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Please be aware that this old REACH registration data factsheet is no longer maintained; it remains frozen as of 19th May 2023.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

Use of this information is subject to copyright laws and may require the permission of the owner of the information, as described in the ECHA Legal Notice.

REACH

N-[[3,5-dichloro-2-fluoro-4-(1,1,2,3,3,3-hexafluoropropoxy)phenyl]carbamoyl]-2,6-difluorobenzamide

EC number: 601-779-5 | CAS number: 121451-02-3

Life Cycle description
Uses advised against

Toxicological information

Neurotoxicity

Currently viewing:

Administrative data

Description of key information

NOEL > 300 mg/kg bw/day (rat) male/female, Maurissen et. al. (2004)

Key value for chemical safety assessment

Effect on neurotoxicity: via oral route

Link to relevant study records

Reference

Endpoint:

neurotoxicity: chronic oral

Type of information:

experimental study

Adequacy of study:

key study

Study period:

02 February 2002 - 20 February 2004

Reliability:

1 (reliable without restriction)

Rationale for reliability incl. deficiencies:

guideline study

Reason / purpose for cross-reference:

reference to same study

Qualifier:

according to guideline

Guideline:

other: EPA OPPTS 870.6200 (Neurotoxicity Screening Battery) and EPA OPPTS 870.4300 (Combined Chronic Toxicity / Carcinogenicity)

Deviations:

Qualifier:

according to guideline

Guideline:

other: OECD Guideline 424 (Neurotoxicity Study in Rodents) and OECD Guideline 453 (Combined Chronic Toxicity / Carcinogenicity Studies)

Deviations:

Qualifier:

according to guideline

Guideline:

other: EU Method Part B. (Combined Chronic Toxicity / Carcinogenicity Test)

Deviations:

Qualifier:

according to guideline

Guideline:

other: JMAFF, Combined Chronic Toxicity/Oncogenicity Study and Subchronic Oral Toxicity Study) 1985

Deviations:

GLP compliance:

yes

Limit test:

Species:

rat

Strain:

Fischer 344

Sex:

male/female

Details on test animals or test system and environmental conditions:

TEST ANIMALS
- Age at study initiation: 6 weeks
- Housing: The animals were housed 2-3 per cage during acclimatisation and two per cage during the main study in stainless steel cages.
- Diet: Certified canine diet in meal form, provided ad libitum.
- Water: Municipal water, provided ad libitum.
- Acclimation period: Animals were acclimated to the laboratory for one week prior to the start of the study.

ENVIRONMENTAL CONDITIONS
- Temperature (° C): 22 ± 1 °C (with a maximum permissible excursion range of ± 3°C)
- Humidity (%): 40 to 70 %
- Air changes (per hr): approximately 12-15 times/hour
- Photoperiod (hrs dark / hrs light): A 12-hour light/dark photocycle was maintained for all animal rooms with lights on at 6:00 a.m. and off at 6:00 p.m.

Route of administration:

oral: feed

Vehicle:

unchanged (no vehicle)

Details on exposure:

DIET PREPARATION
Diets were prepared by serially diluting a concentrated test material- feed mixture (premix) with ground feed. Premixes were mixed periodically throughout the study based on stability data. Initial concentrations of test material in the diet were calculated from historical body weights and feed consumption data. Subsequently, the concentrations of the test material in the feed were adjusted weekly for the first 13-weeks of the study and at 4-week intervals thereafter, based upon the most recent body weight and feed consumption data.

Analytical verification of doses or concentrations:

yes

Details on analytical verification of doses or concentrations:

- Homogeneity: The homogeneity of the low-dose female and high-dose male test material- feed mixtures were determined prior to the start of dosing and at approximately 1.5, 3, 8, 12, 18, and 24 months. Additional analytical analyses were conducted at approximately 8, 9, 14, and 20 months to verify homogeneity of the diet mixes.
- Stability: Previous 90-day toxicity study in rats (Yano and Dryzga, 2002) demonstrated that the test material was stable for at least 42 days in rodent chow at concentrations ranging from 0.005 % to 5 %. In this study, an additional stability evaluation was conducted at 0.0001 % to cover the concentration ranges of the administered diets.
- Concentration Verification: Analyses of the premix, all dose levels and the 0 (control) mg/kg bw/day diets were determined prior to the start of exposure and at approximately 1.5, 3, 8, 12, 18, and 24 months. Additional concentration analyses were also conducted at approximately 14, 18, and 20 months. The method for analysing the test material in feed was a solvent extraction method followed by analyses using liquid chromatography- mass spectrometry (LC-MS) and solvent standards incorporating an internal standard.
- Retainer Samples: Reference samples (one/sex/dose/mix) were retained and stored in sealed vials in a manner consistent with the sample retention policy of the laboratory.

Duration of treatment / exposure:

One year.

Frequency of treatment:

Daily.

Dose / conc.:

0 mg/kg bw/day

Remarks:

Basis:
nominal in diet

Dose / conc.:

0.1 mg/kg bw/day

Remarks:

Basis:
nominal in diet

Dose / conc.:

1 mg/kg bw/day

Remarks:

Basis:
nominal in diet

Dose / conc.:

75 mg/kg bw/day

Remarks:

Basis:
nominal in diet

Dose / conc.:

300 mg/kg bw/day

Remarks:

Basis:
nominal in diet

No. of animals per sex per dose:

Ten per sex per dose.

Control animals:

yes, plain diet

Details on study design:

- Dose selection rationale: Selected dose levels for the combined chronic toxicity/oncogenicity/neurotoxicity study were 0, 0.1, 1.0, 75, and 300 mg/kg bw/day, and these dose levels were approved by the USEPA. The high dose was selected based on the data from the 90-day dietary toxicity study and the limited pharmacokinetic study. Data from these studies indicated that GI absorption was saturated at an actual dose of 78-86 mg/kg bw (targeted concentration of 100 mg/kg bw) and this conclusion was supported by numerous toxicology data points, especially liver weights. The highest-dose level was selected at 300 mg/kg bw/day rather than 100 mg/kg bw/day because, although the dose was not proportional to toxicity or the AUCs, additional test material was absorbed at doses higher than 100 mg/kg bw/day.
Therefore, a conservative approach to dose level selection was to challenge the rats to this higher dose level. Treatment-related effects induced in rats given 300 mg/kg bw/day were expected to be as follows: 1) slight reduction in body weight, 2) increased liver weights, and 3) possible alterations in clinical pathology parameters. Similar effects of a lesser magnitude were expected to occur in rats given 75 mg/kg bw/day.
- Dosing route rationale: Probable routes of human exposure to the test material are via accidental ingestion during application or manufacture. Thus, administration of the test material via the diet represents an appropriate means of exposure.
- Rationale for animal assignment: Animals were stratified by pre-exposure body weight and then randomly assigned to treatment groups using a computer program.

Observations and clinical examinations performed and frequency:

CAGE SIDE OBSERVATIONS: Yes
- Time schedule: A cage-side (general) clinical examination was conducted at least once a day, typically at the same time each day (usually in the morning). Moribund animals not expected to survive until the next observation periods were humanely euthanized that day. In addition, all animals were observed for morbidity, mortality, and the availability of feed and water at least twice daily.
- Cage side observations included, but not limited to: Decreased/increased activity, repetitive behaviour, vocalization, incoordination/limping, injury, neuromuscular function (convulsion, fasciculation, tremor, twitches), altered respiration, blue/pale skin and mucous membranes, severe eye injury (rupture), alterations in faecal consistency, and faecal/urinary quantity.

BODY WEIGHT: Yes
- Time schedule for examinations: Body weights were recorded prior to each FOB session.

FOOD CONSUMPTION AND COMPOUND INTAKE: Yes
- Food consumption: Feed consumption data were collected weekly during the first 13 weeks of the study and then at approximately monthly intervals thereafter for all animals. Feed containers were weighed at the start and end of a measurement cycle and consumption was calculated using the following equation:
Feed consumption (g/day) = (initial weight of feed containers - final weight of feed containers) / [(No. of days in measurement cycle)(No. of animals per cage)]
- Test material intake (TMI): TMI was calculated using test material concentrations in the feed, actual body weights (BW) and feed consumption using the following equation:
TMI = [(feed consumption) * (1000) * (% of test material in feed/100)] / {[(current BW + previous BW) / 2] / 1000}

FOOD EFFICIENCY:
Feed efficiency was calculated using body weight gain and feed consumption data from the first 13 weeks of the study using the following equation:
Feed efficiency = (g feed consumed/day) / (g body weight gain/day)

OPHTHALMOSCOPIC EXAMINATION: Yes
- Time schedule for examinations: The eyes of all animals were examined by a veterinarian pre-exposure and prior to the scheduled necropsy using indirect ophthalmoscopy.
- Dose groups that were examined: All dose groups were examined.
- Method: One drop of 0.5 % tropicamide ophthalmic solution was instilled in each eye to produce mydriasis prior to the indirect ophthalmic examinations. Eyes were also examined by a prosector during necropsy using a moistened glass slide pressed to the cornea.

Neurobehavioural examinations performed and frequency:

FUNCTIONAL OBSERVATIONAL BATTERY: Yes
- Parameters checked: The functional observational battery (FOB) included hand-held and open-field observations, measurements of hind- and forelimb grip performance, hind limb landing, foot splay, and rectal temperature.
> Hand-Held and Open-Field Observations:
Hand-held and open-field observations included a careful physical examination and sensory evaluation according to an established format. Open-field observations and sensory evaluations were made in a clear plastic box (50 cm x 50 cm). Observations were dictionary based, and contained most of the common physical and neurologic abnormalities seen in toxicity studies. Because not all potential observations are contained in the dictionary, free- field descriptions were also allowed.
Cage-side evaluation included: Unusual body movements (e.g. tremors, convulsions), abnormal behaviour (e.g. circling, stereotypy) and posture, and resistance to removal (a ranked observation).
> Grip Performance:
Hind limb grip performance was tested according to the procedure described by Mattsson et al. (1986). Briefly, the observer placed the animal’s forelegs on a plastic bench and the hind feet were set on a horizontal screen attached to an electronic strain gauge (Cha tillon, Greensboro, North Carolina). The observer then smoothly but firmly pulled backward on the tail until the animal’s grip on the screen was broken. An electronic strain gauge reading was used to record the animal’s resistance to the pull in grams. The average of three trials was used for statistical analysis. Forelimb grip performance was similarly tested, except that a bench was not used. The animal was placed so that the forefeet were on the screen and the hind feet were suspended approximately 10 cm above the smooth horizontal plastic surface.
Instrument Calibration: A standard 500 g weight attached to a fine gauge wire was suspended from the load cell and the resulting mass was recorded in grams. A 1 % tolerance (i.e., 500 ± 5 g) was acceptable and was checked just before and just after testing.
> Landing Foot Splay:
The landing foot splay procedure was similar to that published by Edwards and Parker (1977). A form containing the study, animal identification, and date was used to capture the data. The outer-most toe of each hind foot was marked with ink. The animal was then dropped from a height of 30 cm onto the recording sheet. This procedure was repeated three times, and the toes were re-inked as necessary before each trial. The distance from centre-to-centre of the ink marks for each trial was measured (cm), and the average of the three splay values were used for statistical analysis.
> Rectal Temperature:
Rectal temperature was measured with a rectal thermistor that was calibrated at 37 °C before and after the study. The instrument was recalibrated if the temperature recordings differed from the reference thermometer by more than ± 0.5 °C. The thermistor was placed approximately 4 cm into the rectum for about 10-15 seconds. Temperature was then recorded.

MOTOR ACTIVITY: Yes
Twenty-four motor activity chambers (also referred to as cages), visually isolated from each other, were located in a quiet, dimly lit room. Each motor activity cage consisted of a clear plastic circular alley similar to the one developed by Richelle et al. (1967) and Fontaine et al. (1966). An infrared photobeam bisected the cage so that the beam crossed the alley in two locations. Each animal was tested individually and no entry into the MA test room was allowed during the testing period. Each test session consisted of six 8 minute epochs, totalling 48 minutes of testing per animal per test session. This duration was chosen based on the results of validation studies indicating that activity counts of control animals approached asymptote within 30-40 minutes in Fischer 344 rats (Marable and Andrus, 2001). Activity counts for each epoch were recorded. Each beam break lasting more than 100 msec constituted an activity count. This minimum duration was set to discount activities such as tail-flicking, rearing, head-bobbing, etc. A positive control report for motor activity was used.
> Motor Activity Cage Calibration: Cages used for testing were calibrated prior to testing each day. Calibration was performed with a rod attached to a rotary motor that broke the infrared beam at a constant speed. The duration of each beam break was calculated to ensure equivalence across chambers.
> Motor Activity Cage Allocation: Rats were allocated to the motor activity cages such that counterbalancing of treatment groups and sexes across cages and test times was maximized.

Sacrifice and (histo)pathology:

NECROPSY/GROSS PATHOLOGY
Five randomly pre-selected rats/sex/group were evaluated for neuropathologic effects. Rats fasted and submitted alive for necropsy were given an intraperitoneal injection of 0.2 mL heparin (10,000 USP/mL) per 100 grams body weight approximately 10 minutes prior to perfusion. Rats were anesthetised by inhalation of isoflurane vapours. While under deep anaesthesia, the heart was exposed, the left ventricle cannulated, and the right atrium was incised. Rats were perfused by gravity pressure with 0.05 M phosphate buffer containing sodium nitrite followed by a phosphate-buffered fixative solution of 1.5 % glutaraldehyde – 4 % formaldehyde (c. 540 mOs).
Tissues were examined for gross pathologic alterations by a veterinary pathologist assisted by a team of trained individuals. The brain, head, spinal column with spinal cord, fore- and hind limbs, and tail were trimmed to remove excessive skin and muscle; muscles from the hind limbs were reflected to further expose the nerves. All tissues were immersed in the glutaraldehyde/formaldehyde fixative. Transponders were removed and placed in jars with tissues.
In addition, thoracic and abdominal viscera were collected and preserved in the glutaraldehyde/formaldehyde fixative. These tissues were disposed of after the issuance of this report as it was determined that their retention or further examination were not warranted.

HISTOPATHOLOGY
Tissues for neuropathologic evaluation were prepared from all rats in the control and high-dose groups. Nine cross-sections of the brain were prepared to include the following regions: olfactory bulb, cerebrum (frontal, parietal, temporal and occipital areas), thalamus/hypothalamus, midbrain, pons, cerebellum, and medulla oblongata. In addition, sections were prepared from the trigeminal ganglion and nerve, pituitary gland, eyes with optic nerves, spinal cord (cervical and lumbar), olfactory epithelium, and skeletal muscles (gastrocnemius and anterior tibial). These tissues were processed by standard histologic procedures, embedded in paraffin, sectioned approximately six µm thick and stained with hematoxylin and eosin. Spinal nerve roots (cervical and lumbar), dorsal root ganglia (cervical and lumbar), and peripheral nerves (sciatic, tibial [proximal and distal - at the knee and calf muscle branches] and sural) were osmicated, embedded in epoxy resin, sectioned approximately 2-3 µm thick and stained with toluidine blue.
Tissues were evaluated by a veterinary pathologist using a light microscope. Histopathologic findings were subjectively graded as appropriate to assess the potential effects of treatment with the test material with regard to the contribution of a specific lesion to the health status of an animal. Very slight and slight grades were used for conditions present in excess of the normal textbook appearance of an organ/tissue, but were of minimal severity and were not expected to significantly affect the function of the specific organ/tissue involved nor have a significant effect on the overall health of the animal. Lesions of these severities involved only a minor portion of the affected organ and the two grades were differentiated based upon the frequency that the change was noted.
Lesions of moderate severity involved up to 30 % of the parenchyma and may have had an effect upon the function of the organ but not to the extent that it was considered likely to result in clinical manifestations of disease. Categories of severe or very severe were available for potential use for lesions of greater severity.

Statistics:

Data interpretation was guided by the following statement of the ICH Harmonized Tripartite Guideline (European Community, Japan, U.S.A.): "Significance tests (inferential statistics) can be used only as a support for the interpretation of results. The interpretation itself must be based on biological plausibility. It is unwise to assume that a difference from control values is not biologically relevant simply because it is not ‘statistically significant.’ To a lesser extent it can be unwise to assume that a ‘statistically significant’ difference must be biologically relevant" (ICH, 1993).
See "Any other information on materials and methods incl. tables" for details on the statistical analysis employed in this study.

Clinical signs:

no effects observed

Description (incidence and severity):