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Please be aware that this old REACH registration data factsheet is no longer maintained; it remains frozen as of 19th May 2023.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

Oral (OECD 407), rat: NOAEL 1000 mg/kg bw/day
Inhalation: not applicable (endpoint waived)
Dermal: not applicable (endpoint waived)

Key value for chemical safety assessment

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:
02 February 2004 - 30 March 2004
Reliability:
1 (reliable without restriction)
Rationale for reliability incl. deficiencies:
other: see 'Remark'
Remarks:
Study conducted in compliance with agreed protocols, with no or minor deviations from standard test guidelines and/or minor methodological deficiencies, which do not affect the quality of the relevant results. The study report was conclusive, done to valid guidelines and the study was conducted under GLP conditions.
Qualifier:
according to guideline
Guideline:
OECD Guideline 407 (Repeated Dose 28-Day Oral Toxicity Study in Rodents)
Deviations:
no
Qualifier:
according to guideline
Guideline:
EU Method B.7 (Repeated Dose (28 Days) Toxicity (Oral))
Deviations:
no
Qualifier:
according to guideline
Guideline:
other: EPA OPPTS 870.3050 (Repeated dose 28-day oral toxicity study in rodents)
Deviations:
no
GLP compliance:
yes
Limit test:
no
Species:
rat
Strain:
Wistar
Sex:
male/female
Details on test animals or test system and environmental conditions:
TEST ANIMALS
- Strain: Wistar Crl:(Wl) BR (outbred, SPF-Quality).
- Age at study initiation: Approximately 6 weeks.
- Weight at study initiation: Mean group body weights on day 1 of treatment: males 191 – 194 g; females 159 – 162 g.
- Housing: Group housing of 5 animals per sex per cage in stainless steel suspended cages with wire mesh floors (55 x 34 x 21.5 (hieight) cm). During activity monitoring, animals were individually housed overnight in Macrolon plastic cages (type III, height 15 cm) with sterilised sawdust provided as bedding.
- Diet: Standard pelleted laboratory animal diet, ad libitum.
- Water: Tap water, ad libitum.
- Acclimation period: Acclimatisation period was at least 5 days before start of treatment under laboratory conditions.

ENVIRONMENTAL CONDITIONS
- Temperature (°C): 21.0 ± 3.0 °C (range 18.7 – 25.3 °C).
- Humidity (%):30 – 70 % (range 32 – 93 %).
- Air changes (per hr): Approximately 15 air charges per hour.
- Photoperiod (hrs dark / hrs light): 12 hours of artificial fluorescent light and 12 hours of darkness per day.
Route of administration:
oral: gavage
Vehicle:
propylene glycol
Details on oral exposure:
PREPARATION OF DOSING SOLUTIONS
Formulations (w/w) were prepared daily within 4 hours prior to dosing and were homogenised to a visually acceptable level.
Storage conditions: Ambient temperature.

DOSE
- Dose volume: 5 mL/kg bw. Actual dose volumes were calculated weekly according to the latest recorded body weight.

VEHICLE
- Justification for use and choice of vehicle (if other than water): Propylene glycol was selected for use based on trial formulations performed at the test facility.
Analytical verification of doses or concentrations:
yes
Details on analytical verification of doses or concentrations:
SAMPLING
- Concentrations: Samples of week 1 formulations were analysed to check homogeneity (highest and lowest concentration) and accuracy of preparation (all concentrations). Stability in vehicle over 4 hours was also determined (highest and lowest concentration).
- Sampling method: Samples (approximately 500 mg), which were taken from the formulations using a pipette, were accurately weighed into volumetric flasks (25 or 100 mL). For determination of accuracy, duplicate samples were taken at 50 % height or at 90, 50 and 10 % height. For determination of homogeneity, duplicate samples were taken at 90, 50 and 10 % height. For determination of stability, duplicate samples were taken at 50 % height.
The flasks were filled to the mark with acetonitrile and the samples of the 1000 mg/kg day dose group were sonicated for 5 minutes in order to dissolve the test material. The solutions were further diluted with acetonitrile to obtain concentrations within the calibration range. Samples and solutions containing the test material were protected from light as much as possible using amber glassware.

ANALYTICAL METHOD
- Method: Quantitative analyses were based on the largest peak of the HPLC chromatogram of the test material.
- Conditions:
> Column: Symmetry Shield RP-8; 150 x 4.6 mm; dp = 5 µm
> Mobile phase: 100 % Acetonitrile
> Flow: 1 mL/min
> Injection volume: 100 µL
- Detection method: Spectrophotometric (wavelength of 273 nm).
- Calibration coefficient of correlation: r > 0.99 for each calibration curve.
- Procedural recovery samples: 96 – 109 %; test concentrations were not corrected for recovery.

RESULTS
- Concentration: Formulations were found to be in agreement with the target concentrations (97 to 101 % of target concentration).
- Homogeneity: Formulations were considered to be homogenous (≤ 0.9 % relative standard deviation).
- Stability: Formulations were found to be stable for at least 4 hours when stored at room temperature and not protected from light.
Duration of treatment / exposure:
28 days.
Frequency of treatment:
Daily.
Remarks:
Doses / Concentrations:
0, 50, 150 and 1000 mg/kg/day
Basis:
actual ingested
No. of animals per sex per dose:
5 animals per sex per dose.
Control animals:
yes, concurrent vehicle
Details on study design:
- Dose selection rationale: The dose levels were selected on the basis of a 5-day dose range finding study where three males per dose level were treated at 150 and 1000 mg/kg (dose volume 5 mL/kg bw).
The following parameters were observed: mortality, clinical signs, body weight, food consumption, macroscopic examination and organ weights (liver and kidneys).
All parameters appeared to be normal and no overt signs of toxicity were observed.
Based on the results of this range finding study, dose levels selected for the main study were: 50, 150 and 1000 mg/kg body weight/day.
- Rationale for animal assignment (if not random): Prior to commencement of treatment, randomisation took place by computer-generated random algorithm according to body weight, with all animals within ± 20 % of the sex mean.
Observations and examinations performed and frequency:
CAGE SIDE OBSERVATIONS: Yes
- Time schedule: At least twice daily.
- Observations: Mortality and viability.

DETAILED CLINICAL OBSERVATIONS: Yes
- Time schedule: Once daily. Additional observations were made outside the home cage in a standard arena once prior to the start of treatment and on a weekly basis thereafter.
- Observations:
- Scoring: Symptoms were graded according to fixed scales and the time of onset, degree and duration were recorded:
Maximum grade 1: grade 0 = absent, grade 1 = present;
Maximum grade 3 or 4: grade 1 = slight, grade 2 = moderate, grade 3 = severe, grade 4 = very severe.

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

FOOD CONSUMPTION: Yes
- Time schedule: Determined weekly.

WATER CONSUMPTION: No. Although subjective appraisal was maintained during the study, no quantitative investigation was introduced as no effect was suspected.

HAEMATOLOGY: Yes
- Time schedule for collection of blood: Samples were collected immediately prior to scheduled necropsy.
- Anaesthetic used for blood collection: Yes (iso-flurane).
- Animals fasted: Yes (overnight, maximum 20 hours before sampling).
- How many animals: All rats were sampled from each dose group.
- Parameters checked:
Haematology: Erythrocytes count, haemoglobin, haematocrit, mean corpuscular volume, mean corpuscular haemoglobin, mean corpuscular haemoglobin concentration, platelet count, red cell distribution width, total leucocytes count and differential leucocyte count (neutrophils, lymphocytes, monocytes, eosinophils and basophils).
Clotting Potential: Prothrombin time and partial thromboplastin time.

CLINICAL CHEMISTRY: Yes
- Time schedule for collection of blood: Samples were collected immediately prior to scheduled necropsy.
- Animals fasted: Yes (overnight, maximum 20 hours before sampling).
- How many animals: All rats were sampled from each dose group.
- Parameters checked: Alanine aminotransferase, alkaline phosphatase, aspartate aminotransferase, bilirubin (total), chloride, cholesterol (total), creatinine, glucose, phosphorus, protein (total), protein (albumin), urea, calcium, potassium and sodium.

NEUROBEHAVIOURAL EXAMINATION: Yes
- Time schedule for examinations: Once during week 4.
- Dose groups that were examined: All groups.
- Battery of functions tested:
Hearing ability, pupillary reflex, static righting reflex and grip strength (Score: 0 = normal/present, 1 = abnormal/absent).
Motor activity test (recording period: 12 hours during overnight for individual animals, using a computerised monitoring system).
Sacrifice and pathology:
NECROPSY: All animals surviving to the end of the observation period were deeply anaesthetised using iso-flurane and subsequently exsanguinated.

GROSS PATHOLOGY: Yes
Tissues and organs examined (fixed in neutral phosphate buffered 4 % formaldehyde solution: Adrenal glands, aorta, brain (cerebellum, mid-brain, cortex), caecum, cervix, clitoral gland, colon, duodenum, epididymides, eyes with optic nerve and Harderian gland, female mammary gland area, femur including joint, heart, ileum, jejunum, kidneys, larynx, lacrimal gland (exorbital), liver, lung (infused with formalin), lymph nodes (mandibular, mesenteric), nasopharynx, oesophagus, ovaries, pancreas, Peyer's patches (jejunum, ileum) if detectable, pituitary gland, preputial gland, prostate gland, rectum, salivary glands (mandibular, sublingual), sciatic nerve, seminal vesicles, skeletal muscle, skin, spinal cord (cervical, midthoracic, lumbar), spleen, sternum with bone marrow, stomach, testes, thymus, thyroid including parathyroid, tongue, trachea, urinary bladder, uterus, vagina and all gross lesions.

ORGAN WEIGHTS: Yes
Organs examined: Adrenal glands, brain, epididymides, heart, kidneys, liver, spleen, testes and thymus.

HISTOPATHOLOGY: Yes
Tissues and organs examined: All organs and tissues samples (detailed above) were processed, embedded and cut at a thickness of 2 - 4 µm and stained with haematoxylin and eosin.
The following slides were examined by a pathologist: All tissues and organs collected at the scheduled sacrifice from all animals of the control and the highest dose group; all gross lesions of all animals.
Statistics:
The following statistical methods were used to analyse the data:
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 Student's t-test was applied for motor activity data.
The Steel-test (many-to-one rank test) was applied when the data could not be assumed to follow a normal distribution.
The exact Fisher-test was applied to frequency data.
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.
Clinical signs:
no effects observed
Mortality:
no mortality observed
Body weight and weight changes:
no effects observed
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 examined
Ophthalmological findings:
not examined
Haematological findings:
no effects observed
Clinical biochemistry findings:
no effects observed
Urinalysis findings:
not examined
Behaviour (functional findings):
no effects observed
Organ weight findings including organ / body weight ratios:
no effects observed
Gross pathological findings:
no effects observed
Histopathological findings: non-neoplastic:
no effects observed
Histopathological findings: neoplastic:
not examined
Details on results:
CLINICAL SIGNS AND MORTALITY
No mortality occurred during the study period.
There were no clinical signs of toxicity or behavioural changes over the 28-day observation period that were considered to be related to treatment.
General colonic spasms were seen in one female dosed at 50 mg/kg/day in week 3, on two consecutive days. This finding was incidental and in the absence of a dose relationship it was considered not to be related to treatment.
Incidental findings included scabs and chromodacryorrhoea in males, and alopecia and scabs in females. These findings are commonly noted in rats of this age and strain that are housed and treated under the conditions in this study. At the incidence observed, these were considered signs of no toxicological significance.

BODY WEIGHT AND WEIGHT GAIN
Body weights and body weight gain for treated animals remained in the same range as controls over the 4-week study period.

FOOD CONSUMPTION
Food consumption before or after allowance for body weight was similar between treated and control animals.

HAEMATOLOGY
No toxicologically relevant changes occurred in haematological parameters of treated rats.
Increases of neutrophil counts with concurrently reduced lymphocyte counts were noted in females among the dose groups without a treatment related distribution. This shift in type of white blood cells was considered to be a secondary non-specific response to stress and to be of no toxicological significance.
Minor statistically significant differences in mean corpuscular haemoglobin concentration arising between control and treated males receiving 150 mg/kg/day were considered to have arisen by chance and not to represent a change of biological significance.

CLINICAL CHEMISTRY
There were no differences noted between control and treated rats that were considered to be related to treatment with the test material.

Values in treated males and females achieving a level of statistical significance when compared to controls were considered to have arisen as a result of biological variation, and in the absence of a treatment-related distribution or corroborative findings in the opposite sex, were considered to be of no toxicological significance.

NEUROBEHAVIOUR
No changes were observed in hearing ability, pupillary reflex, static righting reflex and grip strength in the animals treated with test material, when compared to control animals.
The variation in motor activity did not indicate a relationship with treatment.
A statistically significant decrease in motor activity measured in the lower sensor of males dosed at 150 mg/kg/day occurred in the absence of a dose-related response, similar changes of the low or high sensors, and/or supportive clinical signs. Therefore, it was considered to be of no toxicological relevance.

ORGAN WEIGHTS
No toxicologically significant changes were noted regarding organ weights and organ/body weight ratios.
Statistically significant changes between absolute and relative spleen weights of females treated at 150 mg/kg/day, and changes in relative adrenal weights in females dosed at 50 and 150 mg/kg/day, when compared to controls, were considered not to be a sign of toxicity, based on the absence of a treatment-related distribution or corroborative findings in the opposite sex.

GROSS PATHOLOGY
Necropsy did not reveal any toxicologically relevant alterations.
Incidental findings among control and treated animals included pelvic dilation of the kidneys, nodules in the epididymides, adrenal glands grown together with kidney, dark red discolouration and foci of the thymus, and dark red discoloured and enlarged mandibular lymph nodes. These findings are occasionally seen among rats used in these types of studies. In the absence of a treatment-related distribution they were considered to be changes of no toxicological significance.
Watery fluid in the uterus, found in two control, one low dose and one high dose females, is related to a stage in the oestrous cycle and is a normal finding.

HISTOPATHOLOGY: NON-NEOPLASTIC
There were no microscopic findings recorded which could be attributed to treatment with the test material.
All microscopic findings were within the range of background pathology encountered in Wistar rats of this age and strain and occurred at similar incidences and severity in both control and treated rats.
Dose descriptor:
NOAEL
Effect level:
1 000 mg/kg bw/day (actual dose received)
Based on:
test mat.
Sex:
male/female
Basis for effect level:
other: see 'Remark'
Critical effects observed:
not specified
Conclusions:
Under the conditions of the test, no significant, toxicologically relevant effects were observed in rats treated up to 1000 mg/kg bw/day. On this basis, the NOAEL was determined to be 1000 mg/kg bw/day.
Executive summary:

The subacute toxicity of the test material was determined using Wistar strain rats in a 28 day oral toxicity study performed under GLP conditions and in line with the standardised guidelines OECD 407, EU Method B.7 and EPA OPPTS 870.3050.

Groups of rats (5 animals per sex per dose) were exposed to the test material at doses of 50, 150 and 1000 mg/kg bw/day via oral gavage for 28 consecutive days. A control group (5 animals per sex per dose) was exposed to the vehicle, propylene glycol, only.

The following parameters were evaluated: mortality and clinical signs (daily), functional observation tests, body weight and food consumption (weekly). Clinical pathology and macroscopic examinations were carried out at termination, along with the determination of organ weights and histopathologic evaluation on a selection of tissues.

No mortality occurred. There were no changes at determination of clinical appearance, performance of functional observations, body weight and food consumption measurements, or alterations during clinical laboratory investigations, macroscopic examination, organ weight determination and microscopic examination that were considered to be an effect of treatment.

Under the conditions of the test, no significant, toxicologically relevant effects were observed in rats treated up to 1000 mg/kg bw/day. Based on these results the NOAEL was determined to be 1000 mg/kg bw/day.

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:
The study was performed under GLP conditions and in accordance with standardised guidelines. The study was therefore assigned a reliability score of 1 in accordance with the principles for assessing data quality as defined by Klimisch et al. (1997). The quality of the dataset is therefore considered to be high.

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

Repeated Dose Toxicity: Oral

The subacute toxicity of the test material was determined using Wistar strain rats in a 28 day oral toxicity study (Hooiveld, 2004) performed under GLP conditions and in line with the standardised guidelines OECD 407, EU Method B.7 and EPA OPPTS 870.3050.

Groups of rats (5 animals per sex per dose) were exposed to the test material at doses of 50, 150 and 1000 mg/kg bw/day via oral gavage for 28 consecutive days. A control group (5 animals per sex per dose) was exposed to the vehicle, propylene glycol, only.

The following parameters were evaluated: mortality and clinical signs (daily), functional observation tests, body weight and food consumption (weekly). Clinical pathology and macroscopic examinations were carried out at termination, along with the determination of organ weights and histopathologic evaluation on a selection of tissues.

No mortality occurred. There were no changes at determination of clinical appearance, performance of functional observations, body weight and food consumption measurements, or alterations during clinical laboratory investigations, macroscopic examination, organ weight determination and microscopic examination that were considered to be an effect of treatment.

Under the conditions of the test, no significant, toxicologically relevant effects were observed in rats treated up to 1000 mg/kg bw/day. Based on these results the NOAEL was determined to be 1000 mg/kg bw/day.

Repeated Dose Toxicity: Inhalation

In accordance with point 8.6.1, column 1 of Annex VIII of Regulation (EC) No. 1907/2006, testing for this endpoint should be performed using an appropriate route of exposure. A 28 day oral study was conducted to fulfil the subacute toxicity endpoint, since this was considered to be the most appropriate route of exposure for the substance. Exposure via inhalation is considered to be negligible based on the vapour pressure (0.0045 Pa at 20 °C) and particle size (10 % < 25.46 µm) of the substance; it is therefore considered that testing via this route is not appropriate and has been omitted on this basis.

Repeated Dose Toxicity: Dermal

In accordance with point 8.6.1, column 1 of Annex VIII of Regulation (EC) No. 1907/2006, testing for this endpoint should be performed using an appropriate route of exposure. A 28 day oral study was conducted to fulfil the subacute toxicity endpoint, since this was considered to be the most appropriate route of exposure for the substance. Furthermore dermal absorption is considered unlikely given that the substance is very lipophilic (log Pow 11.4); it is therefore considered that testing via this route is not appropriate and has been omitted on this basis.


Justification for selection of repeated dose toxicity via oral route - systemic effects endpoint:
Only one study is available.

Justification for classification or non-classification

Repeat Dose Oral Toxicity

The available data on the subacute oral toxicity of the test substance does not meet the criteria for classification according to Regulation (EC) 1272/2008 or Directive 67/548/EEC, and is therefore conclusive but not sufficient for classification.