Registration Dossier

Administrative data

Endpoint:
short-term repeated dose toxicity: oral
Type of information:
migrated information: read-across based on grouping of substances (category approach)
Adequacy of study:
key study
Study period:
1994
Reliability:
1 (reliable without restriction)
Rationale for reliability incl. deficiencies:
other: GLP, guideline study (Note- The exposure period was only 28 days.)
Cross-referenceopen allclose all
Reason / purpose:
reference to same study
Reason / purpose:
reference to other study

Data source

Reference
Reference Type:
study report
Title:
Unnamed
Year:
1994

Materials and methods

Test guideline
Qualifier:
according to
Guideline:
other: Japanese Ministry of Health and Welfare Guidelines (1986)
Deviations:
no
Remarks:
Not specified in report
Principles of method if other than guideline:
Not applicable
GLP compliance:
yes
Limit test:
no

Test material

Reference
Name:
Unnamed
Type:
Constituent
Details on test material:
Test Material: The test material was prepared at the appropriate concentrations as a solution in Arachis oil BP. The test material was stable in this vehicle for at least 10 days. Formulations were prepared weekly and stored at 4 degrees C in the dark. Samples of each formulation were taken for analysis of 1-nitropropane. The analytical results were within +/- 9% of the nominal concentrations. Purity >98.5%.

Test animals

Species:
rat
Strain:
Sprague-Dawley
Sex:
male/female
Details on test animals and environmental conditions:
Animals: Animals (male and female Sprague-Dawley CD rats) were acclimated for 8 days before use. A total of 30/sex were accepted into the study. At the beginning of the study, males weighed 121 - 161 g and females weighed 121-159 g, and were approximately 5-6 weeks old. The animals were allowed free access to food (except for the night prior to urine collection, when it was withdrawn) and water. The diet and drinking water did not contain any contaminants that might have influenced the study. Rats were maintained on a 12 hour light/dark cycle. Animals were randonly allocated to 6 groups (5/sex/group).

Administration / exposure

Route of administration:
oral: gavage
Vehicle:
arachis oil
Details on oral exposure:
The test material was administered daily, for up to twenty-eight consecutive days, by gavage using a stainless steel cannula attached to a disposable plastic syringe. Control animals were treated in an identical manner with 2 ml/kg/day of Arachis oil B.P. Animals from satellite groups 5 and 6 were maintained for a further fourteen days treatment-free period following termination of treatment. The volume of test and control material administered to each animal was based on the most recent bodyweight and was adjusted at weekly intervals.
Analytical verification of doses or concentrations:
yes
Details on analytical verification of doses or concentrations:
The stability and homogeneity of the test material formulations were determined by Safepharm Analytical Laboratory. Samples were taken of each test material formulation and were analysed for concentration of 1 -Nitropropane at Safepharm Analytical Laboratory.
Duration of treatment / exposure:
28 days, post exposure 14 days
Frequency of treatment:
daily
Doses / concentrations
Remarks:
Doses / Concentrations:0, 10, 30, 100 mg/kg/dayBasis:other: nominal
No. of animals per sex per dose:
5/sex/group
Control animals:
yes, concurrent vehicle
Details on study design:
Test conduct: Four groups of animals were dosed with 0 (control), 10, 30 or 100 mg/kg/day active material for 28 consecutive days by gavage, and then were terminated. Controls were dosed with 2 ml/kg/day Arachis oil BP. The two additional groups (satellite animals) were dosed with 0 or 100 mg/kg/day test material for 28 days and then were allowed to recover for 14 days before termination.
Positive control:
No

Examinations

Observations and examinations performed and frequency:
All animals were examined for signs of toxicity before dosing and 1 and 5 hours after dosing on weekdays and before dosing and 1 hour after treatment on weekends. During the treatment-free period, satellite animals were observed twice daily on weekdays and once daily on weekends.Body weights were recorded on day 0 and on days 7, 14, 21 and 28. Satellite animals also were weighed on days 35 and 42 (at termination). Food consumption was recorded for each cage (5 animals) at weekly intervals. Water intake was visually inspected. Clinical chemistry (urea, total protein, albumin, albumin/globulin ratio, sodium , potassium, chloride, calcium, inorganic phosphorus, alkaline phosphatase, alanine aminotransferase, aspartate aminotransferase, glucose, gamma glutamyl transpeptidase, triglycerides, total chloesterol, total bilirubin and creatinine) and hematological analyses (hematocrit, hemoglobin, erythrocyte count, total and differential leukocyte count, platelet count, mean corpuscular hemoglobin, mean corpuscular volume, mean corpuscular hemoglobin concentration, methemoglobin concentration, reticulocyte count and clotting time) were performed on blood collected from the lateral tail vein of all survivors from the control and high dose groups (both main study and satellite animals) at termination. Blood for hematological and blood chemistry analyses was collected into tubes containing potassium EDTA or lithium heparin, respectively (with the exception of blood for clotting time analysis, which was collected into sodium citrate). Animals were not fasted prior to blood collection. Urinalysis (volume, specific gravity, pH, protein, glucose, ketones, bilirubin, urobilinogen, reducing substances and blood) was performed on control and high dose main study animals during week 4 and from satellite animals during the final week of the study. Urine samples were collected over a period of approximately 16 hours, by housing the rats in metabolism cages. Animals did not have access to food during the collection period.
Sacrifice and pathology:
At termination, all surving animals were euthanized and were subjected to a full external and internal examination. The adrenals, brain, testes, ovaries, heart, kidneys, liver, pituitary and spleen were weighed. The organs, plus the aorta (thoracic), bone and bone marrow (femur and sternum), cecum, colon, duodenum, eyes, gross lesions, ileum, jejunum, lungs, lymph nodes (cervial and mesenteric), muscle (skeletal), esophagus, pancreas, prostate, rectum, salivary glands, sciatic nerve, seminal vesicles, skin (hind limb), stomach, thymus, thyroid/parathyroid, trachea, urinary bladder and uterus were fixed in 10% buffered formalin. Microscopic examinations were performed on tissues from control and high dose animals. All lesions from other groups were also examined microscopically.
Other examinations:
None
Statistics:
Statistical analyses: Absolute and relative organ weights, hematological, blood chemistry, weekly body weight gain and quantitative urinalysis data were analyzed by one was analysis of variance (ANOVA) incorporating the Fmax test for homogeneity of variance. Data with heterogenous variances were analyzed with the Kruskal Wallis non-parametric analysis of variance and the Mann Whitney U-Test. The critical level of significance was P < 0.05.

Results and discussion

Results of examinations

Clinical signs:
effects observed, treatment-related
Mortality:
mortality observed, treatment-related
Body weight and weight changes:
effects observed, treatment-related
Food consumption and compound intake (if feeding study):
effects observed, treatment-related
Food efficiency:
not examined
Water consumption and compound intake (if drinking water study):
no effects observed
Ophthalmological findings:
not examined
Haematological findings:
no effects observed
Clinical biochemistry findings:
no effects observed
Urinalysis findings:
no effects observed
Behaviour (functional findings):
no effects observed
Organ weight findings including organ / body weight ratios:
effects observed, treatment-related
Gross pathological findings:
effects observed, treatment-related
Histopathological findings: non-neoplastic:
no effects observed
Histopathological findings: neoplastic:
no effects observed
Details on results:
Effects at 100 mg/kg: One high dose male was killed in extremis on day 27. This animal had exhibited hunched posture, lethargy and ataxia on day 16. This animal exhibited dark kidneys, thickening of the forestomach and sloughing of the glandular gastric epithelium at necropsy. Animals showed signs of increased salivation one hour after dosing on day 15. Sporadic incidents were then noted approximately 5 minutes after dosing from day 16 on. This lasted for up to one hour after dosing. Red/brown staining around the mouth was apparent from day 15 on. Two females developed hunched posture on day 28. Body weight gain was reduced in main study males during weeks 2 and 4. This was not observed in females or satellite males. Food consumption of main study males was reduced by 13% during the last week of the study. A reduction in food intake was not observed in satellite males. Water consumption did not appear to be altered. Hemoglobin, hematocrit and erythrocyte counts were reduced and white blood cells (lymphocytes) and clotting time were increased in main study females (with respect to the study control). However, hemoglobin, hematocrit and erythrocyte counts were within the normal ranges for rats of the same strain and age. Mean corpuscular hemoglobin concentration was reduced in main study males. Methemoglobin and lymphocyte counts were increased in satellite males and hematocrit was decreased in satellite females. Plasma urea was increased in main study males with respect to study controls, but not to historical controls. The albumin/globulin ratio and triglycerides were elevated and alkaline phosphatase was less than control in main study females. Urine volume was increased and urine specific gravity was decreased in main study females. Absolute and relative brain weights were increased in main study animals, with only female relative weights not achieving statistical significance. Main study females also had increased absolute and relative kidney weights and decreased relative ovary weights. The absolute weight of the pituitary was decreased and the relative heart weight was increased in main study males with respect to the study control (but not historical controls). There was no pathological evidence of brain toxicity. The incidences of all morphological changes in the organs that were examined were similar to control. Effects at 30 mg/kg: There were no deaths or clinical signs of toxicity. Males had increased body weight gains during weeks 2 and 4. There was no effect of treatment on food consumption. Water consumption did not appear to be altered. Urine volume was increased and urine specific gravity was decreased in main study females. Changes in these parameters were dose-dependent. Absolute brain and liver weights were increased in main study females and males, respectively. Effects at 10 mg/kg: There were no deaths or clinical signs of toxicity. Body weight gains and food consumption were unaffected by treatment. Water consumption did not appear to be altered. There was an increase in methemoglobin in main study males. Alkaline phosphatase was less than control in main study females. Urine specific gravity was decreased in main study females. The absolute weights of the adrenals and heart were increased in main study females. Controls: A dark accessory lobe of the liver was found in one female at necropsy.

Effect levels

open allclose all
Dose descriptor:
NOEL
Effect level:
30 mg/kg bw/day (nominal)
Sex:
male/female
Basis for effect level:
clinical signs
mortality
body weight and weight gain
food consumption and compound intake
organ weights and organ / body weight ratios
gross pathology
Dose descriptor:
LOAEL
Effect level:
100 mg/kg bw/day (nominal)
Sex:
male/female
Basis for effect level:
clinical signs
mortality
body weight and weight gain
food consumption and compound intake
organ weights and organ / body weight ratios
gross pathology

Target system / organ toxicity

Critical effects observed:
not specified

Any other information on results incl. tables

The dose levels were chosen based on a 14-day study with 10, 50, 150 and 250 mg/kg test material.  All animals treated with 250 mg/kg and one animal treated with 150 mg/kg were killed in extremis. No effects were noted at 10 mg/kg.

A detailed justification for setting the NOAEL (actually a NOEL) at 30 mg/kg was provided by the investigators.
  Effects considered to be related to treatment with 100 mg/kg/day were hunched posture, lethargy, ataxia and death. Effects considered possibly related to treatment with 100 mg/kg/day were reduced body weight gain and increased brain weight.  The increase in the absolute brain weight of females treated with 30 mg/kg/day was not considered to be biologically significant since an increase in the relative weight was not observed.

Changes in clincial chemistries, hematologies, urinalysis and weights of organs other than the brain were not considered to be related to treatment since there were no corresponding morphologic changes and/or the changes were not dose-dependent.

Applicant's summary and conclusion

Conclusions:
Clinical findings in a few animals dosed with 100 mg/kg were consistent with toxicity to the nervous system. Organ weight data revealed significant increases in brain weights in high dose animals. However, there were no treatment-related morphologic changes in the brain. Kidney weights were increased in high dose animals and urinalyses suggested that females treated with 30 and 100 mg/kg produced increased amounts of dilute urine. However, blood chemistries and histopathology revealed no evidence of kidney toxicity. Therefore, there was "no convincing evidence of treatment-related renal effects." None of the other changes observed were considered to be related to treatment since they were not dose-dependent.Oral administration of the test material, 1-Nitropropane, to rats for a period of twenty-eight consecutive days at dose levels of up to 100 mg active ingredient/kg/day resulted in toxicologically significant effects at 100 mg active ingredient/kg/day. No such effects were detected at 30 or 10 mg active ingredient/kg/day and, the "No Observed Effect, Level " (NOEL) is, therefore, considered to be 30 mg active ingredient/kg/day.
Executive summary:

None