 |
78 |
f |
89.4 |
62.2 |
113 |
0 |
104.8 |
200 |
78 |
f |
73.9 |
54.4 |
91 |
0 |
87.7 |
1000 |
78 |
f |
40.3 |
36.3 |
81+ |
1 |
29.1 |
4000 |
78 |
f |
41.2 |
37.5 |
90 |
2+ |
18.4 |
0 |
103/104 |
f |
61.3 |
45.9 |
114 |
0 |
35.5 |
50 |
103/104 |
f |
61.3 |
49.0 |
114 |
1 |
53.7 |
200 |
103/104 |
f |
76.6 |
53.8 |
102 |
0 |
70.9 |
1000 |
103/104 |
f |
45.7 |
41.2 |
92 |
0 |
26.4 |
4000 |
103/104 |
f |
43.4 |
43.6 |
102 |
2++ |
41.5 |
+ difference against control for p<0.05
++ difference against control for p<0.01
Table 9: Activities and inhibition of cholinesterases (CHE)
Dose |
Week |
Sex |
CHE/Brain |
CHE/Plasma |
CHE/Erythrocytes |
|||
ppm |
|
|
U/g |
% Inhibition |
kU/L |
% Inhibition |
kU/L |
% Inhibition |
0 |
27 |
m |
- |
|
0.50 |
|
1.06 |
|
50 |
27 |
m |
- |
|
0.52 |
|
1.06 |
|
200 |
27 |
m |
- |
|
0.52 |
|
0.88 |
17 |
1000 |
27 |
m |
- |
|
0.50 |
|
0.65+ |
39 |
3000 |
27 |
m |
- |
|
0.45 |
10 |
0.32++ |
70 |
0 |
52/53 |
m |
10.64 |
|
0.62 |
|
1.14 |
|
50 |
52/53 |
m |
10.46 |
2 |
0.68 |
|
1.15 |
|
200 |
52/53 |
m |
10.56 |
|
0.63 |
|
1.01 |
11 |
1000 |
52/53 |
m |
10.26 |
4 |
0.61 |
2 |
1.00 |
12 |
3000 |
52/53 |
m |
9.54 |
10 |
0.56 |
10 |
0.57++ |
50 |
0 |
78 |
m |
- |
|
0.72 |
|
1.24 |
|
50 |
78 |
m |
- |
|
0.71 |
|
1.35 |
|
200 |
78 |
m |
- |
|
0.71 |
1 |
1.16 |
6 |
1000 |
78 |
m |
- |
|
0.63 |
13 |
0.84+ |
32 |
3000 |
78 |
m |
|
|
0.55 |
24 |
0.55++ |
55 |
0 |
103/104/105 |
m |
9.88 |
|
0.66 |
|
0.83 |
|
50 |
103/104/105 |
m |
9.87 |
|
0.95 |
|
0.76 |
8 |
200 |
103/104/105 |
m |
9.86 |
|
0.86 |
|
0.70 |
16 |
1000 |
103/104/105 |
m |
9.47 |
4 |
0.65 |
2 |
0.50++ |
40 |
3000 |
103/104/105 |
m |
8.50 |
14 |
0.55 |
17 |
0.15++ |
82 |
0 |
26/27 |
f |
- |
|
2.47 |
|
0.77 |
|
50 |
26/27 |
f |
- |
|
2.24 |
9 |
0.83 |
|
200 |
26/27 |
f |
- |
|
2.15 |
13 |
0.70 |
9 |
1000 |
26/27 |
f |
- |
|
2.57 |
|
0.38++ |
51 |
4000 |
26/27 |
f |
- |
|
2.30 |
7 |
0.08++ |
90 |
0 |
52/53 |
f |
11.28 |
|
2.39 |
|
1.24 |
|
50 |
52/53 |
f |
10.82+ |
4 |
2.21 |
8 |
1.17 |
6 |
200 |
52/53 |
f |
10.80 |
4 |
2.39 |
|
0.96+ |
23 |
1000 |
52/53 |
f |
9.41++ |
17 |
2.73 |
|
0.58++ |
53 |
4000 |
52/53 |
f |
7.07++ |
37 |
2.49 |
|
0.34++ |
73 |
0 |
78 |
f |
- |
|
2.18 |
|
1.19 |
|
50 |
78 |
f |
- |
|
2.03 |
7 |
1.18 |
1 |
200 |
78 |
f |
- |
|
2.12 |
3 |
1.01 |
15 |
1000 |
78 |
f |
- |
|
2.45 |
|
0.45++ |
62 |
4000 |
78 |
f |
- |
|
2.22 |
|
0.13++ |
89 |
0 |
103/104/105 |
f |
10.22 |
|
2.25 |
|
0.74 |
|
50 |
103/104/105 |
f |
10.02 |
2 |
2.31 |
|
0.66 |
11 |
200 |
103/104/105 |
f |
9.86 |
4 |
1.95 |
13 |
0.73 |
1 |
1000 |
103/104/105 |
f |
8.91++ |
13 |
2.11 |
6 |
0.28++ |
62 |
4000 |
103/104/105 |
f |
6.02++ |
41 |
2.08 |
8 |
0.08++ |
89 |
+ difference against control for p<0.05
++ difference against control for p<0.01
Table 10: Determination of urine pH week 98/99
|
pH mean value |
|
Sex |
male |
female |
Dose (ppm) |
||
0 |
7.55 |
7.20 |
50 |
7.57 |
7.47 |
200 |
7.32 |
7.12 |
1000 |
7.45 |
7.64 |
3000 |
7.34 |
- |
4000 |
- |
7.67 |
Table 11: Urinalyses
Dose (ppm) |
Week |
Sex |
VOL (ml) |
Density (g/L) |
PROT*Vol (mg) |
PROT (g/L) |
0 |
26 |
m |
13.9 |
1023 |
20.3 |
1.78 |
50 |
26 |
m |
14.8 |
1021 |
30.1 |
2.88 |
200 |
26 |
m |
12.2 |
1025 |
56.5 |
5.23 |
1000 |
26 |
m |
10.8 |
1032 |
26.7 |
3.29 |
3000 |
26 |
m |
15.6 |
1020 |
31.1 |
3.07 |
0 |
52 |
m |
6.3 |
1036 |
33.5 |
5.73 |
50 |
52 |
m |
8.2 |
1035 |
57.9 |
7.95 |
200 |
52 |
m |
7.0 |
1039 |
101.2 |
13.71 |
1000 |
52 |
m |
11.2 |
1032 |
61.4 |
9.03 |
3000 |
52 |
m |
9.3 |
1034 |
58.2 |
7.84 |
0 |
79 |
m |
7.1 |
1037 |
35.2 |
5.35 |
50 |
79 |
m |
7.8 |
1030 |
42.9 |
5.49 |
200 |
79 |
m |
12.3 |
1023 |
76.1 |
6.53 |
1000 |
79 |
m |
7.5 |
1025 |
36.4 |
5.99 |
3000 |
79 |
m |
8.4 |
1028 |
43.5 |
6.37 |
0 |
103 |
m |
11.6 |
1024 |
75.9 |
7.70 |
50 |
103 |
m |
8.6 |
1031 |
77.2 |
8.98 |
200 |
103 |
m |
10.9 |
1024 |
84.1 |
8.65 |
1000 |
103 |
m |
8.8 |
1030 |
91.4 |
9.67 |
3000 |
103 |
m |
11.8 |
1024 |
75.7 |
6.64 |
0 |
26 |
f |
8.5 |
1026 |
5.5 |
0.68 |
50 |
26 |
f |
14.0 |
1016 |
2.8 |
0.25 |
200 |
26 |
f |
12.5 |
1022 |
2.4 |
0.24+ |
1000 |
26 |
f |
6.9 |
1028 |
1.8 |
0.31 |
4000 |
26 |
f |
7.3 |
1026 |
1.9 |
0.34 |
0 |
52 |
f |
8.4 |
1029 |
13.3 |
1.25 |
50 |
52 |
f |
9.0 |
1021 |
3.2 |
0.37 |
200 |
52 |
f |
10.7 |
1022 |
4.5 |
0.45 |
1000 |
52 |
f |
12.5 |
1015+ |
2.5 |
0.32+ |
4000 |
52 |
f |
8.4 |
1030 |
1.8 |
0.29+ |
0 |
79 |
f |
5.8 |
1026 |
10.1 |
1.85 |
50 |
79 |
f |
8.0 |
1020+ |
6.3 |
0.91 |
200 |
79 |
f |
11.2+ |
1019+ |
10.9 |
1.25 |
1000 |
79 |
f |
10.8+ |
1015+ |
6.0 |
0.66 |
4000 |
79 |
f |
10.6 |
1018+ |
3.3 |
0.42 |
0 |
103 |
f |
8.4 |
1024 |
46.0 |
4.94 |
50 |
103 |
f |
10.0 |
1021 |
42.5 |
4.80 |
200 |
103 |
f |
7.9 |
1024 |
23.6 |
2.85 |
1000 |
103 |
f |
9.8 |
1022 |
19.7 |
1.16+ |
4000 |
103 |
f |
7.9 |
1029 |
7.5 |
1.14+ |
+ difference against control for p<0.05
++ difference against control for p<0.01
Table 12: Absolute and relative organ weights (mg/100 g bw) – interim kill
Dose |
Sex |
Body weight |
Brain |
Adrenals |
Heart |
Liver |
Spleen |
Kidneys |
Testes |
Ovaries |
Absolute organ weights |
g |
mg |
mg |
mg |
mg |
mg |
mg |
mg |
mg |
|
0 |
m |
517 |
2052 |
54 |
1402 |
17938 |
853 |
2910 |
3844 |
- |
50 |
m |
557 |
2159+ |
53 |
1534 |
19013 |
944 |
3220 |
4062 |
- |
200 |
m |
528 |
2064 |
54 |
1467 |
17702 |
881 |
3085 |
3815 |
- |
1000 |
m |
531 |
2128 |
52 |
1560 |
19351 |
801 |
3054 |
3652 |
- |
3000 |
m |
519 |
2105 |
56 |
1439 |
21255+ |
715 |
2998 |
4051 |
- |
0 |
f |
312 |
1912 |
84 |
1023 |
10016 |
535 |
1994 |
- |
172 |
50 |
f |
302 |
1890 |
83 |
1031 |
9690 |
561 |
2057 |
- |
173 |
200 |
f |
296 |
1920 |
83 |
1059 |
9442 |
476 |
2061 |
- |
197 |
1000 |
f |
285 |
1879 |
80 |
988 |
10299 |
684 |
1983 |
- |
204 |
4000 |
f |
261++ |
1827 |
69 |
974 |
11901++ |
488 |
1856 |
- |
162 |
Relative organ weights |
Body weight |
Brain |
Adrenals |
Heart |
Liver |
Spleen |
Kidneys |
Testes |
Ovaries |
|
g |
mg |
mg |
mg |
mg |
mg |
mg |
mg |
mg |
||
0 |
m |
517 |
400 |
10 |
272 |
3480 |
166 |
561 |
749 |
- |
50 |
m |
557 |
390 |
10 |
276 |
3424 |
170 |
580 |
731 |
- |
200 |
m |
528 |
393 |
10 |
279 |
3347 |
167 |
585 |
723 |
- |
1000 |
m |
531 |
403 |
10 |
296 |
3649 |
151 |
579 |
697 |
- |
3000 |
m |
519 |
406 |
11 |
278 |
4094+ |
137 |
578 |
782 |
- |
0 |
f |
312 |
620 |
27 |
331 |
3235 |
173 |
642 |
- |
55 |
50 |
f |
302 |
628 |
28 |
342 |
3205 |
186 |
681 |
- |
57 |
200 |
f |
296 |
652 |
28 |
357 |
3196 |
195 |
696 |
- |
68 |
1000 |
f |
285 |
660 |
28 |
347 |
3615+ |
235 |
696 |
- |
71 |
4000 |
f |
261++ |
704++ |
27 |
375+ |
4584++ |
186 |
711+ |
- |
63 |
+ difference against control for p<0.05
++ difference against control for p<0.01
Table 13: Absolute and relative organ weights (mg/100 g bw) – terminal kill
Dose |
Sex |
Body weight |
Brain |
Adrenals |
Heart |
Liver |
Spleen |
Kidneys |
Testes |
Ovaries |
Absolute organ weights |
g |
mg |
mg |
mg |
mg |
mg |
mg |
mg |
mg |
|
0 |
m |
534 |
2176 |
64 |
1866 |
18013 |
1217 |
3491 |
3785 |
- |
50 |
m |
507 |
2191 |
68 |
1960 |
17363 |
1098 |
3429 |
3929 |
- |
200 |
m |
508 |
2172 |
58 |
1830 |
18083 |
1157 |
3611 |
3847 |
- |
1000 |
m |
513 |
2165 |
71 |
18368 |
18368 |
1035+ |
3407 |
4048 |
- |
3000 |
m |
483++ |
2118 |
78 |
19494 |
19494 |
937++ |
3369 |
4029 |
- |
0 |
f |
318 |
1931 |
84 |
11395 |
11395 |
721 |
2295 |
- |
216 |
50 |
f |
318 |
1957 |
77 |
11784 |
11784 |
714 |
2380 |
- |
173 |
200 |
f |
312 |
1947 |
82 |
11347 |
11347 |
738 |
2328 |
- |
181 |
1000 |
f |
308 |
1938 |
69 |
11579 |
11579 |
671 |
2197 |
- |
193 |
4000 |
f |
282++ |
1914 |
71 |
13476++ |
13476++ |
585++ |
2190 |
- |
194 |
Relative organ weights |
Body weight |
Brain |
Adrenals |
Heart |
Liver |
Spleen |
Kidneys |
Testes |
Ovaries |
|
g |
mg |
mg |
mg |
mg |
mg |
mg |
mg |
mg |
||
0 |
m |
534 |
410 |
12 |
351 |
3369 |
229 |
655 |
712 |
- |
50 |
m |
507 |
435 |
14 |
387 |
3416 |
229 |
680 |
776 |
- |
200 |
m |
508 |
434 |
12 |
367 |
3608 |
230 |
721 |
756 |
- |
1000 |
m |
513 |
426 |
14 |
364 |
3608 |
203 |
669 |
793 |
- |
3000 |
m |
483++ |
483++ |
17 |
400+ |
4050++ |
196+ |
706 |
835++ |
- |
0 |
f |
318 |
617 |
27 |
448 |
3608 |
229 |
728 |
- |
68 |
50 |
f |
318 |
625 |
25 |
444 |
3758 |
229 |
754 |
- |
54 |
200 |
f |
312 |
633 |
27 |
440 |
3674 |
239 |
754 |
- |
58 |
1000 |
f |
308 |
638 |
23 |
437 |
3773 |
219 |
716 |
- |
62 |
4000 |
f |
282++ |
682++ |
25 |
470 |
4795++ |
208 |
781 |
- |
69 |
+ difference against control for p<0.05
++ difference against control for p<0.01
Table 14: Number of rats with remarkable non-neoplastic lesions – interim kill
Sex |
|
m |
m |
m |
m |
|
f |
f |
f |
f |
f |
Dose |
(ppm) |
0 |
50 |
200 |
1000 |
3000 |
0 |
50 |
200 |
1000 |
4000 |
Organ/Finding |
Number Animals |
10 |
10 |
10 |
10 |
10 |
10 |
10 |
10 |
10 |
10 |
Adrenal Glands - increased vacuolation in zona fascicu.* |
§4 |
6 |
4 |
6 |
10 |
§2 |
0 |
0 |
0 |
9 |
|
Liver - hepatocellular hypertrophy (centilobul.) |
§0 |
0 |
0 |
6 |
10 |
§0 |
0 |
0 |
3 |
10 |
|
Urinary Bladder - transitional hyperplasia |
#0 |
0 |
2 |
2 |
3 |
§0 |
0 |
0 |
0 |
3 |
|
*= increased cytoplasmic vacuolisation in zona fasciculata
# trend (p < 0.05)
§ trend (p < 0.01)
Table 15: Number of rats with remarkable non-neoplastic lesions – terminal kill
Sex |
|
|
m |
m |
m |
m |
|
|
f |
f |
f |
f |
f |
Dose |
(ppm) |
|
0 |
50 |
200 |
1000 |
3000 |
|
0 |
50 |
200 |
1000 |
4000 |
Organ/Finding |
Number Animals |
|
50 |
50 |
50 |
50 |
50 |
|
50 |
50 |
50 |
50 |
50 |
Liver - eosinophilic foci |
# |
3 |
3 |
1 |
2 |
10 |
# |
|
|
3 |
3 |
3 |
|
- focal degenerative changes (periportal) |
§ |
|
|
|
2 |
13 |
|
|
|
|
|
1 |
|
- clear cell foci |
|
36 |
34 |
31 |
35 |
28 |
|
6 |
9 |
6 |
9 |
1 |
|
- cytoplasmic changes (periportal) |
§ |
|
|
|
2 |
22 |
|
|
|
|
|
|
|
- cytoplasmic changes (centrilobular) |
§ |
|
|
1 |
|
14 |
§ |
1 |
3 |
3 |
8 |
38 |
|
- hepatocellular hypertrophy (centilobul.) |
|
1 |
3 |
|
1 |
4 |
# |
|
|
1 |
3 |
40 |
|
- peliotic foci |
|
|
|
|
|
|
§ |
|
|
|
|
3 |
|
- single cell necrosis |
|
|
|
|
|
|
§ |
4 |
7 |
5 |
5 |
16 |
|
Urinary Bladder - transitional hyperplasia (focal) |
§ |
|
|
|
|
4 |
|
1 |
|
|
|
3 |
|
- simple transitional hyperplasia (diffuse) |
§ |
|
|
3 |
|
11 |
§ |
|
|
|
|
20 |
|
Skeletal Muscle - atrophy |
# |
7 |
2 |
9 |
10 |
11 |
§ |
|
|
1 |
2 |
9 |
|
- fibre degeneration (thigh) |
|
4 |
5 |
6 |
3 |
6 |
# |
2 |
|
1 |
|
7 |
|
Sciatic Nerve - degenerative myelinopathy |
§ |
21 |
25 |
27 |
32 |
43 |
§ |
17 |
20 |
13 |
23 |
38 |
|
Thyroid Gland - follicular hyperplasia |
§ |
1 |
1 |
2 |
3 |
7 |
|
2 |
|
|
2 |
1 |
|
- mineralization of follicular colloid |
# |
33 |
28 |
31 |
35 |
39 |
§ |
13 |
13 |
17 |
17 |
43 |
|
Adrenal Glands - increased vacuolation in zona fascicu.* |
§ |
5 |
7 |
7 |
3 |
20 |
|
1 |
2 |
2 |
3 |
4 |
|
- degenerative changes in the cortex |
|
5 |
3 |
3 |
3 |
3 |
|
19 |
14 |
11 |
8 |
4 |
|
Pituitary Gland (pars distalis) - hyperplasia (focal) |
|
10 |
6 |
4 |
13 |
10 |
|
18 |
13 |
9 |
11 |
9 |
|
Stomach (glandular mucosa) - dilated glands |
|
6 |
4 |
8 |
7 |
6 |
# |
7 |
7 |
6 |
8 |
16 |
|
Tongue - necrotizing arteritis |
|
7 |
12 |
7 |
11 |
9 |
|
7 |
7 |
4 |
2 |
0 |
|
Heart - cardiomyopathy |
|
43 |
45 |
48 |
44 |
48 |
|
37 |
37 |
33 |
28 |
22 |
|
Kidneys - chronic progressive nephropathy |
|
47 |
47 |
48 |
44 |
48 |
|
41 |
32 |
36 |
26 |
21 |
|
- diffuse urothelial hyperplasia |
|
10 |
18 |
16 |
8 |
15 |
|
35 |
35 |
30 |
21 |
19 |
|
Spleen - pigment storage |
|
19 |
24 |
31 |
20 |
27 |
§ |
9 |
9 |
11 |
15 |
23 |
|
# trend (p < 0.05)
§ trend (p < 0.01)
Table 16: Tumor incidences – Number with neoplastic lesions – terminal kill
Sex |
|
Males |
Females |
||||||||
Dose |
(ppm) |
0 |
50 |
200 |
1000 |
3000 |
0 |
50 |
200 |
1000 |
4000 |
Organ/Findings |
No.animals |
50 |
50 |
50 |
50 |
50 |
50 |
50 |
50 |
50 |
50 |
Liver |
No. exam. |
50 |
49 |
50 |
50 |
50 |
50 |
49 |
50 |
50 |
50 |
- Hepatocellular Adenoma |
|
1 |
|
1 |
|
|
|
1 |
|
|
1 |
- Hcpatocellular Adcnocarcinoma |
|
1 |
|
|
|
|
|
|
|
|
|
-Cholangioma |
|
|
|
|
|
|
|
|
|
1 |
|
- Cholangiocarcinoma |
|
|
1 |
|
|
|
|
|
|
|
|
Heart |
No. exam. |
50 |
49 |
50 |
50 |
50 |
50 |
49 |
50 |
50 |
50 |
- Endocardial Tumor (mal.) |
|
2 |
1 |
|
|
|
|
1 |
|
|
|
Kidneys |
No. exam. |
50 |
49 |
50 |
50 |
50 |
50 |
49 |
50 |
50 |
50 |
-Adenoma |
|
|
|
|
1 |
|
|
|
|
|
|
- Lipoma |
|
|
|
|
|
1 |
|
|
|
|
|
- Liposarcoma |
|
|
|
1 |
|
|
|
|
|
|
|
Urinary Bladder |
No. exam. |
50 |
48 |
50 |
50 |
50 |
50 |
49 |
48 |
49 |
49 |
- Transitional Cell Papilloma |
|
|
|
|
|
|
§ |
|
|
|
2 |
- Transitional Cell Carcinoma |
|
|
|
|
|
|
|
|
|
|
1 |
Urethra (residual)&) |
No. exam. |
|
|
|
|
1 |
|
|
|
|
|
- Transitional Cell Carcinoma |
|
|
|
|
|
1 |
|
|
|
|
|
LungsNo. Exam. |
No. exam. |
50 |
49 |
50 |
50 |
50 |
50 |
49 |
50 |
50 |
50 |
- AlvcolarffironchiolarAdenoma
|
|
1 |
|
|
|
|
|
|
|
|
|
- Alveolar/UronchiolarCarcinoma
|
|
|
1 |
1 |
|
|
|
|
|
|
|
Metastasis primary siteunknown |
|
|
1 |
|
|
|
|
|
1 |
|
|
Stomach No. Exam. |
No. exam. |
50 |
49 |
50 |
50 |
50 |
50 |
49 |
50 |
50 |
50 |
- Forestomach Papilloma |
|
|
|
|
|
|
|
|
|
|
1 |
Colon |
No. exam. |
50 |
48 |
50 |
50 |
50 |
50 |
49 |
49 |
49 |
49 |
- Fibrosarcoma |
|
|
|
|
|
|
|
1 |
|
|
|
Intestine (residual)&) |
No. exam. |
|
|
|
|
|
|
|
1 |
2 |
2 |
-Fibroma |
|
|
|
|
|
|
|
|
|
1 |
|
Pancreas |
No. exam. |
48 |
49 |
50 |
50 |
50 |
50 |
49 |
50 |
50 |
50 |
- Islet Cell Adenoma |
|
1 |
1 |
2 |
3 |
|
1 |
|
|
|
|
- Acinar Cell Adenoma |
|
|
|
|
|
1 |
|
|
|
1 |
|
Thymus |
No. exam. |
49 |
49 |
50 |
50 |
50 |
48 |
49 |
50 |
50 |
48 |
- Thymoma (benigne) |
|
1 |
1 |
|
1 |
|
|
I |
2 |
|
1 |
- Thymoma (maligne) |
|
1 |
|
|
|
|
|
|
|
|
|
Mesent. Lymph Node Mesent. Lymph NodeNo. Exam. |
No. exam. |
49 449 |
49 5049 |
50 50 |
50 50 |
50 50 |
50 50 |
49 49 |
50 50 |
49 49 |
50 50 |
- Hemangioma |
|
1 |
1 |
1 |
|
1 |
|
|
|
|
|
Spleen |
No. exam. |
50 |
49 |
50 |
50 |
50 |
50 |
49 |
50 |
49 |
50 |
- Hemangiosarcoma |
|
1 |
|
|
|
|
|
|
|
|
|
Skin/Other&) |
No. exam. |
9 |
3 |
5 |
5 |
4 |
6 |
2 |
2 |
2 |
2 |
- Schwannoma (mal.) |
|
|
1 |
|
|
|
|
|
|
|
|
- Fibroma |
|
|
|
|
1 |
|
|
|
|
|
|
- Fibrosarcoma |
|
1 |
|
|
|
|
|
1 |
|
|
|
- Squamous Cell Carcinoma |
|
2 |
1 |
|
1 |
|
|
|
|
|
|
- Keratoacanthoma |
|
3 |
|
2 |
|
1 |
|
|
|
|
|
- Papilloma |
|
|
|
|
|
1 |
|
|
|
|
|
- Hair Follicle Tumor (ben.) |
|
|
|
1 |
|
|
|
|
|
|
|
-Lipoma |
|
|
|
|
|
|
1 |
|
|
|
|
- Basal Cell Carcinoma |
|
|
|
|
|
|
|
|
1 |
|
|
Mammary Gland/Region |
No. exam. |
- |
- |
- |
- |
- |
50 |
49 |
50 |
50 |
50 |
-Adenoma |
|
|
|
|
|
|
|
1 |
|
|
|
- Adenocarcinoma |
|
|
|
|
|
|
3 |
1 |
6 |
|
2 |
- Adenocarcinoma arising in a fibroadenoma |
|
|
|
|
|
|
1 |
|
|
|
1 |
- Fihroadenoma |
|
|
|
|
|
|
15 |
10 |
7 |
6 |
5 |
- Tumor Mixed (mal.) |
|
|
|
|
|
|
1 |
|
|
1 |
1 |
Brain |
No. exam. |
50 |
49 |
50 |
50 |
50 |
50 |
49 |
50 |
50 |
50 |
- Astrocytoma (ben.) |
|
1 |
|
|
|
|
|
|
|
|
|
- Granular Cell Tumor (ben.) |
|
|
|
|
|
1 |
|
1 |
|
|
|
Testes |
No. exam. |
50 |
49 |
50 |
50 |
50 |
- |
- |
- |
- |
- |
- Leydig Cell Adenoma |
|
1 |
4 |
|
1 |
2 |
|
|
|
|
|
Prostate |
No. exam. |
50 |
49 |
50 |
50 |
50 |
- |
- |
- |
- |
- |
- Fibrosarcoma |
|
|
|
|
1 |
|
|
|
|
|
|
Ovaries |
No. exam. |
- |
- |
- |
- |
- |
50 |
49 |
50 |
50 |
50 |
- Granulosa Cell Tumor (ben.) |
|
|
|
|
|
|
1 |
|
|
|
|
- Granulosa Cell Tumor (mal.) |
|
|
|
|
|
|
|
|
|
|
1 |
- Tumor/Sex Cord (ben.) |
|
|
|
|
|
|
|
1 |
1 |
1 |
|
Uterus Uterus |
No. exam. No. Exam. |
|
|
|
|
|
50 |
49 550 |
50 49 |
50 50 |
50 50 |
-Adenoma |
|
|
|
|
|
|
|
|
|
1 |
|
- Adenocarcinoma |
|
|
|
|
|
|
§ 2 |
2 |
3 |
3 |
8 |
- Adenosquamous Carcinoma |
|
|
|
|
|
|
1 |
1 |
1 |
|
|
- Squamous Cell Carcinoma |
|
|
|
|
|
|
|
|
|
|
1 |
- Schwannoma (mal.) |
|
|
|
|
|
|
1 |
|
1 |
|
|
- Stromal Sarcoma |
|
|
|
|
|
|
|
1 |
|
1 |
1 |
- Stromal Polyp |
|
|
|
|
|
|
7 |
7 |
4 |
5 |
6 |
- Glandular Polyp |
|
|
|
|
|
|
1 |
|
3 |
|
|
Vagina |
No. exam. |
- |
- |
- |
- |
- |
50 |
49 |
50 |
50 |
50 |
- Granular Cell Tumor (ben.) |
|
|
|
|
|
|
|
|
|
|
1 |
Adrenal Glands |
No. exam. |
50 |
49 |
50 |
50 |
50 |
50 |
49 |
50 |
50 |
50 |
- Cortical Adenoma |
|
|
|
1 |
|
1 |
|
1 |
|
|
|
- Cortical Adenocarcinoma |
|
|
|
|
|
|
1 |
|
|
|
|
- Medullary Tumor (ben.) |
|
10 |
9 |
5 |
8 |
10 |
1 |
3 |
1 |
1 |
|
- Medullary Tumor (mal.) |
|
2 |
1 |
1 |
1 |
|
|
|
1 |
|
|
Pituitary Gland |
No. exam. |
49 |
49 |
50 |
50 |
50 |
50 |
49 |
50 |
50 |
50 |
- Adenoma (Pars Distalis) |
|
5 |
6 |
11 |
5 |
8 |
24 |
18 |
24 |
14 |
11 |
- Adenoma (Pars Intermedia) |
|
|
|
|
|
|
1 |
|
|
|
1 |
- Adenocarcinoma (Pars Distalis) |
|
|
|
|
|
|
1 |
|
1 |
|
|
Thyroid Gland |
No. exam. |
50 |
49 |
50 |
50 |
50 |
50 |
49 |
50 |
50 |
50 |
- C-Cell Adenoma |
|
3 |
7 |
2 |
2 |
3 |
3 |
2 |
3 |
4 |
3 |
- C-Cell Adenocarcinoma |
|
1 |
|
|
|
|
|
|
|
|
|
- Follicular Adenoma |
|
# |
|
|
1 |
2 |
1 |
1 |
1 |
1 |
1 |
- Follicular Adenocarcinoma |
|
|
|
|
|
1 |
|
|
|
|
|
Bone/NOS&) |
|
|
|
|
|
|
|
|
1 |
1 |
|
- Osteosarcoma |
|
|
|
|
|
|
|
|
1 |
|
|
- Schwannoma (maligne) |
|
|
|
|
|
|
|
|
|
1 |
|
Systemic Tumors |
No. exam. |
50 |
49 |
50 |
50 |
50 |
50 |
49 |
50 |
50 |
50 |
- Lymphoma (maligne) |
|
2 |
1 |
|
2 |
|
|
|
1 |
1 |
|
- Myeloid Leukemia |
|
|
1 |
|
1 |
|
|
|
|
|
|
- Histiocytic Sarcoma |
|
2 |
1 |
|
1 |
1 |
|
|
|
|
|
* in case of two tumors in a paired organ one tumor was taken into consideration
&) only investigated in case of macro lesions
# positive trend p<0.05
§ positive trend p<0.01
NOS = not otherwise specified
no. = number
exam. = examinations
Table 17: Number of Animals with neoplastic lesions – terminal kill
Sex |
|
male |
female |
||||||||
Dose ppm |
|
0 |
50 |
200 |
1000 |
3000 |
0 |
50 |
200 |
1000 |
4000 |
Organ/Finding |
No. animals |
50 |
50 |
50 |
50 |
50 |
50 |
50 |
50 |
50 |
50 |
Body cavities &) |
No. exam. |
5 |
3 |
6 |
6 |
3 |
3 |
3 |
3 |
4 |
6 |
- Mesothelioma (benign) |
|
1 |
|
|
|
|
|
|
|
|
|
- Lipoma |
|
|
|
1 |
|
|
1 |
|
|
1 |
|
- Fibroma |
|
|
|
|
|
|
1 |
|
|
|
|
Clitorial glands &) |
No. exam. |
|
|
|
|
|
|
1 |
1 |
|
|
- Squamous cell carcinoma |
|
|
|
|
|
|
|
|
1 |
|
|
Number of benign tumors |
|
29 |
29 |
27 |
23 |
32 |
58 |
47 |
47 |
36 |
33 |
Malignant tumors |
|
15 |
10 |
3 |
7 |
3 |
11 |
8 |
18 |
7 |
16 |
Total tumors |
|
44 |
39 |
30 |
30 |
35 |
69 |
55 |
65 |
43 |
49 |
* in case of two tumors in a paired organ one tumor was taken into consideration
&) only investigated in case of macro lesions
# positive trend p<0.05
§ positive trend p<0.01
NOS = not otherwise specified
no. = number
exam. = examinations
Table 18: Animals with tumors – terminal kill
Sex |
Males |
Females |
||||||||
Dose ppm |
0 |
50 |
200 |
1000 |
3000 |
0 |
50 |
200 |
1000 |
4000 |
|
lntercurrent Deaths |
|||||||||
No. of animals examined |
21 |
19 |
21 |
17 |
13 |
12 |
20 |
24 |
20 |
22 |
Animals with tumors |
12 |
11 |
13 |
7 |
5 |
10 |
13 |
19 |
12 |
15 |
Animals with benign tumors only |
5 |
7 |
10 |
3 |
4 |
8 |
7 |
9 |
7 |
5 |
Animals with malignant tumors only |
5 |
5 |
3 |
2 |
|
1 |
4 |
5 |
2 |
3 |
Animals with benign and malignant tumors |
2 |
|
|
2 |
1 |
1 |
2 |
5 |
3 |
7 |
|
Terminal Kill |
|||||||||
No. of animals examined |
29 |
31 |
29 |
33 |
37 |
38 |
30 |
26 |
30 |
28 |
Animals with tumors |
17 |
16 |
13 |
16 |
21 |
31 |
20 |
22 |
16 |
15 |
Animals with benign tumors only |
10 |
11 |
13 |
13 |
19 |
25 |
19 |
15 |
14 |
10 |
Animals with malignant tumors only |
2 |
2 |
|
2 |
1 |
1 |
|
|
|
1 |
Animals with benign and malignant tumors |
5 |
3 |
|
1 |
1 |
5 |
1 |
7 |
2 |
4 |
|
All Animals |
|||||||||
No. of animals examined |
50 |
50 |
50 |
50 |
50 |
50 |
50 |
50 |
50 |
50 |
Animals with tumors |
29 |
27 |
26 |
23 |
26 |
41 |
33 |
41 |
28 |
30 |
Animals with benign tumors only |
15 |
18 |
23 |
16 |
23 |
33 |
26 |
24 |
21 |
15 |
Animals with malignant tumors only |
7 |
7 |
3 |
4 |
1 |
2 |
4 |
5 |
2 |
4 |
Animals with benign and malignant tumors |
7 |
3 |
|
3 |
2 |
6 |
3 |
12 |
5 |
11 |
Table 19: Occurrence of tumor bearing animals in time
Sex |
|
Males |
Females |
|||||||||
Dose ppm |
|
0 |
50 |
200 |
1000 |
3000 |
0 |
50 |
200 |
1000 |
4000 |
|
Number of Animals with Neoplasms |
|
|
|
|
|
|
|
|
|
|
|
|
Dying Prematurely |
|
12 |
11 |
13 |
7 |
5 |
10 |
13 |
19 |
12 |
15 |
|
Neoplasms |
benign |
11 |
9 |
13 |
7 |
7 |
13 |
11 |
17 |
11 |
14 |
|
|
maligne |
7 |
5 |
3 |
4 |
1 |
3 |
7 |
9 |
5 |
11 |
|
|
total |
18 |
14 |
16 |
11 |
8 |
16 |
18 |
26 |
16 |
25 |
|
Number of Animals with Tumors |
||||||||||||
Week |
0-13 |
|
|
1 |
|
|
|
|
|
|
|
|
|
13-26 |
|
|
|
|
|
|
|
|
|
|
|
|
27-39 |
|
|
|
|
|
|
|
|
|
|
|
|
40-52 |
|
|
|
|
1 |
|
|
1 |
1 |
|
|
|
53-65 |
|
1 |
2 |
|
|
|
|
|
|
2 |
1 |
|
66-78 |
|
1 |
|
1 |
1 |
|
1 |
2 |
4 |
1 |
2 |
|
79-91 |
|
4 |
2 |
6 |
2 |
2 |
3 |
3 |
8 |
4 |
7 |
|
92-Necropsy |
|
6 |
7 |
6 |
3 |
3 |
6 |
7 |
6 |
5 |
5 |
Applicant's summary and conclusion
- Conclusions:
- CLP: STOT RE 2, H372 Liver
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