Registration Dossier

Data platform availability banner - registered substances factsheets

Please be aware that this old REACH registration data factsheet is no longer maintained; it remains frozen as of 19th May 2023.

The new ECHA CHEM database has been released by ECHA, and it now contains all REACH registration data. There are more details on the transition of ECHA's published data to ECHA CHEM here.

Diss Factsheets

Administrative data

Description of key information

- 90-day, oral, rat (OECD 408): NOAEL >= 1000 mg/kg bw/day (m/f)


 The available information comprises an adequate and reliable study, and is thus sufficient to fulfil the standard information requirements set out in Annex VII, 8.5, of Regulation (EC) No. 1907/2006.

Key value for chemical safety assessment

Repeated dose toxicity: via oral route - systemic effects

Link to relevant study records
Reference
Endpoint:
sub-chronic toxicity: oral
Type of information:
experimental study
Adequacy of study:
key study
Study period:
22 Jun - 25 Nov 2021
Reliability:
1 (reliable without restriction)
Rationale for reliability incl. deficiencies:
guideline study
Qualifier:
according to guideline
Guideline:
OECD Guideline 408 (Repeated Dose 90-Day Oral Toxicity Study in Rodents)
Version / remarks:
adopted in 2018
Deviations:
yes
Remarks:
minor deviations related to body weight at arrival, blood sample collection, epididymal cauda weighing, and new version (version 7.5.1) of computerised system Pristima (details were given under material and methods); no influence on study outcome
GLP compliance:
yes (incl. QA statement)
Limit test:
no
Species:
rat
Strain:
Sprague-Dawley
Sex:
male/female
Details on test animals or test system and environmental conditions:
TEST ANIMALS
- Source: Envigo RMS B.V., Horst, Netherlands
- Females nulliparous and non-pregnant: yes
- Age at arrival: 27 - 29 days
- Weight at arrival: 96.6 - 110.0 g (males) and 84.2 - 104.4 g (females); The body weights of the animals at arrival were slightly outside the range indicated in the study protocol (96.6 - 110.0 g for males, 84.2 - 104.4 g for females, instead of 75 - 99 g).
- Fasting period before study: no
- Housing: Up to 5 animals of the same sex per cage in clear polysulfone solid bottomed cages
- Diet: A commercially available laboratory rodent diet (4 RF 21, Mucedola S.r.l., Settimo Milanese (MI), Italy) was offered ad libitum throughout the study.
- Water: Drinking water was supplied ad libitum to each cage via water bottles.
- Acclimation period: An acclimatisation period of 26 days was allowed before the start of treatment, during which time the health status of the animals was assessed by thorough observations.

DETAILS OF FOOD AND WATER QUALITY:
There was no information available to indicate that any non-nutrient substance likely to influence the effect of the test item was present in the drinking water or the diet.

ENVIRONMENTAL CONDITIONS
- Temperature (°C): 22 ± 2 °C
- Humidity (%): 55 ± 15%
- Air changes (per hr): 15 - 20
- Photoperiod (hrs dark / hrs light): 12 / 12

IN-LIFE DATES: 22 Jun - 26 Oct 2021
Route of administration:
oral: gavage
Vehicle:
corn oil
Details on oral exposure:
PREPARATION OF DOSING SOLUTIONS:
The required amount of test item was suspended in the vehicle. The preparations were made at 7/8-day intervals at concentrations of 25, 75 and 250 mg/mL, calculated and expressed in terms of test item as supplied.

VEHICLE
- Justification for use and choice of vehicle (if other than water): The vehicle was corn oil, since the test item was not-miscible in aqueous media
- Amount of vehicle (if gavage): The test item was administered orally by gavage at a dose volume of 4 mL/kg bw/day. The dose was administered to each animal on the basis of the most recently recorded body weight and the volume administered was recorded for each animal.

Analytical verification of doses or concentrations:
yes
Details on analytical verification of doses or concentrations:
The analytical method was validated previously in the range from 25 to 250 mg/mL. Linearity, accuracy and precision were within the limits stated in the validation protocol (r > 0.99; accuracy 85 - 115%; precision CV < 10%).
A 28 h stability at room temperature and a 10 d stability at room temperature were verified in the range from 25 to 250 mg/mL. According to the laboratory SOPs, suspensions are considered to be stable if concentration and homogeneity, after the defined period of storage, are still acceptable (85 - 115% for concentration and CV < 10% for homogeneity).
Samples of the preparations prepared on Day 1,Week 4 and Week 13 were analysed to check the concentration and homogeneity. Results of the analyses were within the acceptability limits stated in the laboratory SOPs for suspensions (85 - 115% for concentration and CV < 10% for
homogeneity).
Duration of treatment / exposure:
13 weeks followed by a recovery period of 4 weeks for 5 males and 5 females from groups 1 (control) and 4 (high dose group)
Frequency of treatment:
once daily, 7 days/week
Dose / conc.:
100 mg/kg bw/day (nominal)
Dose / conc.:
300 mg/kg bw/day (nominal)
Dose / conc.:
1 000 mg/kg bw/day (nominal)
No. of animals per sex per dose:
10; Extra 5 males and 5 females in groups 1 (control) and 4 (high dose group) for recovery period
Control animals:
yes, concurrent vehicle
Details on study design:
- Dose selection rationale: Dose levels have been selected in consultation with the Sponsor, based on information from preliminary non-GLP study.
- Fasting period before blood sampling for clinical biochemistry: yes
- Post-exposure recovery period: Recovery period of 4 weeks for 5 males and 5 females from groups 1 (control) and 4 (high dose group)
Positive control:
No
Observations and examinations performed and frequency:
CAGE SIDE OBSERVATIONS: Yes
- Time schedule: at least daily (animals were checked for mortality early in each working day and also in the afternoon)

DETAILED CLINICAL OBSERVATIONS: Yes
- Time schedule: weekly
Once before commencement of treatment and at least once per week during the study from the start of treatment, each animal was given a detailed clinical examination. Each animal was observed in an open arena. The test included observation and record of changes in gait and posture, reactivity to handling, presence of clonic or tonic movements, stereotypies or bizarre behaviour and effects on the autonomic nervous system (e.g. lachrymation, piloerection, unusual respiratory pattern). Changes in fur, skin, eyes, mucous membranes, occurrences of secretions and excretions was also recorded. Once during Week 12 of treatment and once during Week 4 of recovery an evaluation of sensory reactivity to stimuli of different modalities (e.g. auditory, visual and proprioceptive stimuli) and an assessment of grip strength was also performed.

BODY WEIGHT: Yes
- Time schedule for examinations: Each animal was weighed on the day of allocation to treatment group, on the day that treatment commenced, weekly thereafter and just prior to necropsy.

FOOD CONSUMPTION:
- Food consumption for each animal determined and mean daily diet consumption calculated as g food/kg body weight/day: No
The weight of food consumed by each cage of rats was recorded at weekly intervals following allocation. The group mean daily intake per rat was calculated.

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

WATER CONSUMPTION AND COMPOUND INTAKE: No

OPHTHALMOSCOPIC EXAMINATION: Yes
- Time schedule for examinations: prior to the commencement of treatment; during Week 13 of treatment (high dose and control groups)
- Dose groups that were examined: all animals (before the commencement of treatment), animals of the high dose and control groups (during week 13)
Both eyes of all animals were examined prior to the commencement of treatment by means of an ophthalmoscope, and by a slit-lamp microscope, after the instillation of 0.5% Tropicamide (Visumidriatic®, Visufarma, Rome, Italy). Observations carried out on the spare animals are not tabulated in the report and will be archived with the raw data of the study. The eyes of all animals from high dose and control groups were re-examined during Week 13 of treatment.

HAEMATOLOGY: Yes
- Time schedule for collection of blood: prior to necropsy and at the end of the recovery period
- Anaesthetic used for blood collection: Yes (isoflurane)
- Animals fasted: Not specified
- How many animals: all animals
- Parameters checked: haematocrit, haemoglobin, red blood cell count, reticulocyte count, mean red blood cell volume, mean corpuscular haemoglobin, mean corpuscular haemoglobin concentration, white blood cell count, differential leucocyte count (neutrophils, lymphocytes, eosinophils, basophils, monocytes, large unstained cells), platelets, prothrombin time
- Blood samples for haematology evaluations were collected from the sublingual vein, instead of the abdominal vena cava.

CLINICAL CHEMISTRY: Yes
- Time schedule for collection of blood: prior to necropsy and at the end of the recovery period
- Animals fasted: Not specified
- How many animals: all animals
- Parameters checked: alkaline phosphatase, alanine aminotransferase, aspartate aminotransferase, gamma-glutamyltransferase, urea, blood urea nitrogen, creatinine, bile acids, glucose, triglycerides, inorganic phosphorus, total bilirubin, total cholesterol, HDL, LDL, total protein, albumin, globulin, A/G ratio, sodium, potassium, calcium, chloride
- Blood samples for clinical chemistry evaluations were collected from the sublingual vein, instead of the abdominal vena cava.

PLASMA/SERUM HORMONES/LIPIDS: Yes
- Time of blood sample collection: prior to necropsy and at the end of the recovery period
- Animals fasted: Not specified
- How many animals: all animals; as no treatment-related effects were apparent following 13 wks treatment, samples collected at the end of recovery period were not analysed
- Parameters checked: T3, T4 and TSH
- Blood samples were collected from the sublingual vein, instead of the abdominal vena cava.

URINALYSIS: No

NEUROBEHAVIOURAL EXAMINATION: Yes
- Time schedule for examinations: Once during Week 12 of treatment and once during Week 4 of recovery (sensory activity, grip strength); once during Week 13 and once during Week 4 of recovery (motor activity; MA)
- Dose groups that were examined: all animals
- Battery of functions tested: Sensory reactivity to stimuli of different modalities (e.g. auditory, visual and proprioceptive stimuli) were measured; Motor activity measurements were performed using a computer generated random order

OTHER:
Oestrus cycle: From the first day of Week 12 up to the end of the treatment period and at the end of the recovery period, vaginal smears were taken daily in the morning. The vaginal smear data were examined to determine potential anomalies of the oestrus cycle.
Sacrifice and pathology:
Organ weights of the following organs: adrenal glands, brain (cerebrum, cerebellum, medulla/pons), coagulating glands, epididymides, one epididymal cauda, heart, kidneys, liver, ovaries, pituitary gland, prostate gland, seminal vesicles, spleen, testes, thymus, thyroid gland, uterus - cervix

Histopathological examination of the following organs: adrenal glands, aorta, bone marrow (from sternum), brain (cerebrum, cerebellum, medulla/pons), caecum, coagulating glands, colon, duodenum, epididymides, one epididymal cauda, heart, ileum, jejunum (including Peyer's patches), kidneys, liver, lungs (including mainstem bronchi), lymph nodes - cervical, lymph nodes - mesenteric, mammary area, oesophagus, ovaries, oviducts, pancreas, parathyroid glands, pituitary gland, prostate gland, rectum, salivary glands, sciatic nerve, seminal vesicles, skin, spina cord (at 3 levels), spleen, stomach, testes, thymus, thyroid gland, trachea, urinary bladder, uterus - cervix, vagina

- One epididymal cauda was weighed separately from epididymides only in Groups 1 and 4 where sperm analysis was carried out and not in Group 2 and 3 animals, since no sperm analysis was carried out in these animals. However, as in the study protocol it was not clearly indicated to weigh one cauda for sperm analysis only, this is a protocol deviation, although no critical data are missing.
Statistics:
Standard deviations were calculated as considered appropriate. For continuous variables the significance of the differences amongst groups was assessed by analysis of variance. Differences between each treated group and the control group were assessed by Dunnett’s test using a pooled error variance. The homogeneity of the data was verified by Bartlett’s test before Dunnett’s test. If the data were found to be inhomogeneous a Modified t test (Cochran and Cox) was applied.
The mean values, standard deviations and statistical analysis were calculated from the actual values in the computer without rounding off. Statistical analysis of histopathological finding was carried out by means of a non-parametric Kolmogorov-Smirnov test.
Clinical signs:
effects observed, non-treatment-related
Description (incidence and severity):
Salivation, considered non-adverse, was occasionally observed during the treatment phase of the study in 7 out of 10 males and 2 out of 10 females treated at 300 and in all males and 12 out of 15 females dosed at 1000 mg/kg bw/day. It was also episodically recorded in one control male and one dosed at 100 mg/kg bw/day.
One female treated at 100 mg/kg bw/day presented dyspnoea on a single occasion and rales for almost every day starting from Day 39. Slight hairloss on the head was observed in 3 out of 15 females dosed at 1000 mg/kg bw/day, while one female of the same group showed moderate swelling of the neck associated with a palpable mass. These clinical signs were not considered to be treatment-related due to the occasional occurrence and/or the absence of dose relation.
No clinical signs were observed during the recovery phase.
No treatment-related changes were found at the weekly clinical examination, which included an evaluation of neurotoxicity.
Slight statistically significant decreases and /or increases in the number of rearing were occasionally observed during treatment-period in males and females receiving 100, 300 and 1000 mg/kg bw/day, when compared to controls. Due to the occasional occurrence of these changes, to their inconsistent direction and absence of dose-relation, they were not considered to be treatment-related.
Mortality:
mortality observed, non-treatment-related
Description (incidence):
Premature death occurred in one male treated at 300 mg/kg bw/day (mid-dose), found dead on Day 93 following blood sampling procedure, and one female receiving 1000 mg/kg bw/day (high dose), found dead on Day 79.
No clinical signs were observed prior to death in the male animal. Dark colour of lungs was observed at macroscopic examination and congestion of lungs at histopathological evaluation. The cause of death can be related to the bleeding procedure.
The female showed salivation on Days 77 and 78. No abnormalities were detected at macroscopic and microscopic examinations. Based on the above data, the cause of death of this animal and the relationship to treatment cannot be established.
Body weight and weight changes:
no effects observed
Description (incidence and severity):
When compared to control animals, no treatment-related changes were noted in mean body weights in both genders, during the treatment and recovery periods of the study.
Food consumption and compound intake (if feeding study):
no effects observed
Description (incidence and severity):
No treatment-related changes were observed in food consumption in male and female animals during the treatment and recovery periods.
Food efficiency:
not examined
Water consumption and compound intake (if drinking water study):
not examined
Ophthalmological findings:
no effects observed
Description (incidence and severity):
Before the start of treatment, no animals showed ocular abnormality at the ophthalmoscopic examination. Both eyes of all animals from high dose and control groups were re-examined during Week 13 of treatment (on study Day 86). No ocular findings were detected.
Haematological findings:
effects observed, non-treatment-related
Description (incidence and severity):
Dosing Phase
No treatment-related changes were recorded. The statistically significant differences between control and high dose females (haemoglobin and lymphocytes) were within the range of expected biological variation, therefore they were considered to be incidental.
The reduction of prothrombin time in males dosed at 1000 mg/kg bw/day (5%) and the prolongation of the same parameter in females receiving 100 and 300 mg/kg bw/day (5%, both) were not dose-related and/or of minimal severity, therefore they were considered to be unrelated to treatment.

Recovery Phase
No changes were recorded.
Clinical biochemistry findings:
effects observed, non-treatment-related
Description (incidence and severity):
Dosing Phase
Treatment-related changes of some hepatic parameters (increases of alkaline phosphatase and alanine aminotransferase and decreases of protein, albumin and globulin) were recorded in a number of treated animals. The severity of the findings observed was not considered to be suggestive of tissue/organ injury, they could rather reflect an organ adaptation and were not considered to be adverse. Any other differences in the serum chemistry parameters, including those determined to be statistically significant, were within the range of the control group data and/or due to biological variation and/or not dose-related, therefore they were considered to be incidental.

Thyroid stimulating hormone was slightly higher than controls in females dosed at 1000 mg/kg bw/day (33%). Since no other related findings were observed (i.e. T3 and T4 decrease, thyroid weight and/or histopathological changes) and TSH values were within the range of the control group data, the above change was considered to be incidental.

Recovery Phase
At the end of the recovery period, no differences between control and treated animals were observed, confirming complete reversibility.
Endocrine findings:
not specified
Description (incidence and severity):
The following ED-related parameters were investigated in the study:
- thyroid hormones
- accessory sex organ weight and histopathology
- sex organ weight and histopathology
- estrus cyclicity
- thyroid weight and histopathology
- sperm analysis
- vaginal smears
- adrenal weight and histopathology
- brain weight
- pituitary gland weight and histopathology

For details, please refer to the respective result fields and the endpoint summary.
Urinalysis findings:
not examined
Behaviour (functional findings):
effects observed, non-treatment-related
Description (incidence and severity):
No relevant differences that could be considered treatment-related were observed at functional tests (sensory reactivity, landing footsplay, grip strength) performed at the end of treatment and recovery periods.
A slight statistically significant increase of mean grip strength was recorded in females treated at 100, 300 and 1000 mg/kg bw/day at the end of treatment period. This increase was also observed in females treated with 1000 mg/kg bw/day at the end of the recovery phase, when compared to controls. These increases did not show any clear dose-relation or association with other findings, therefore they were not considered to be treatment-related.
Motor activity measurements performed at the end of the treatment and recovery periods did not show any differences between treated animals and controls.
Immunological findings:
not examined
Organ weight findings including organ / body weight ratios:
effects observed, non-treatment-related
Description (incidence and severity):
Final sacrifice
There were no changes that could be considered adverse in organ weights when compared to the controls. Any organ weight variations, including the increase in relative mean liver weights in high dose animals of both sexes (11% and 8% for males and females, respectively, for relative liver weight to body weight) and mid-dose females (14% for relative liver weight to body weight) were not correlated to any histopathological changes, in the range of expected spontaneous changes in rats of the same age and considered not adverse.

Recovery sacrifice
After 4 weeks of recovery, no treatment-related organ weight changes were reported in animals of both sexes previously treated with the high dose, when compared with controls.
Gross pathological findings:
effects observed, non-treatment-related
Description (incidence and severity):
Final sacrifice
There were no treatment-related macroscopic observations at the end of the treatment period. Any macroscopic observations, including the subcutaneous mass with dark creamy content observed during necropsy of a high dose female confirmed at histopathological evaluation as malignant adenocarcinoma of the mammary gland, had a comparable incidence in control and treated groups and/or are characteristically seen in untreated rats of the same age and were considered incidental and unrelated to treatment.

Recovery sacrifice
There were no treatment-related macroscopic observations at the end of the recovery period. Any macroscopic observations were within the range of occasionally observed and expected spontaneous changes in rats of the same age and therefore considered unrelated to treatment.
Neuropathological findings:
not examined
Histopathological findings: non-neoplastic:
effects observed, non-treatment-related
Description (incidence and severity):
Final sacrifice
There were no treatment-related microscopic observations at the end of the treatment period.
There were no test item-related microscopic observations in the testis (stage aware evaluation on PAS-stained slides).
There was no test item-related effect on oestrus cycle stages.
Any microscopic observations had a comparable incidence in control and treated groups and/or are characteristically seen in untreated rats of the same age and were considered incidental and unrelated to treatment.
Histopathological findings: neoplastic:
not examined
Other effects:
no effects observed
Description (incidence and severity):
No treatment-related anomalies were observed in the oestrus cycle of the treated females during the last 2 weeks of the treatment period and at the end of recovery phase.

Sperm motility, concentration and morphology did not show any significant differences between control and high dose group.
Dose descriptor:
NOAEL
Effect level:
>= 1 000 mg/kg bw/day (nominal)
Based on:
test mat.
Sex:
male/female
Basis for effect level:
other: no adverse effects were observed up to the highest dose
Critical effects observed:
no
Endpoint conclusion
Endpoint conclusion:
no adverse effect observed
Dose descriptor:
NOAEL
1 000 mg/kg bw/day
Study duration:
subchronic
Species:
rat

Repeated dose toxicity: inhalation - systemic effects

Endpoint conclusion
Endpoint conclusion:
no study available
Study duration:
subacute

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
Study duration:
subacute

Repeated dose toxicity: dermal - local effects

Endpoint conclusion
Endpoint conclusion:
no study available

Additional information

Repeated dose toxicity – oral

Subchronic

The subchronic oral toxicity of the registered substance was investigated in a study performed according to OECD guideline 408 under GLP conditions (Croda Europe Limited, 2022). The registered substance was administered once daily via gavage to groups of 10 Sprague-Dawley rats per sex at doses of 100, 300, and 1000 mg/kg bw in corn oil for a period of 13 weeks. A control group received the vehicle alone. Additionally, satellite groups of 5 animals per sex (control and high dose) were used to investigate the reversibility of effects during a 4 week post-exposure recovery period.

One male treated at 300 mg/kg bw/day (mid-dose), and one female receiving 1000 mg/kg bw/day (high dose) were found dead on Days 93 and 79, respectively. On the basis of the in vivo and post mortem data, the cause of death and its relation to treatment cannot be established for the high dose female, while it can be ascribed to the blood sampling procedure for the mid-dose male.

Besides salivation, no clinical signs were observed. Due to the occasional occurrence and/or absence of dose-relationship, these effects were not considered as treatment-related. No clinical signs were observed during the recovery phase. Observation of animals at removal from the cage and in an open arena did not reveal any treatment related changes.

No signs indicating neurotoxic effects were seen during the in vivo phase of the study. The slight statistically significant variations in mean rearing events number or sensor reactivity recorded in treated males and/or females at the end of treatment period or recovery period did not show any clear dose-relation, consistency between sexes or association with other findings, therefore they were not considered to be of toxicological relevance.

Body weight, food consumption and oestrus cycle were not affected by treatment. No lesions were recorded at ophthalmological examination.

No treatment-related changes were recorded at the end of treatment and recovery periods in haematological and coagulation parameters. Changes of some hepatic parameters were recorded in a number of treated animals, mainly dosed at 300 and 1000 mg/kg bw/day. The severity of the findings observed was not considered to be suggestive of tissue/organ injury, they could rather reflect an organ adaptation and were considered to be non adverse. No treatment-related changes were recorded for thyroid hormones levels.

No treatment-related or adverse changes were observed in terminal body weight and absolute and relative organ weights of treated animals that completed the treatment or recovery periods. The slight increase in relative mean liver weights in high dose animals of both sexes and mid-dose females, recorded at the end of the dosing period, was not correlated to any histopathological changes and was considered not adverse.

There were no treatment-related macroscopic and microscopic observations at the end of treatment and recovery periods. There were no test item-related microscopic observations in the testis (stage aware evaluation on PAS-stained slides). There was no test item-related effect on oestrus cycle stages. No significant differences between control and treated groups were observed at sperm analysis.

In conclusion, some treatment-related effects were observed in animals dosed at 300 and 1000 mg/kg bw/day (salivation, alterations of some hepatic parameters, very slight increments in liver weights) following treatment with the registration substance, when administered by oral gavage for 13 consecutive weeks at the dosages of 100, 300 and 1000 mg/kg bw/day. However, due to their low magnitude and in the absence of correlated histopathological changes, these effects were not considered to be adverse. Based on these findings, a No Observed Adverse Effect Level (NOAEL) of 1000 mg/kg bw/day was concluded for this study.

Justification for classification or non-classification

According to Regulation (EC) No. 1272/2008 the data on repeated dose toxicity are conclusive but not sufficient for classification.