Triphenylphosphine

Neurotoxicity

Description of key information

Neurotoxicity - oral route:

Experimental test data from several species are available, suggesting that rats are relatively insensitive towards the test substance and dogs being the most sensitive species.

Under the conditions of a 90-day combined Repeated Dose Toxicity Study with a Neurotoxicity study, the oral administration of the test item in an aqueous vehicle by gavage to Wistar rats revealed no sings of neurotoxicity up to the highest tested dose of 120 mgt/kg bw/d [BASF, 2002].

However, the oral application of the test substance suspended in an oily vehicle over 28 days caused clinical signs of neurotoxicity in rabbits with a NOAEL of <100 mg/kg bw/d [BASF, 1973] and dogs with a NOAEL of 1 mg/kg bw/d [M&T Chemicals, 1983], which were accompanied with axonal degeneration in the dog study (LOAEL 5 mg/kg bw/d).

Neurotoxicity - inhalation route:

An aerosol of the test substance in xylol produced signs of neurological impairment at 28 mg/m3 in a 4-wk inhalation study in dogs which correlated with neurotoxic tissue effects (NOAEC 10 mg/m3) [M&T Chemicals, 1983].

Neurotoxicity - dermal route:

No data available.

Key value for chemical safety assessment

Effect on neurotoxicity: via oral route

Link to relevant study records

Reference

Endpoint:

neurotoxicity

Type of information:

experimental study

Adequacy of study:

weight of evidence

Reliability:

2 (reliable with restrictions)

Rationale for reliability incl. deficiencies:

other: acceptable test method based on scientific principles with detailed documentation, only 2 animals per dose group

Reason / purpose for cross-reference:

reference to same study

Principles of method if other than guideline:

Procedure: Evaluation of effects produced by oral gavage. This study was conducted to assess the oral toxicity of the test substance when administered via gastric intubation in a corn oil vehicle to male and female dogs. Four dose levels and two different durations of treatment were used in the study; the specific test regimes were as follows: 2 dogs (1 male, 1 female) per group received 20 doses over four weeks at 1, 5, 10 and 20 milligrams test substance per kilogram (mg/kg); 2 doses were administered to 2 dogs (l male, 1 female) per group at concentrations of 1, 5 and 20 mg/kg. These shorter duration treatments were followed by a four week recovery period. A concurrent vehicle control group (1 male, 1 female) received 20 doses of corn oil, the volume was comparable to that given the test substance treated animals.
Each batch of treated corn oil was prepared fresh daily and the level of test substance in the vehicle was determined by gas chromatographic and/or high pressure liquid chromatographic methods.
Body weight measurements, physical observations and neurological examinations were performed pre-test and weekly throughout the study. Complete gross postmortem examinations were performed and selected sections of nervous system tissue were preserved from all animals for hisopathological evaluation.

GLP compliance:

not specified

Limit test:

no

Specific details on test material used for the study:

- Purity: 94.9%

Species:

dog

Strain:

Beagle

Sex:

male/female

Details on test animals or test system and environmental conditions:

TEST ANIMALS
- Age at study initiation: 6 - 7 months initiation of dosing.
- Weight at study initiation: males: 11.2 kg, females 9.3 kg.
- Housing: individually in metal grid cages.
- Identification: Each dog was assigned a unique identification number which appeared on an ear tag and on the animals cage. In addition, each animal bore an ear tattoo with a USDA number.
- Diet: standard laboratory diet (approx. 400 g/day).
- Water: city tap water ad libitum.
- Acclimation period: Yes, minimum 4 weeks (4 - 5 months at receipt and 6 - 7 months at initiation of dosing).
- Immunization: against distempter, hepatitis and leptospirosis by supplier.
- Parasites: fecal examination conducted during pretest an all animals during the equilibration period.

ENVIRONMENTAL CONDITIONS
- Photoperiod (hrs dark / hrs light): 12 hours/12 hours.

Route of administration:

oral: gavage

Vehicle:

corn oil

Details on exposure:

- Appropriate amounts of the test substance were dissolved in Mazola corn oil.
- Dosing solutions were prepared daily. Individual doses were adjusted on the basis of the most recent weekly body weight data.
- Dosing volume: 1 ml/kg across all groups.

Analytical verification of doses or concentrations:

yes

Details on analytical verification of doses or concentrations:

Sampling: 10 ml samples were taken from each dosing solution at least once a week for analytical verification of dosing solution concentrations.
Analysis of the daily dosing solutions of the test substance in corn oil was performed using a gas chromatograph.

Duration of treatment / exposure:

20 doses during 4 weeks exposure period

Frequency of treatment:

2 - 5 days/week (totally 20 doses

Dose / conc.:

1 mg/kg bw/day (nominal)

Dose / conc.:

5 mg/kg bw/day (nominal)

Dose / conc.:

10 mg/kg bw/day (nominal)

Dose / conc.:

20 mg/kg bw/day (nominal)

No. of animals per sex per dose:

1

Control animals:

yes, concurrent vehicle

Observations and clinical examinations performed and frequency:

- Animals were observed twice daily for incidence of abnormal signs.
- Observations of any unusual conditions were recorded when made.
- Full lphysical and neurological assessments were performed weekly.
- Individual body weights were recorded pretest and weekly during exposure and/or recovery period.

Sacrifice and (histo)pathology:

- Tissues fixed: brain, sciatic nerve with muscle, spinal cord
- Fixative: all tissues - 10% neutral buffered formalin.
- Histopathology: slides of tissue sections were prepared and examined microscopically from all animals; sections of all tissues were prepared with H&E stain and luxol fast blue stain.
- Sciatic nerve (right and left): 1 longitudinal secion (on each); 2 transverse sections (on each).
- Cervical spinal cord: 1 longitudinal section; 2 transverse sections
- Lumbar spinal cord: 1 longitudinal section; 2 transverse sections

Statistics:

no data

Clinical signs:

effects observed, treatment-related

Description (incidence and severity):

No physical signs of toxicity but neurological observations revealed the following signs: At 5, 10, and 20 mg/kg (20 exposures), male dogs showed indications of neurological impairment: failure in the extensor postural thrust reflex with tendency to knuckle over, unsteadiness and weakness in the hind limbs, failure of the patellar reflexes. Indications of similar neurological lesions were found in two female dogs having received 20 doses of 1 and 2 doses of 5 mg/kg, but not at the higher doses. The effects noted in male dogs were considered treatment-related as no such signs were observed in the controls and the other groups. The cause of the effects in the low-dosed female dogs was unclear.

Mortality:

no mortality observed

Description (incidence):

All animals survived.

Body weight and weight changes:

no effects observed

Description (incidence and severity):

Body weight development was normal.

Gross pathological findings:

no effects observed

Description (incidence and severity):

No treatment-related post-mortem findings.

Neuropathological findings:

effects observed, treatment-related

Description (incidence and severity):

The sections prepared from male and female dogs after 20 exposures to 10 or 20 mg/kg bw revealed mild to moderate generalised vacuolative axonal lesion in the ventral and lateral funicule of the spinal cord. No such findings in the sciatic nerve.
Only isolated damaged axons were observed in other groups including controls. Also in a histopathological reevaluation compound related axonal degeneration was evident only at 10 and 20 mg/kg bw.

Dose descriptor:

NOAEL

Effect level:

1 mg/kg bw/day (nominal)

Sex:

male/female

Dose descriptor:

LOAEL

Effect level:

5 mg/kg bw/day (nominal)

Sex:

male/female

Basis for effect level:

clinical signs

Critical effects observed:

yes

Lowest effective dose / conc.:

5 mg/kg bw/day (nominal)

System:

nervous system

Treatment related:

yes

Dose response relationship:

yes

Relevant for humans:

yes

Executive summary:

This study was conducted to assess the oral toxicity of the test substance when administered via gastric intubation in a corn oil vehicle to male and female dogs.Four dose levels and two different durations of treatment were used in the study; the specific test regimes were as follows: 2 dogs (1 male, 1 female) per group received 20 doses over four weeks at 1, 5, 10 and 20 milligrams test substance per kilogram (mg/kg); 2 doses were administered to 2 dogs (l male, 1 female) per group at concentrations of 1, 5 and 20 mg/kg. These shorter duration treatments were followed by a four week recovery period. A concurrent vehicle control group (1 male, 1 female) received 20 doses of corn oil, the volume was comparable to that given the test substance treated animals.

Each batch of treated corn oil was prepared fresh daily and the level of test substance in the vehicle was determined by gas chromatographic and/or high pressure liquid chromatographic methods.

Body weight measurements, physical observations and neurological examinations were performed pre-test and weekly throughout the study. Complete gross postmortem examinations were performed and selected sections of nervous system tissue were preserved from all animals for hisopathological evaluation.

All animals survived the duration of the study. Signs of neurological impairment were noted in male dogs receiving 20x 5, 10 or 20 mg/kg test substance doses. Similar findings were noted in two female dogs which received 1 or 5 mg/kg doses for twenty or two treatments, respectively; however, these findings were not noted in female dogs receiving either two or twenty treatments at higher dose levels.

Microscopic pathology evaluations (H&E, and luxol fast blue stains) revealed mild to moderate generalized vacuolative axonal lesions in the ventral and lateral funicle of the spinal cord at both sexes of dogs receiving twenty doses of 20 mg/kg. Similar lesions, although minimal and focal in nature, were noted in spinal cord sections in dogs receiving 1 or 10 mg/kg test substance. In

the group receiving 1 mg/kg (2 or 20 doses) only females exhibited this change. These lesions were not seen in any dog receiving only corn oil.

Evaluations of body weight, general physical observations (non-neurological) and gross postmortem results were considered unremarkable.

Endpoint conclusion

Endpoint conclusion:

adverse effect observed

Dose descriptor:

NOAEL

1 mg/kg bw/day

Study duration:

subacute

Species:

dog

Effect on neurotoxicity: via inhalation route

Link to relevant study records

Reference

Endpoint:

neurotoxicity

Type of information:

experimental study

Adequacy of study:

weight of evidence

Reliability:

2 (reliable with restrictions)

Rationale for reliability incl. deficiencies:

other: acceptable test method based on scientific principles with detailed documentation, only 2 animals per dose group

Reason / purpose for cross-reference:

reference to same study

Principles of method if other than guideline:

This study was conducted to assess the inhalation toxicity of the test substance when administered as a respirable aerosol in a xylene vehicle to male and female beagle dogs. Four atmospheric concentration levels and two different durations of treatment were used in the study; the specific test regimes were as follows: two dogs (1 male, 1 female) per group received twenty 6-hour exposures over four weeks at exposure levels of 0.5, 3.2. 9.7 and 28 milligrams test substance per meter cubed (mg/m3); two 6-hour exposures were administered to two dogs (1 male, l female) per group at concentrations of 0.3, 3.6 and 24 mg/m3 test substance. These shorter duration treatments were followed by a four week recovery period. A concurrent vehicle control group (1 male, 1 female dog) received twenty 6-hour exposures to150 parts per million (v/v in air) of xylene, a level of solvent which was comparable to that in each test exposure chamber. Chambers were monitored analytically for test substance levels at least 3 times/chamber/day. Particle size distribution were determined periodically throughout the study. Body weight measurements, physical observations and neurological examinations were performed pre-test and weekly throughout the study. Complete gross postmortem examinations were performed and selected sections of nervous system tissue were preserved from all test animals for histopathological evaluation

GLP compliance:

not specified

Limit test:

no

Specific details on test material used for the study:

- Purity 94.9%
- Expiry Date: December, 1982.
- Storage Infonnatfon: 60 - 85°F.

Species:

dog

Strain:

Beagle

Sex:

male/female

Details on test animals or test system and environmental conditions:

TEST ANIMALS
- Source: Marshall Research Animals, Inc. North Rose, New York 14516
- Age at study initiation: 6-7 months.
- Weight at study initiation: 9.6-13.2 kg in males and 8.2-10.4 kg in females

Route of administration:

inhalation: aerosol

Vehicle:

other: xylene

Mass median aerodynamic diameter (MMAD):

< 0.5 µm

Details on exposure:

- Whole-body exposure using chambers of 10 m3 which received an aerosol produced from the test substance dissolved in xylene (complete air change in the chamber every 3.3 minutes).
- Chambers were monitored analytically for test substance levels 3x/d per chamber and particle-size distribution periodically.
- The count medium diameter of the aerosol particles was found to be 0.5 um or less, i.e. respirable.

Analytical verification of doses or concentrations:

yes

Details on analytical verification of doses or concentrations:

- Analytical concentrations of 0, 0.5, 3.2, 9.7, 28 mg/m3 (as aerosol in xylene, aerosol particles 0.5 um) or for only 2 times to 0.5, 3.2 or 28 mg/m3 (post exposure observation period 30 days).
- The concentration of xylene was 150 ppm or less in all chambers (solvent not specified, isomer mixture??).

Duration of treatment / exposure:

5 weeks

Frequency of treatment:

3 - 5 days/week, 6 hours/day (20 exposures)

Dose / conc.:

0.5 mg/m³ air (nominal)

Remarks:

as aerosol in xylene

Dose / conc.:

3.2 mg/kg bw/day (nominal)

Remarks:

as aerosol in xylene

Dose / conc.:

9.7 mg/kg bw/day (nominal)

Remarks:

as aerosol in xylene

Dose / conc.:

28 mg/kg bw/day (nominal)

Remarks:

as aerosol in xylene

No. of animals per sex per dose:

1

Control animals:

yes, concurrent vehicle

Observations and clinical examinations performed and frequency:

Body weight determined once weekly. Animal observation twice daily, full physical assessment once weekly.

Neurobehavioural examinations performed and frequency:

Neurological assessment once weekly.

Sacrifice and (histo)pathology:

Dogs were sacrificed and necropsy performed. Histopathologic examination was focussed on the spinal cord and sciatic nerve.

Statistics:

no data

Clinical signs:

effects observed, treatment-related

Description (incidence and severity):

At 28 mg/m3 (20 exposures), both dogs showed indications of neurological impairment: failure in the extensor postural thrust reflex with tendency to knuckle over, unsteadiness and weakness in the hind limbs. (This effect was already noted in one male dog having received only two exposures.) The locomotor reflex in the male dog appeared also unsteady and uncoordinated (ataxia).
At 10 mg/m3 erratic placement of limbs on week 1 and 4 was observed in one male dog.
In one male receiving only 2 exposures of 3 mg/m3 some limb placement incoordination throughout the study was seen.
Only the effects at the highest dose were considered by the authors to be treatment-related (in other dose groups effects not persistent, no severe signs, animals were only exposed twice).

Mortality:

no mortality observed

Description (incidence):

All animals survived.

Body weight and weight changes:

no effects observed

Description (incidence and severity):

Body weight development was normal.

Neuropathological findings:

effects observed, treatment-related

Description (incidence and severity):

The sections prepared from the high-dosed dogs revealed moderate generalised vacuolative degeneration of axons in the ventral and lateral funicule of both the cervical and lumbar spinal cord. No such findings in the sciatic nerve. No such lesions were noted in the control animals having received xylene alone.
The above mentioned neuronal lesions were seen in the longitudinal sections, recognised with both H&E and luxol fast blue stains, characterised by multiple dilated or vacuolated axon sheaths with accumulations of scattered eosinophilic debris and occasional dark rounded bits of what appeared to have been nuclear chromatin.
Similar damages in the lower dose groups, but marginal in nature, were not considered to correlate with test substance exposure since in a histopathological reevaluation also historical control animals showed some axonal lesions. Sporadic degeneration is an expected finding in the spinal cord of "normal" animals.

Dose descriptor:

NOAEC

Effect level:

10 mg/m³ air (nominal)

Sex:

male/female

Dose descriptor:

LOAEC

Effect level:

28 mg/m³ air (nominal)

Sex:

male/female

Basis for effect level:

clinical signs

Critical effects observed:

yes

Lowest effective dose / conc.:

28 mg/m³ air

System:

nervous system

Treatment related:

yes

Dose response relationship:

yes

Relevant for humans:

yes

Executive summary:

This study was conducted to assess the inhalation toxicity of the test substance when administered as a respirable aerosol in a xylene vehicle to male and female beagle dogs.

 

Four atmospheric concentration levels and two different durations of treatment were used in the study; the specific test regimes were as follows: two dogs (1 male, 1 female) per group received twenty 6-hour exposures over four weeks at exposure levels of 0.5, 3.2. 9.7 and 28 milligrams test substance per meter cubed (mg/m3); two 6-hour exposures were administered to two dogs (1 male, 1 female) per group at concentrations of 0.3, 3.6 and 24 mg/m3 test substance. These shorter duration treatments were followed by a four week recovery period. A concurrent vehicle control group (1 male, 1 female dog) received twenty 6-hour exposures to 150 parts per million (v/v in air) of xylene, a level of solvent which was comparable to that in each test exposure chamber.

Chambers were monitored analytically for test substance levels at least 3 times/chamber/day. Particle size distribution were determined periodically throughout the study. Body weight measurements, physical observations and neurological examinations were performed pre-test and weekly throughout the study. Complete gross postmortem examinations were performed and selected sections of nervous system tissue were preserved from all test animals for histopathological evaluation.

All animals survived the duration of the study. Signs of neurological impairment were noted during the study in male dogs receiving 2 exposures at 24 mg/m3 test substance or 20 exposures at 28 mg/m3 test substance and in the female dogs receiving 20 exposures at 28 mg/m3 test substance.

Histopathological examinations"(H&E, and luxol fast blue stains) revealed moderate generalized vacuolative degeneration of the ventral and lateral funicle of the cervical and lumbar spinal cord of the dogs which received twenty exposures to 28 mg/m3 test substance. Similar lesions, although minimal and focal in nature were noted in spinal cord sections of dogs receiving 0.5 or 3.2 mg/m3test substance. In the lowest treatment group animals, this lesion was observed in one male following 20 exposures and in·one female following only two exposures. These lesions were not seen in any vehicle control dog. Evaluations of body weight, general physical observations (non-neurological) and gross postmortem results were considered unremarkable.

Endpoint conclusion

Endpoint conclusion:

adverse effect observed

Dose descriptor:

NOAEC

10 mg/m³

Study duration:

subacute

Species:

dog

Effect on neurotoxicity: via dermal route

Endpoint conclusion

Endpoint conclusion:

no study available

Additional information

Neurotoxicity - oral route:

Experimental test data from several species are available, suggesting that rats are relatively insensitive towards the test substance and dogs being the most sensitive species. Therefore, the endpoint conclusion is based on the experimental data on dogs. Moreover, the vehicle had an influence on the toxicity of the test substance since oily suspensions produced adverse effects at lower concentrations than aqueous suspensions, presumably due to higher bioavailability.

BASF (2002) reported a 3-month neurotoxicity gavage study according to OECD guideline 408 testing aqueous suspension of the test substance with 0.5% CMC. Male and female rats were given doses of 6, 60 or 120 mg/kg over a study period of 91 days . The oral administration of the test substance caused changes associated with induction of the microsomal enzyme system in the liver, increased liver weights and centrilobular hypertrophy of hepatocytes. Decreases in alanine and aspartate aminotransferase activities were seen. This fall in alanine and aspartate aminotransferase is considered not to be an adverse toxic effect. No signs of neurotoxicity were observed up to the highest tested dose. The NOAEL was found to be 6 mg/kg and the LOAEL was found to be 60 mg/kg for male and female rats.

BASF (2003) reported a neurotoxicity screening study on rats. Male rats were given orally (gavage) doses of 700 and 2100 mg/kg bw over a study period of 28 days. No neuropathological changes in the central and peripheral nervous system were obtained, whereas during clinical examinations and concerning clinical pathology severe signs of general toxicity were observed.

In the course of a 28-day repeated dose toxicity study by BASF (2003) the test substance was administered to groups of 3-4 rabbits in the form of A) oily suspensions as well as B) aqueous suspensions in carboxymethyl cellulose (CMC) in doses of 100, 200 and 400 mg/kg bw/d or 100, 200, 400 and 800 mg/kg bw/d, respectively. Repeated applications of the test substance produced hyperexcitability, convulsions with subsequent atonia, lataral decubitus and unsteady locomotion from 100 mg/kg or 200 mg/kg bw/day onwards, when the oily or aqueous vehicle was used, respectively. Mortality was observed from 200 mg/kg onwards with both vehicles. The degree of the symptoms of neurological impairment was dose- and vehicle-dependent. The test substance appeared to be absorbed more readily from the oily phase than from the aqueous phase in the subacute experiment similar to the acute studies. Repeated doses of the test substance led to a transient deterioration of the red blood cell count as well as of liver and kidney function, and sometimes also of the differential white blood cell count. However, these changes were reversible even during the study or during the four-week followup period. After doses of 800 and 400 mg/kg, the animals which died showed signs of heart enlargement and in several cases, fine necrotic foci and fatty degeneration of the liver.

M&T Chemicals Inc. (1983) reported a 4-week oral toxicity study using dogs in which the test substance was applied in corn oil. Doses used were 1, 5, 10 or 20 mg/kg bw/day. All animals survived; body weight development was normal. No physical signs of toxicity but neurological observations revealed the following signs: At 5, 10, and 20 mg/kg (20 exposures), male dogs showed indications of neurological impairment: failure in the extensor postural thrust reflex with tendency to knuckle over, unsteadiness and weakness in the hind limbs, failure of the patellar reflexes. Similar signs of neurological impairment were found in two female dogs having received 20 doses of 1 mg/kg and 2 doses of 5 mg/kg, but not at the higher doses. The effects noted in male dogs were considered treatment-related as no such signs were observed in the controls and the other groups. The cause of the effects in the low-dosed female dogs was unclear. Macroscopic pathology: No treatment-related post-mortem findings. Histopathology: The sections prepared from male and female dogs after 20 exposures to 10 or 20 mg/kg bw revealed mild to moderate generalised vacuolative axonal lesion in the ventral and lateral funicule of the spinal cord. No such findings in the sciatic nerve. Only isolated damaged axons were observed in other groups including controls. In a histopathological reevaluation compound related axonal degeneration was evident only at 10 and 20 mg/kg bw. The NOAEL was 1 mg/kg bw/day for male and female dogs (based on clinical signs of ataxia/impairment of reflexes).

BASF reported an acute delayed neurotoxicity GLP-study in hen according to the EPA guideline 163 (1978) (BASF, 1981). Ten animals per group were gavaged with 0; 1500; 3000; 6000; or 9000 mg/kg bw test substance in corn oil and observed for 21 days post treatment. Ten animals dosed with 500 mg/kg bw of tri-ortho-cresyl phosphate (TOCP) were used as positive control. No test substance-treated animal died. The treatment had no effect on food consumption and body weights. There was no clear evidence of ataxia at a dose of 1500 mg/kg bw and no signs of ataxia at 3000 mg/kg bw, but clearly treatment-related neuropathological changes in the spinal cord were found even at the low dose level. Severe neuropathological morphological changes (i.e. treatment-related changes mainly in the spinal cord but also in the sciatic nerve) occurred in birds of the high-dose groups (6000 and 9000 mg/kg bw). First signs of ataxia appeared at day 5 after administration in four birds given 6000 mg/kg bw and three birds given 9000 mg/kg bw. Doubtful (grade 1) signs of ataxia were observed in one bird given 1500 mg/kg bw, three birds given 6000 mg/kg bw, and one bird given 9000 mg/kg bw. The ataxia in birds given the test substance was unusual in that the first signs were observed on day 5 and there was an apparent improvement towards the end of the observation period. All birds given TOCP developed positive signs of ataxia, the first sign occurring on day 8, with one bird being sacrificed since it was in a moribund state.

Neurotoxicity - inhalation route:

An aerosol of the test substance in xylol produced signs of neurological impairment at 28 mg/m3 in a 4-wk inhalation study in dogs which correlated with neurotoxic tissue effects (NOAEC 10 mg/m3) [M&T Chemicals, 1983]. This study was conducted to assess the inhalation toxicity of the test substance when administered as a respirable aerosol in a xylene vehicle to male and female beagle dogs. Four atmospheric concentration levels and two different durations of treatment were used in the study; the specific test regimes were as follows: two dogs (1 male, 1 female) per group received twenty 6-hour exposures over four weeks at exposure levels of 0.5, 3.2. 9.7 and 28 milligrams test substance per meter cubed (mg/m3); two 6-hour exposures were administered to two dogs (1 male, 1 female) per group at concentrations of 0.3, 3.6 and 24 mg/m3 test substance. These shorter duration treatments were followed by a four week recovery period. A concurrent vehicle control group (1 male, 1 female dog) received twenty 6-hour exposures to 150 parts per million (v/v in air) of xylene, a level of solvent which was comparable to that in each test exposure chamber. Chambers were monitored analytically for test substance levels at least 3 times/chamber/day. Particle size distribution were determined periodically throughout the study. Body weight measurements, physical observations and neurological examinations were performed pre-test and weekly throughout the study. Complete gross postmortem examinations were performed and selected sections of nervous system tissue were preserved from all test animals for histopathological evaluation. All animals survived the duration of the study. Signs of neurological impairment were noted during the study in male dogs receiving 2 exposures at 24 mg/m3 test substance or 20 exposures at 28 mg/m3 test substance and in the female dogs receiving 20 exposures at 28 mg/m3 test substance. Histopathological examinations"(H&E, and luxol fast blue stains) revealed moderate generalized vacuolative degeneration of the ventral and lateral funicle of the cervical and lumbar spinal cord of the dogs which received twenty exposures to 28 mg/m3 test substance. Similar lesions, although minimal and focal in nature were noted in spinal cord sections of dogs receiving 0.5 or 3.2 mg/m3test substance. In the lowest treatment group animals, this lesion was observed in one male following 20 exposures and in one female following only two exposures. These lesions were not seen in any vehicle control dog. Evaluations of body weight, general physical observations (non-neurological) and gross postmortem results were considered unremarkable.

Neurotoxicity - dermal route:

No data available.

Neurotoxicity - other routes:

The aim of the study was to determine the biochemical, neuropathological, and clinical effects of single doses of the test substance in the European ferret, a mammalian species shown to be susceptible to organophosphorus-induced neurotoxicity. Eight 12-week-old ferrets were each injected subcutaneously with either 250 or 500 test substance/kg bw, or with the peanut oil/ethyl ether vehicle. Twenty-four hours after dosing, the brains of 5 animals from each dose group were examined for NTE and AChE activities. The remaining 3 animals in each group were observed for 6 days for the development of clinical signs, after which their brains were processed for the presence of axonal degeneration using the Fink-Heimer silver impregnation method. Single injections of the test substance had no effect on the activities of whole-brain NTE or AChE 24 h after injection, The animals observed for clinical signs showed increasing trunk and hindlimb ataxia beginning 4 days after injection, culminating in foreand hindlimb paralysis 6 days after injection . All brains exposed to either dose of the test substance showed widespread axonal degeneration extending from the brainstem and cerebellum into midbrain and forebrain areas.

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. Clinical signs of neurological impairment and neuropathological effects were seen in the oral and inhalation studies depending on the species and the vehicle used. Based on a worst case consideration, the substance is considered to be classified for repeated dose toxicity STOT RE Cat 1 under Regulation (EC) No. 1272/2008, as amended for the tenth time in Regulation (EU) No 2017/776.