A mixture of cis- and trans-cyclohexadec-8-en-1-one

Endpoint summary

Description of key information

In the key subacute feeding study in male and female Wistar rats according to OECD guideline 407, the NOAEL was 232.25 mg/kg bw/day in males and 250.75 mg/kg bw/day in females.

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:

key study

Study period:

1986-05-27 to to 1986-10-24

Reliability:

2 (reliable with restrictions)

Rationale for reliability incl. deficiencies:

guideline study with acceptable restrictions

Qualifier:

according to guideline

Guideline:

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

Deviations:

yes

Remarks:

compared to the current version, some endpoints were not examined

Qualifier:

according to guideline

Guideline:

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

Version / remarks:

1981-05-12

Deviations:

yes

Remarks:

compared to the current version, some endpoints were not examined

Principles of method if other than guideline:

Compared to the current version of OECD guideline 407, some endpoints were not examined (no analytical verification of test concentrations, no detailed clinical observations, no functional observations, not all organs according to OECD 407 examined).

- A few clinical chemistry determinations in the plasma were not carried out in a single male or female rat because the sample had clotted or because the sample was lost at bleeding.
- Urinary volume and density were not recorded in one female of the control group because unreliable values were obtained.
- The ovaries of one rat of the low-dose group were not weighed because of the presence of cystically dilated follicles.

GLP compliance:

yes (incl. QA statement)

Limit test:

no

Species:

rat

Strain:

Wistar

Remarks:

Bor: WISW (SPF;Cpb)

Details on species / strain selection:

The rat is the commonly used species for repeated-dose toxicity studies. The Wistar strain is a common laboratory strain with a good historical control data base.

Sex:

male/female

Details on test animals or test system and environmental conditions:

TEST ANIMALS
- Females nulliparous and non-pregnant: Yes
- Age at study initiation: 6-7 weeks
- Weight at study initiation: 146.1 - 146.4 g (males) and 121.4 - 121.7 g (females)
- Housing: 5 per sex per cage in suspended, stainless steel cages, fitted with wire-screen bottom and front (during acclimatation)
- Diet: Ad libitum (basal diet for rats and mice)
- Water: Ad libitum (tap water)
- Acclimation period: 13 days

ENVIRONMENTAL CONDITIONS
- Temperature (°C): 22 + 2
- Humidity (%): At least 40
- Air changes (per hr): 10
- Photoperiod (hrs dark / hrs light): 12/12

IN-LIFE DATES: From day 0 to day 28

Route of administration:

oral: feed

Details on route of administration:

The oral route is the likely route for human exposure.

Vehicle:

unchanged (no vehicle)

Details on oral exposure:

PREPARATION OF DOSING SOLUTIONS: The diet with the highest concentration was prepared first by adding the appropriate amount of the test substance (liquidized at 30°C) to the stock diet. The diets with the lower concentrations were obtained by diluting the top-dose diet with stock diet. Homogeneity of the diets was achieved by mixing for 2 minutes in a mechanical blender (Stephan cutter). Batches of 4 kg of each of the diets were prepared twice during the study. Immediately after preparation of the diets, samples were taken and subsequently stored at -20°C.

DIET PREPARATION
- Rate of preparation of diet: Twice during the study, on day 0 and day 14 of the study.
- Mixing appropriate amounts with powdered stock diet
- Storage temperature of food: The diets were stored at approx. 15°C until use.

Analytical verification of doses or concentrations:

no

Duration of treatment / exposure:

28 days

Frequency of treatment:

Continuously (feed)

Dose / conc.:

1 190.5 mg/kg bw/day (actual dose received)

Remarks:

15000 ppm in feed, corresponding to 1027-1341 mg/kg bw/day (females)

Dose / conc.:

1 198.75 mg/kg bw/day (actual dose received)

Remarks:

15000 ppm in feed, corresponding to 1004-1320 mg/kg bw/day (males)

Dose / conc.:

250.75 mg/kg bw/day (actual dose received)

Remarks:

3000 ppm in feed, corresponding to 219-274 mg/kg bw/day (females)

Dose / conc.:

232.25 mg/kg bw/day (actual dose received)

Remarks:

3000 ppm in feed, corresponding to 194-269 mg/kg bw/day (males)

Dose / conc.:

49.5 mg/kg bw/day (actual dose received)

Remarks:

600 ppm in feed, corresponding to 43-55 mg/kg bw/day (females)

Dose / conc.:

45.75 mg/kg bw/day (actual dose received)

Remarks:

600 ppm in feed, corresponding to 38-53 mg/kg bw/day (males)

No. of animals per sex per dose:

5 animals per sex per dose

Control animals:

yes, plain diet

Details on study design:

- Dose selection rationale: As the oral LD50 was >10000 mg/kg bw, the test substance was tested up to a limit dose of 1000 mg/kg bw/day. For mid and low doses, a 5-fold spacing was used.
- Fasting period before blood sampling for clinical biochemistry: Yes, for the 24-day collection (deprivation of water for 24 hours and of food during the last 16 hours of this period)

Positive control:

Not included

Observations and examinations performed and frequency:

CAGE SIDE OBSERVATIONS: Yes
- Time schedule: Daily
- Cage side observations included: General condition, behaviour, signs of ill health or reaction to treatment

DETAILED CLINICAL OBSERVATIONS: No

BODY WEIGHT: Yes
- Time schedule for examinations: Initially and on day 7, 14 ,21 and 28 of the study.

FOOD CONSUMPTION AND COMPOUND INTAKE:
- Food consumption for each animal determined and mean daily diet consumption calculated as g food/kg body weight/day: No (Food intake was measured per cage (5 animals) weekly)
- 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: No (efficiency of food utilization was calculated and expressed as gram weight gain per gram food consumed)

WATER CONSUMPTION AND COMPOUND INTAKE: Yes (water intake was measured on a cage basis (5 animals))
- Time schedule for examinations: During the first week of the study

OPHTHALMOSCOPIC EXAMINATION: No

HAEMATOLOGY: Yes
- Time schedule for collection of blood: On day 22
- Anaesthetic used for blood collection: Not specified
- Animals fasted: Not specified
- How many animals: All animals
- Parameters checked in table 1 were examined.

CLINICAL CHEMISTRY: Yes
- Time schedule for collection of blood: On day 24 (blood glucose) and on day 28
- Anaesthetic used for blood collection: Ether anaesthesia (terminal blood collection on day 28)
- Animals fasted: Yes, for the 24-day collection (deprivation of water for 24 hours and of food during the last 16 hours of this period)
- How many animals: All animals
- Parameters checked in table 2 were examined.

PLASMA/SERUM HORMONES/LIPIDS: No

URINALYSIS: Yes
- Time schedule for collection of urine: From day 24 to day 25
- Metabolism cages used for collection of urine: Not specified
- Animals fasted: Yes (Animals were deprived of water for 24 hours and of food for 16 hours. Urine was collected from individual animals during the last 16 hours of the deprivation period)
- Parameters examined: Volume and density of the urine

NEUROBEHAVIOURAL EXAMINATION: No

IMMUNOLOGY: No

Sacrifice and pathology:

GROSS PATHOLOGY: Yes (see table 3)

HISTOPATHOLOGY: Yes (see table 4)

Statistics:

Data an body weights were evaluated by one-way analysis of co-variance followed by Dunnett's multiple comparison test. Data on red blood cell variables, clinical chemistry values, volume and density of the urine, and organ weights were evaluated by one-way analysis of variance (ANWK) followed by Dunnett's multiple comparison test. Differential white blood cell counts were analyzed by the Mann-Whitney U-test. The histopathological changes were examined by Fisher's exact probability test.

Clinical signs:

no effects observed

Mortality:

no mortality observed

Body weight and weight changes:

effects observed, treatment-related

Description (incidence and severity):

Body weights were significantly decreased in the top-dose group in both sexes (up to 23%, >10% during the whole study in males and up to 10% in females). Body weights were relatively low in males of the mid-dose group (non-significant, up to 7% lower than the control group)

Food consumption and compound intake (if feeding study):

effects observed, treatment-related

Description (incidence and severity):

Food intake was decreased in the top-dose group in both sexes (up to 22% in males and up to 11% in females), whereas females tended to recover at the end of the study.

Food intake was relatively low in males of the mid-dose group (up to 10% lower than the control group).

Food efficiency:

effects observed, treatment-related

Description (incidence and severity):

Food conversion efficiency was decreased in the top-dose group in both sexes (up to 50% in males females) whereas females tended to recover at the end of the study.

Food conversion efficiency was relatively low in males of the mid-dose group (up to 21% lower compared to the control).

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

no effects observed

Ophthalmological findings:

not examined

Haematological findings:

effects observed, treatment-related

Description (incidence and severity):

15000 ppm: Red blood cell count and packed cell volume were increased in males, while haemoglobin concentration showed a tendency toward higher values. Packed cell volume was increased, MCHC was slightly decreased and red blood cell count and haemoglobin concentration showed a tendency toward higher values in females. These effects are considered to be a result of the lower body weight gain in this group.

3000 ppm: A decrease in thrombocytes count was observed in males which was considered incidential as no dose-dependence was observed.

600 ppm: A shift in the lymphocytes /neutrophils ratio was observed in females which was considered incidential.

Clinical biochemistry findings:

effects observed, treatment-related

Description (incidence and severity):

15000 ppm: Plasma urea and creatinine concentrations were increased in males. GPT activity was increased and albumin concentration decreased in females.

600 ppm: A decrease in GOT activity was observed in males which was considered incidential as no dose-dependence was observed.

Endocrine findings:

not examined

Urinalysis findings:

effects observed, treatment-related

Description (incidence and severity):

The urinary volume was relatively low whereas the density was increased in the top-dose group in both sexes.

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):

15000 ppm: The absolute and relative weights of the liver were increased in both sexes (50% increase of the relative liver weight in males and 30% increase of the relative liver weight in females). The absolute and relative weights of the adrenals were decreased in females (28% decrease of relative adrenal weight compared to the control). Decreases observed in the absolute weights of the spieen, heart, ovaries and testes in the top-dose group are considered to be secondary to the reduction in body weight gain in this group.

3000 ppm: The absolute and relative weights of the liver were increased in males (21% increase of the relative liver weight)

Gross pathological findings:

no effects observed

Neuropathological findings:

not examined

Histopathological findings: non-neoplastic:

effects observed, treatment-related

Description (incidence and severity):

15000 ppm: Moderate hepatocellular cytoplasmic alterations in both sexes (5/5 male animals and 4/5 female animals)

3000 ppm: Very slight hepatocellular cytoplasmic alterations in 1/5 males

Histopathological findings: neoplastic:

no effects observed

Other effects:

no effects observed

Key result

Dose descriptor:

NOAEL

Effect level:

250.75 mg/kg bw/day (actual dose received)

Based on:

test mat.

Sex:

female

Basis for effect level:

body weight and weight gain

clinical biochemistry

food consumption and compound intake

food efficiency

histopathology: non-neoplastic

organ weights and organ / body weight ratios

Key result

Dose descriptor:

NOAEL

Effect level:

232.25 mg/kg bw/day (actual dose received)

Based on:

test mat.

Sex:

male

Basis for effect level:

body weight and weight gain

food consumption and compound intake

food efficiency

histopathology: non-neoplastic

organ weights and organ / body weight ratios

Key result

Critical effects observed:

no

Conclusions:

In a subacute feeding study according to OECD guideline 407, the NOAEL was 232.25 mg/kg bw/day in male and 250.75 mg/kg bw/day in female Wistar rats.

Executive summary:

In a GLP compliant study according to OECD guideline 407, the toxicity of the test item was examined in four groups of 5 male and 5 female Wistar rats, which received the test substance in the diet at dose levels of 0 (control), 600, 3000 and 15000 ppm (corresponding to 45.75, 232.25 and 1198.75 mg/kg bw/day in males and 49.5, 250.75 and 1190.5 mg/kg bw/day in females) for a period of 4 weeks. General condition, behaviour and survival were not adversely affected by the administration of the test item. Growth, food intake and food conversion efficiency were decreased in the top-dose group in both sexes, and were relatively low in males of the mid-dose group. Red blood cell count and packed cell volume were increased in the top-dose group (more pronounced in males than in females). This effect was considered secondary to the lower food consumption and body weight gain. In the top-dose group, increases were observed in plasma urea and creatinine concentrations in males and in GPT activity in females, whereas albumin concentration was decreased in females.The urine concentration test showed an increased urinary density in the top-dose group in both sexes. As changes were not accompanied by changes in kidney weight or in microscopic renal morphology and the urine concentration test did not provide any indication of impaired kidney function, it was concluded that no nephrotoxic effect of the test substance was found. The weight of the liver was increased in males and females of the top-dose group and in males of the mid-dose group. The weight of the adrenals was decreased in females of the top-dose group. Gross examination at autopsy did not reveal any treatment-related changes. Upon microscopic examination, hepatocellular cytoplasmic alterations were observed in all males and all females of the top-dose group and in a single male of the mid-dose group.

 

Based on the results, the no observed adverse effect level (NOAEL) of the test item in the present study was found to be 232.25 mg/kg bw/day (3000 ppm) in males and 250.75 mg/kg bw/day (3000 ppm) in females.

Endpoint conclusion

Endpoint conclusion:

adverse effect observed

Dose descriptor:

NOAEL

232.25 mg/kg bw/day

Study duration:

subacute

Species:

rat

Quality of whole database:

Reliable with restrictions (RL2)

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

Key, subacute feeding study, rat, RL2

In a GLP compliant study according to OECD guideline 407, the toxicity of the test item was examined in four groups of 5 male and 5 female Wistar rats, which received the test substance in the diet at dose levels of 0 (control), 600, 3000 and 15000 ppm (corresponding to 45.75, 232.25 and 1198.75 mg/kg bw/day in males and 49.5, 250.75 and 1190.5 mg/kg bw/day in females) for a period of 4 weeks. General condition, behaviour and survival were not adversely affected by the administration of the test item. Growth, food intake and food conversion efficiency were decreased in the top-dose group in both sexes, and were relatively low in males of the mid-dose group. Red blood cell count and packed cell volume were increased in the top-dose group (more pronounced in males than in females). This effect was considered secondary to the lower food consumption and body weight gain. In the top-dose group, increases were observed in plasma urea and creatinine concentrations in males and in GPT activity in females, whereas albumin concentration was decreased in females. The urine concentration test showed an increased urinary density in the top-dose group in both sexes. As changes were not accompanied by changes in kidney weight or in microscopic renal morphology and the urine concentration test did not provide any indication of impaired kidney function, it was concluded that no nephrotoxic effect of the test substance was found. The weight of the liver was increased in males and females of the top-dose group and in males of the mid-dose group. The weight of the adrenals was decreased in females of the top-dose group. Gross examination at autopsy did not reveal any treatment-related changes. Upon microscopic examination, hepatocellular cytoplasmic alterations were observed in all males and all females of the top-dose group and in a single male of the mid-dose group.

 

Based on the results, the no observed adverse effect level (NOAEL) of the test item in the present study was found to be 232.25 mg/kg bw/day (3000 ppm) in males and 250.75 mg/kg bw/day (3000 ppm) in females.

Justification for classification or non-classification

The available experimental test data are reliable and suitable for classification purposes under Regulation (EC) No 1272/2008. Based on available data on repeated dose toxicity, the test item is not classified and labelled for Specific target organ toxicity (repeated exposure) (STOT-RE) according to Regulation (EC) No 1272/2008 (CLP), as amended for the eighteenth time in Regulation (EU) 2022/692.