4-morpholinecarbaldehyde

Repeated dose toxicity: oral

Administrative data

Endpoint:

short-term repeated dose toxicity: oral

Type of information:

experimental study

Adequacy of study:

key study

Reliability:

1 (reliable without restriction)

Rationale for reliability incl. deficiencies:

other: GLP guideline study

Data source

Reference

Reference Type:

study report

Title:

Unnamed

Year:

1998

Report date:

1998

Materials and methods

Test guideline

Qualifier:

according to guideline

Guideline:

OECD Guideline 407 (Repeated Dose 28-Day Oral Toxicity Study in Rodents)

Deviations:

yes

Remarks:

the weight of prostate and seminal vesicles with coagulating glands as a whole was not determinded

GLP compliance:

yes

Limit test:

no

Test material

Test material information

Constituent 1

Reference substance name:

4-morpholinecarbaldehyde

EC Number:

224-518-3

EC Name:

4-morpholinecarbaldehyde

Cas Number:

4394-85-8

Molecular formula:

C5H9NO2

IUPAC Name:

morpholine-4-carbaldehyde

Details on test material:

Name of test material (as cited in study report): 4-morpholinecarbaldehyde

Test animals

Species:

rat

Strain:

Wistar

Sex:

male/female

Details on test animals or test system and environmental conditions:

TEST ANIMALS
- Source: Dr. Karl Thomae GmbH
- Age at study initiation: 49 days old
- Weight at study initiation: males mean ca. 236 g; females mean ca. 172 g
- Housing: individually in type DK III stainless steel wire mesh cages
- Diet: ad libitum, ground Kliba maintenance diet rat/mouse/hamster
- Water: ad libitum from water bottles
- Acclimation period: 8 (males) or 9 (females) days


ENVIRONMENTAL CONDITIONS
- Temperature: 20-24 °C
- Humidity: 30-70 %
- Photoperiod (hrs dark / hrs light): 12/12

Administration / exposure

Route of administration:

oral: gavage

Vehicle:

water

Remarks:

doubly distilled

Details on oral exposure:

PREPARATION OF DOSING SOLUTIONS:
The test substance was administered as a solution in doubly distilled water. The test substance was weighed in depending on the dose group, then doubly distilled water was filled up to the desired volume. The solutions were mixed by shaking. The mixtures were prepared at least once a week.

VEHICLE
- Concentration in vehicle: 0.5, 2.0 and 10.0 g/100 mL
- Amount of vehicle: 10 ml/kg bw

Analytical verification of doses or concentrations:

yes

Details on analytical verification of doses or concentrations:

The analyses were carried out at the Analytical Department of BASF Aktiengesellschaft. The stability of the test substance in water
was tested over a period of 10 days at room temperature. Concentration analyses were performed in samples of all concentrations at the start of the administration period. As the test substance was administered as a solution, no homogeneity analyses were conducted.

Duration of treatment / exposure:

28 days

Frequency of treatment:

once daily

Doses / concentrations

Remarks:

Doses / Concentrations:
0, 50, 200 and 1000 mg/kg bw/d
Basis:

No. of animals per sex per dose:

5 animals per sex per dose plus another 5 animals per sex in the control and high dose groups (recovery)

Control animals:

yes, concurrent vehicle

Details on study design:

- Dose selection rationale:
In a preceding test study, N-formylmorpholin was administered to groups of 3 male and female Wistar rats by gavage at dose levels of 0, 300, or 1000 mg/kg bw/d for 2 weeks. No substancerelated findings were observed. On this basis, the dose levels (0, 50, 200 and 1000 mg/kg bw/d) were selected for the present study.

- Rationale for selecting satellite groups:
Animals of the highest dose and control group underwent a recovery period of two weeks in order to prove reversibility and/or delayed onset of possible effects.

- Post-exposure recovery period in satellite groups:
2 weeks

Positive control:

none

Examinations

Observations and examinations performed and frequency:

CAGE SIDE OBSERVATIONS / DETAILED CLINICAL OBSERVATIONS:
- The animals were examined for evident signs of toxicity or mortality twice a day (in the morning and in the late afternoon) from Mondays to Fridays and once a day (in the morning) on Saturdays, Sundays and public holidays. Additionally, further clinical examinations were carried out daily just before treatment, less than 1 hour after treatment, between 3 and 4 hours after treatment, as well as once a day during the recovery period.

BODY WEIGHT:
- Time schedule for examinations: The body weight was determined before the start of the administration period in order to randomize the animals. During the administration period and the recovery period the body weight was determined on day 0 (start of administration period) and thereafter at weekly intervals. The difference between the body weight on the respective day of weighing and the body weight on day 0 was calculated as body weight change.

FOOD CONSUMPTION:
- Food consumption was determined weekly over a period of 7 days and calculated as mean food consumption in grams per animal and day.

FOOD EFFICIENCY:
- body weight gain in g/food consumption in g/per unit time x 100 calculated as time-weighted averages

WATER CONSUMPTION:
- Water consumption was observed daily by visual inspection of the water bottles for any overt changes in volume.

HAEMATOLOGY:
- Time schedule for collection of blood: day 24 (males) and 26 (females) for all animals and day 40 (females, recovery) and 41 (males, recovery)
- Anaesthetic used for blood collection: no
- Animals fasted: no
- How many animals: all animals
- Parameters examined: leukocytes, erythrocytes, hemoglobin, hematocrit, mean corpuscular volume, mean corpuscular hemoglobin, mean corpuscular hemoglobin concentration, platelets, differential blood count, prothrombin time (Hepato Quick's test)


CLINICAL CHEMISTRY:
- Time schedule for collection of blood: day 24 (males) and 26 (females) for all animals and day 40 (females, recovery) and 41 (males, recovery)
- Animals fasted: yes. withdrawal of food and water during the collection period
- How many animals: all animals
- Parameters examined: alanine aminotransferase, aspartate aminotransferase, alkaline phosphatase, serum-y-glutamyltransferase, sodium, potassium, chloride, inorganic phosphate, calcium, urea, creatinine, glucose, total bilirubin, total protein, albumin, globulins, triglycerides, cholesterol, magnesium

URINALYSIS:
- Time schedule for collection of urine: day 23 (all animals) and day 36 (females, recovery) and 37 (males, recovery)
- Metabolism cages used for collection of urine: yes
- Animals fasted: no
- Parameters examined: volume, color, turbidity, nitrite, pH, protein, glucose, ketones, urobilinogen, bilirubin, blood, specific gravity, sediment

NEUROBEHAVIOURAL EXAMINATION:
- Time schedule for examinations: Detailed clinical examinations (open field observations) were carried out in all animals prior to the test substance administration (day -1), and on day 7, 14, 21, and in all animals of recovery groups on day 35, each time from about 2.00 p.m. onwards.
Functional observational batteries and motor activity measurements were carried out in all animals on the day of the last administration (day 28), and in all animals of recovery groups on day 42. Functional observational batteries were performed each time from about 2.00 p.m . and motor activity measurements were conducted each time from about 11.00 a.m . (main groups) and 12.30 p.m. (recovery groups) onwards.
- Dose groups that were examined: all animals
- Battery of functions tested: sensory activity / grip strength / motor activity

Sacrifice and pathology:

GROSS PATHOLOGY: Yes as well as organ weights

HISTOPATHOLOGY: Yes: all gross lesions, brain, pituitary gland, thyroid glands with parathyroid glands, thymus, trachea, lungs, heart, liver, spleen, kidneys, adrenal glands, testes/ovaries, uterus/vagina, accessory genital organs (epididymides, prostate, seminal vesicle), stomach (glandular and non-glandular), duodenum, jejunum, ileum, cecum, colon, rectum, urinary bladder, mandibular and mesenteric lymph nodes, sciatic nerve, bone marrow (femur), eyes, spinal cord (cervical, thoracic and lumbar cord)

Other examinations:

Organ weights:
The weight of the anesthetized animals as well as the weights of liver, kidneys, adrenal glands, testes, epididymides, ovaries, brain, thymus, heart, and spleen from all animals sacrificed at scheduled dates were determined .

Statistics:

Means and standard deviations of each test group were calculated for several parameters.
Further statistical analyses were performed but are too large in number to be described here.

Results and discussion

Results of examinations

Clinical signs:

no effects observed

Description (incidence and severity):

No animal died during the study period and no substance-related findings were obtained.

Mortality:

no mortality observed

Description (incidence):

No animal died during the study period and no substance-related findings were obtained.

Body weight and weight changes:

effects observed, non-treatment-related

Description (incidence and severity):

No statistically significant or biologically relevant deviations of body weights were observed. Concerning body weight change data, statistically significantly increased values were seen in males of the high dose recovery group on days 21 (+10.8%), day 28 (+6.3%) and day 42 (+10.1%). These slight deviations were most probably due to the (incidentally) increased food consumption in these animals, but not related to treatment.

Food consumption and compound intake (if feeding study):

effects observed, non-treatment-related

Description (incidence and severity):

Food consumption was statistically significantly increased in high dose males (recovery group) on day 21 (+10.1%) and in high dose females (recovery group) on day 35 (+15.0%). Due to either the single occurrence or the fact that the effect occurred during even in the recovery period, this was assessed as being incidental and not related to treatment.

Food efficiency:

effects observed, treatment-related

Description (incidence and severity):

Food efficiency was statistically significantly decreased in high dose males (main group) on day 28 and statistically significantly increased in high dose females (main group) on day 21. Both findings were assessed as being incidental in nature.

Haematological findings:

effects observed, non-treatment-related

Description (incidence and severity):

At the end of the study significant decreases in white blood cell counts were observed in the peripheral blood of all treated males of the main groups. These findings were not considered to be test substance-related since there was no dose-response relationship and all leukocyte values of the treated animals were within the limits of historical control data. Moreover, statistically significantly decreased white blood cells were observed in the males only. It is very likely that the statistically significant decreases in leukocytes were only due to the high value of the control group. Thus, the lowered white blood cell counts were regarded to be incidental in nature and were not related to treatment. The other hematological and clotting examinations revealed no treatment-related changes.

Clinical biochemistry findings:

effects observed, non-treatment-related

Description (incidence and severity):

At the end of the administration period slightly increased alanine aminotransferase activities were found in the sera of the high dose males and females of the main and recovery groups. After cessation of the test compound administration these findings returned to normal. There are no treatment-related changes in the other clinical chemistry parameters measured.

Urinalysis findings:

effects observed, non-treatment-related

Description (incidence and severity):

There were no treatment-related changes in the urinalytical parameters measured.

Behaviour (functional findings):

effects observed, non-treatment-related

Description (incidence and severity):

Detailed clinical observations (open field observations): No substance-related effects were observed.
Functional observational battery: No substance-related effects were observed
Home cage observations: No substance-related effects were observed.

Open field observations : No abnormalities were obtained in any of the main test groups when observing the animals in the open field. Rearing was statistically significantly decreased in the high dose males (recovery group) on day 28. As no such effect was seen in the main group, this was assessed as being incidental.
Sensorimotor tests/Reflexes: All findings were within the biological range of variation. No treatment-related effect was observed.
Quantitative observations (Grip strength, Landing foot-splay test): Grip strength hindlimbs was statistically significantly decreased in low dose females on day 28. Due to the lack of a dose-response relationship, this was assessed as being incidental. All other findings were within the biological range of variation. No treatment-related effect was observed.
Motor activity measurement: Regarding the overall motor activity no statistically significant or biologically relevant deviation was seen in any of the treatment groups. The comparison of the single intervals with the control group resulted in a statistically significantly higher value in high dose females (recovery group) on day 28 (interval 11). This was, however clearly incidental in nature and not related to treatment.

Organ weight findings including organ / body weight ratios:

effects observed, non-treatment-related

Description (incidence and severity):

The mean absolute weight of the spleen was significantly decreased in males of the low and high dose groups, whereas it was significantly increased in females of the low dose group. The other mean absolute weight parameters of the animals of the main groups did not show significant differences when compared with the concurrent control group. Due to a slightly decreased mean terminal body weight, the mean weight of the kidneys was significantly increased in males of the high dose group. This was not regarded to be treatment related. The mean weight of the spleen was significantly increased in females of the low dose group. The other mean relative weight parameters (when related to terminal body weight) of the animals of the main groups did not show significant differences when compared with the concurrent control group.

Recovery groups:
Due to a slight increase of the mean terminal body weight (+ 2.6%), the mean weight of the liver was also slightly (+ 9.2%) significantly increased in males of the high dose recovery group. This was not regarded to be treatment related. The other mean absolute weight parameters of the animals of the recovery groups did not show significant differences when compared with the concurrent control group. The mean relative weight parameters (when related to terminal body weight) of the animals of the recovery groups did not show significant differences when compared with the concurrent control group.

Gross pathological findings:

effects observed, treatment-related

Description (incidence and severity):

The only gross lesions noted were in the glandular stomach of a low dose male (erosion/ulcer) and in the kidneys of a low dose female (pelvic dilation).

Histopathological findings: non-neoplastic:

no effects observed

Description (incidence and severity):

There were no remarkable microscopic findings noted in any of the organs investigated. All microscopic findings recorded were either single observations, or they occurred in control animals only, or they were recorded at comparable incidence and graded severity in control and high dose males and/or females. There was no indication of a morphologic affection of the organs of the central or peripheral nervous system, the reproductive system, and the immune system.

Recovery groups:
Histopathology correlated the gross lesions in the glandular stomach with an incidental focus of cystic malformation. The focus in the liver turned out as tension lipidosis, a spontaneous observation caused by the traction of the ligamentum hepato-colicum at its insertion site on the surface of the liver. The few other microscopic findings noted in the liver were also of spontaneous origin and unrelated to treatment.

Description (incidence and severity):

There were additional statistically significant intergroup differences in the results of clinical pathology testing. These deviations were marginal, incidental or inconsistent, when compared with the other sex, or lacked a dose-response relationship. Accordingly, these findings were considered to be of no toxicological significance.

Effect levels

Dose descriptor:

NOAEL

Effect level:

1 000 mg/kg bw/day (nominal)

Sex:

male/female

Basis for effect level:

other: see 'Remark'

Target system / organ toxicity

Critical effects observed:

not specified

Any other information on results incl. tables

Chemical analyses of dose formulations revealed that the test item was stable in the vehicle over a period of 10 days when kept at room temperature. The mean analytical concentration values were 100 -105 % of the nominal concentraions.

Applicant's summary and conclusion

Executive summary:

The oral administration of the test item by gavage to rats at the high dose level of 1000 mg/kg bw/day for 4 weeks resulted in slight increases in alanine aminotransferase activity in the sera indicating mild hepatocellular damage. No test substance-related changes were seen in the low and mid dose animals (50 and 200 mg/kg bw/day). The increases in liver enzyme activities in the sera observed at the end of the administration period were reversible during the course of the treatment-free recovery period. The no observed adverse effect level (NOAEL) under the conditions of this study was 1000 mg/kg bw/day.