4-(1,4-dioxa-spiro[4.5]dec-8-yl)-cyclohexanone

Endpoint summary

Description of key information

An oral 28 d toxicity study according to OECD TG 407 in the rat, revealed no adverse efffects and a NOAEL of 1000 mg/kg bw/d (highest dose applied) was derived (reference 7.5.1 -1).

Key value for chemical safety assessment

Toxic effect type:

dose-dependent

Repeated dose toxicity: via oral route - systemic effects

Link to relevant study records

Reference

Endpoint:

short-term repeated dose toxicity: oral

Type of information:

experimental study

Adequacy of study:

supporting study

Study period:

07 January - 12 May 1990

Reliability:

1 (reliable without restriction)

Rationale for reliability incl. deficiencies:

guideline study

Qualifier:

according to guideline

Guideline:

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

Version / remarks:

1995

Deviations:

no

Qualifier:

according to guideline

Guideline:

EU Method B.7 (Repeated Dose (28 Days) Toxicity (Oral))

Version / remarks:

1996

GLP compliance:

yes

Limit test:

no

Species:

rat

Strain:

Wistar

Remarks:

Crl:(WI) BR

Sex:

male/female

Details on test animals or test system and environmental conditions:

TEST ANIMALS
- Source: Charles River, Sulzfeld, Germany
- Females nulliparous and non-pregnant: yes
- Age at study initiation: approx. 6 weeks
- Weight at study initiation: males 195 - 228 g, females 158 - 184 g
- Fasting period before study: not specified
- Housing: 5 animals/cage, individual housing during activity monitoring
- Diet: ad libitum
- Water: ad libitum
- Acclimation period: at least 5 days

DETAILS OF FOOD AND WATER QUALITY: diet batches were analysed for nutrients and contaminants on a regular basis, for water certificates of analysis were performed quarterly

ENVIRONMENTAL CONDITIONS
- Temperature (°C): 21°C
- Humidity (%): 50%
- Air changes (per hr): 15
- Photoperiod (hrs dark / hrs light): 12 hours light / 12 hours dark

IN-LIFE DATES: From: To: 12 February 1998 - 12 March 1998

Route of administration:

oral: gavage

Details on route of administration:

using stainless steel stomach tube

Vehicle:

propylene glycol

Details on oral exposure:

PREPARATION OF DOSING SOLUTIONS: formulations were prepared daily immediately prior to dosing, adjustment was made for specific gravity of vehicle

VEHICLE
- Justification for use and choice of vehicle: based on laboratory trial informations
- Concentration in vehicle: 10, 40 and 200 mg/mL
- Amount of vehicle: 5 mL/kg

Analytical verification of doses or concentrations:

yes

Details on analytical verification of doses or concentrations:

Samples of week 1 formulations were analysed to check stability, homogeneity in vehicle (highest and lowest concentration) and accuracy of preparations.

Duration of treatment / exposure:

28 days

Frequency of treatment:

7 days/week

Dose / conc.:

50 mg/kg bw/day (nominal)

Dose / conc.:

200 mg/kg bw/day (nominal)

Dose / conc.:

1 000 mg/kg bw/day (nominal)

No. of animals per sex per dose:

5

Control animals:

yes, concurrent vehicle

Details on study design:

- Dose selection rationale: based on 5-day dose range finding study
- Fasting period before blood sampling for clinical biochemistry: overnight fasting

Positive control:

none

Observations and examinations performed and frequency:

CAGE SIDE OBSERVATIONS: Yes
- Time schedule: twice daily

DETAILED CLINICAL OBSERVATIONS: Yes
- Time schedule: once prior to treatment, once daily thereafter

BODY WEIGHT: Yes
- Time schedule for examinations: days 1, 8, 15, 22 and 28

FOOD CONSUMPTION:
- weekly

WATER CONSUMPTION:
- subjective, no quantitative investigation

OPHTHALMOSCOPIC EXAMINATION: No

HAEMATOLOGY: Yes
- Time schedule for collection of blood: immediately prior to scheduled post mortem examination
- Anaesthetic used for blood collection: Not specified
- Animals fasted: Yes
- How many animals: 5/sex/group
- Parameters checked: erythrocyte count, haemoglobin, mean corpuscular volume, mean corpuscular haemoglobin concentration, platelet count, red cell distribution width, total leucocyte count, differential leucocyte count, prothrombin time, partial thromboplastin time

CLINICAL CHEMISTRY: Yes
- Time schedule for collection of blood: immediately prior to scheduled post mortem examination
- Animals fasted: Yes
- How many animals: 5/sex/group
- Parameters checked: alanine aminotransferase, aspartate aminotransferase, total bilirubin, total cholesterol, creatinine, glucose, urea, total protein, albumin, alkaline phosphatase, sodium, potassium, chloride, calcium, phosphorus

URINALYSIS: No

NEUROBEHAVIOURAL EXAMINATION: Yes
- Time schedule for examinations: during week 4 of treatment
- Dose groups that were examined: all dose groups
- Battery of functions tested: sensory activity / grip strength / motor activity

IMMUNOLOGY: No, but endpoints addressing potential adverse effects on the immune system were assessed

Sacrifice and pathology:

GROSS PATHOLOGY: Yes

HISTOPATHOLOGY: Yes
Tissues examined: adrenal glands, aorta, brain, caecum, colon, duodenum, epididymides, heart, ileum, jejunum, kidneys, liver, lung, lymph nodes, oesophagus, ovaries, pancreas, Peyer's patches, pituitary gland, prostate gland, rectum, sciatic nerve, spinal cord, spleen, sternum with bone marrow, stomach, testes, thymus, thyroid/parathyroid, trachea, urinary bladder, uterus, all gross lesions

Other examinations:

Organ weights: adrenal glands, brain, epididymides, heart, kidneys, liver, spleen, testes, thymus

Statistics:

Univariate one-way analysis of variance was used to assess the significance of intergroup differences.
If the variables could be assumed to follow a normal distribution, the Dunnett-test (many-to-one t-test) based on a pooled variance estimate was applied for the comparison of the treated groups and the control groups for each sex.
The Steel-test (many-to-one rank test) was applied when the data could not be assumed to follow a normal distribution.
All tests were two-sided and in all cases p < 0.05 was accepted as the lowest level of significance.
Group means were calculated for continuous data and medians were calculated for discrete data (scores) in the summary tables.
Test statistics were calculated on the basis of exact values for means and pooled variances. Individual values, means and standard deviations may have been rounded off before printing. Therefore, two groups may display the same printed means for a given parameter, yet display different test statistics values.

Clinical signs:

effects observed, non-treatment-related

Description (incidence and severity):

Salivation was observed in control and treated animals. This sign is often noted in rats of this age and strain following oral gavage and considered to be related to multiple intra-oesophageal intubation and/or the taste of the test substance. Therefore, this finding was considered not to be a sign of systemic toxicity.
Other findings, including a wound, scabs, diarrhoea and red staining were considered not be treatment related.

Mortality:

no mortality observed

Description (incidence):

No mortality occurred during the study period.

Body weight and weight changes:

effects observed, non-treatment-related

Description (incidence and severity):

The increased body weight gain of females receiving 200 mg/kg bw/d on days 15, 22 and 28, when compared to controls, was considered to have arisen by chance. This finding was considered of no toxicological significance, based on the absence of a treatment-related distribution and any other corroborative findings in these females. Body weights and body weight gain of other dose groups and the male animals remained in the same range as controls over the study period.

Food consumption and compound intake (if feeding study):

no effects observed

Food efficiency:

not examined

Water consumption and compound intake (if drinking water study):

not specified

Ophthalmological findings:

not examined

Haematological findings:

effects observed, non-treatment-related

Description (incidence and severity):

A minor statistically significant difference for erythrocyte count arising between controls and animals receiving 50 mg/kg bw/d was considered to have arisen by chance and not to represent a change of biological significance.

Clinical biochemistry findings:

effects observed, non-treatment-related

Description (incidence and severity):

Values in treated males or females achieving a level of statistical significance when compared to the controls, were considered to have arisen as a result of slightly high control values and in the absence of a treatment- related distribution or corroborative findings in the opposite sex, considered to be of no toxicological significance. Moreover, a decrease in the enzyme level of alanine aminotransferase is considered to be of no biological relevance.

Urinalysis findings:

not examined

Behaviour (functional findings):

no effects observed

Description (incidence and severity):

Any findings noted and the variation in motor activity did not indicate a relation with treatment.

Immunological findings:

not examined

Description (incidence and severity):

Endpoints addressing potential adverse effects on the immune system did not reveal any immunotoxic potential of the test item.

Organ weight findings including organ / body weight ratios:

effects observed, non-treatment-related

Description (incidence and severity):

A slightly increased relative liver weight, achieving a level of statistical significance when compared to controls, was found in females dosed with 1000 mg/kg bw/d. The increase in relative liver weight can reflect a higher metabolic activity in this organ and may thus simply represent an adaptive change related to high dose treatment rather than a toxic effect. There were no relevant findings in histopathology corresponding to the increased relative liver weight and this effect was not found in male animals.

Gross pathological findings:

no effects observed

Description (incidence and severity):

Findings noted among treated animals were considered to be within the range of biological variation for rats of this age and strain and not to represent a change of toxicological significance.

Neuropathological findings:

not examined

Histopathological findings: non-neoplastic:

no effects observed

Description (incidence and severity):

The small number of findings recorded are within the normal range of background alterations which may be seen in untreated rats of this age and strain.

Histopathological findings: neoplastic:

no effects observed

Description (incidence and severity):

The small number of findings recorded are within the normal range of background alterations which may be seen in untreated rats of this age and strain.

Other effects:

not examined

Key result

Dose descriptor:

NOAEL

Effect level:

1 000 mg/kg bw/day (nominal)

Based on:

test mat.

Remarks on result:

other: no adverse effects observed

Dose descriptor:

NOEL

Effect level:

200 mg/kg bw/day (nominal)

Based on:

test mat.

Sex:

female

Basis for effect level:

organ weights and organ / body weight ratios

Remarks on result:

other: considered as adaptive change

Analysis of dose preparations:

Test item formulations in proylene glycol were noted as stable for at least 4 hours and formed a homogeneous suspension at the concentrations tested. Analysis of accuracy of dose preparations revealed mean values of 102%, 103% and 100% for the dose groups 50, 200 and 1000 mg/kg bw/d, respectively. This was considered to represent an acceptable level of accuracy for formulations of this type.

Conclusions:

From the results of the study presented, 1000 mg/kg bw is considered to be the no-observed-adverse-effect-level (NOAEL) and 200 mg/kg bw/d the no-observed-effect-level (NOEL).

Executive summary:

A subacute 28-day toxicity study (OECD 407) was performed with the test item at doses of 0, 50, 200 and 1000 mg/kg bw/d administered orally (gavage) to rats (5 animals/sex/group). The test item was administered daily for 28 days. The following parameters were evaluated: clinical signs and mortality daily, funtional observation tests, body weight and food consumption weekly, clinical pathology and macroscopy at termination, organ weights and histopathology on selected tissues.

Accuracy, stability and homogeneity of test item formulations in the vehicle were demonstrated by analyses. Up to 1000 mg/kg bw/d no treatment-related findings were observed for the parameters evaluated. An increased relative liver weight was observed in females only at the highest dose. This finding was considered to be an adaptive change of the liver related to treatment without toxicological significance (higher metabolic activity).

There were no changes in clinical appearance, functional observations, body weights, food consumption, clinical laboratory investigations, macrosopic and microscopic examination that were considered to be treatment related.

From the results of this study, a NOAEL of 1000 mg/kg bw/d and a NOEL of 200 mg/kg bw/d were concluded.

Endpoint conclusion

Endpoint conclusion:

no adverse effect observed

Dose descriptor:

NOAEL

1 000 mg/kg bw/day

Study duration:

subacute

Species:

rat

Quality of whole database:

OECD 407, GLP

Repeated dose toxicity: inhalation - systemic effects

Endpoint conclusion

Endpoint conclusion:

no study available

Repeated dose toxicity: inhalation - local effects

Endpoint conclusion

Endpoint conclusion:

no study available

Repeated dose toxicity: dermal - systemic effects

Endpoint conclusion

Endpoint conclusion:

no study available

Repeated dose toxicity: dermal - local effects

Endpoint conclusion

Endpoint conclusion:

no study available

Additional information

A subacute 28-day toxicity study (OECD 407) was performed with the test item at doses of 0, 50, 200 and 1000 mg/kg bw/d administered orally (gavage) to rats (5 animals/sex/group). The test item was administered daily for 28 days. The following parameters were evaluated: clinical signs and mortality daily, funtional observation tests, body weight and food consumption weekly, clinical pathology and macroscopy at termination, organ weights and histopathology on selected tissues.

Accuracy, stability and homogeneity of test item formulations in the vehicle were demonstrated by analyses. Up to 1000 mg/kg bw/d no treatment-related findings were observed for the parameters evaluated. An increased relative liver weight was observed in females only at the highest dose. This finding was considered to be an adaptive change of the liver related to treatment without toxicological significance (higher metabolic activity).

There were no changes in clinical appearance, functional observations, body weights, food consumption, clinical laboratory investigations, macrosopic and microscopic examination that were considered to be treatment related.

From the results of this study, a NOAEL of 1000 mg/kg bw/d and a NOEL of 200 mg/kg bw/d were concluded (reference 7.5.1 -1).

Justification for classification or non-classification

Classification, Labeling, and Packaging Regulation (EC) No. 1272/2008

The available experimental test data are reliable and suitable for classification purposes under Regulation 1272/2008. The substance is not considered to be classified for repeated dose toxicity according to Regulation (EC) No 1272/2008 (CLP), as amended for the twelfth time in Regulation (EU) 2019/521.