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Toxicological information

Neurotoxicity

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Administrative data

Description of key information

Hydrocarbons, C9 Aromatics:

Acute CNS effects: exposures to levels below approximately 1000 mg/m3 are unlikely to produce profound acute CNS effects.

 

Sub-chronic CNS effects: NOAEC = 1500 ppm for neurotoxicity.

 

Hydrocarbons, C10 Aromatics:

Acute CNS effects: NOAEC for in rats: 600 mg/m3 (based primarily on volatility). 

Key value for chemical safety assessment

Effect on neurotoxicity: via oral route

Endpoint conclusion
Endpoint conclusion:
no study available

Effect on neurotoxicity: via inhalation route

Link to relevant study records

Referenceopen allclose all

Endpoint:
neurotoxicity: sub-chronic inhalation
Type of information:
experimental study
Adequacy of study:
key study
Study period:
1993
Reliability:
2 (reliable with restrictions)
Rationale for reliability incl. deficiencies:
other: Acceptable well-documented publication which meets basic scientific principles.
Principles of method if other than guideline:
Groups of 20 male rats were exposed via inhalation to 100, 500, or 1500 ppm high flash aromatic naphtha for 6 hrs per day, 5 days per week, for 13 weeks. Neurotoxicity testing was performed at 5, 9, and 13 weeks of exposure. Animals were evaluated for motor activity, fore and hind limb grip strength, audio startle response, thermal response, and hind limb foot splay. Animals were also observed weekly for clinical and behavioural signs, and pharmacotoxic signs.
GLP compliance:
yes
Limit test:
yes
Species:
rat
Strain:
Sprague-Dawley
Sex:
male
Details on test animals or test system and environmental conditions:
TEST ANIMALS
- Source: Charles River Laboratories, Portage, MI
- Age at study initiation:52 days
- Weight at study initiation: 300 g
- Housing: Inidividually housed in wire mesh cages
- Diet (e.g. ad libitum): Purina Certified Rodent Chow No. 5002 ad libitum except during exposure
- Water (e.g. ad libitum): ad libitum
- Acclimation period: 2-3 weeks


ENVIRONMENTAL CONDITIONS
Animal husbandry followed standards by the US Department of Health, Education, and Welfare (1985)
Route of administration:
inhalation: vapour
Details on exposure:
- System of generating particulates/aerosols: Test atmosphere was generated by heating nitrogen to 200°C by passing it through a 1 l stainless steel cylinder with a 1500 W band heater. The nitrogen then passed through a glass column 7.6 cm diameter and 30 cm long packed with glass beads. Test material was delivered by a metering pump into Teflon tubing, to the bottom of the column. The liquid test substance vaporized as it went up the column with the nitrogen. The vapor then went into the test chambers where dilution with the chamber ventilation air produced the desired concentrations.
TEST ATMOSPHERE
- Brief description of analytical method used: Measurements made hourly using gas-phase IR.
- Samples taken from breathing zone: yes
Analytical verification of doses or concentrations:
yes
Details on analytical verification of doses or concentrations:
Concentration of the test material in the test chambers was determined by GC analysis.
Duration of treatment / exposure:
13 weeks
Frequency of treatment:
6 hours/day 5 days/week
Remarks:
Doses / Concentrations:
101 (2.5) ppm
Basis:
analytical conc.
target conc.: 100 ppm
Remarks:
Doses / Concentrations:
432 (2.8) ppm
Basis:
analytical conc.
target conc.: 500 ppm
Remarks:
Doses / Concentrations:
1320 (13) ppm
Basis:
analytical conc.
target conc.: 1500 ppm
No. of animals per sex per dose:
20 male animals per group
Control animals:
yes, concurrent no treatment
Observations and clinical examinations performed and frequency:
CAGE SIDE OBSERVATIONS: Yes
- Time schedule: pretest, monthly intervals
- Cage side observations checked: tremors, convulsions, lacrimation, respiration, urination, defecation, vocalization, piloerection, pupil size

BODY WEIGHT: Yes
- Time schedule for examinations: weekly


Neurobehavioural examinations performed and frequency:
FUNCTIONAL OBSERVATIONAL BATTERY: Yes
- Parameters examined: Fore and hind leg grip strength, thermal response, hind limb foot splay
- Description of procedures: Fore and hind leg grip strength: Force transducers (Grass Instrument Co. Model FT-10) were connected to a chart recorder and mounted on a plexiglass platform. Animals were tested three times and the average used.
Thermal response: An Omnitech, Inc. analgesiometer was used for the hot plate stimulus test. Hind limb foot splay: Diluted tatoo ink was placed on the fifth toe of each hind foot, and the animals dropped from a standard height.
- Time schedule for examinations: 5, 9, 13 weeks after exposure.

LOCOMOTOR ACTIVITY: Yes
- Replicates used: 18-20 animals
- Type of equipment used: Digiscan Activity Meter
- Length of session, number and length of subsessions: Total session time was 0-30 minutes, 0-2 minutes acclimation, then 2-5, and 5-minute intervals
- Total activity: Activity was measured using vertical and horizontal infrared beams. Vertical and horizontal activity was recorded seperately each time an animal broke a beam.

AUDITORY STARTLE REFLEX HABITUATION: Yes
- Number of animals: 20 per dose
- Days of testing: 5, 9 and 13 weeks after exposure initiation
- Type of equipment used: Black acrylic startle box fabricated in the laboratory. The floor of the box was supported by a load cell. Rats were placed in the box, and after five minutes, were stimulated using white noise.
- Number of trials performed: 10
- Length (msec) and intensity (dB) of sound: 110 dBA, 50 msec
- Length of interval between trials: 15 seconds
- Other procedural details: The 2nd through 11th stimuli were measured for amplitude (kg) and latency (ms).
Sacrifice and (histo)pathology:
- Time point of sacrifice: 13 weeks after start of exposure
- Number of animals sacrificed: 10 per group
- Procedures for perfusion: After intraperitoneal anesthesia with barbiturates, heart was perfused with heparizined saline followed by glutaraldehyde, and paraformaldehyde
- Tissues evaluated: proximal sciatic nerves and extensions, tibial, peroneal, and sural nerves, L3 and L4 dorsal root ganglia, dorsal and ventral root fibers, spinal cord, forebrain, cerebrum, midbrain, cerebellum, pons, medulla
- Type of staining: luxol fast blue, hematoxylin, eosin
- Embedding media: osmium tetraoxide
- Number of sections: two
- Number of animals evaluated from each sex and treatment group: 10 animals per group
Statistics:
Kruskal-Wallis one-way analysis of variance, followed if needed by the Mann-Whitney U using a Bonferroni correction were used for analysing body weight, motor activity, and the functional observational battery. Parametric statistics, analysis of variance on the square root activity.
Details on results:
CLINICAL SIGNS AND MORTALITY
No animals died during the study. No clinical signs were considered exposure related.

BODY WEIGHT AND WEIGHT GAIN
The 1500 ppm had depressed weight gain throughout the study. The 500 ppm group also had depressed weight gain, but the difference was only significant at 4 weeks.


NEUROBEHAVIOUR
No signicant differences in motor activity were found in any group. No differences in hind foot splay and grip strength were noted in any group at any time. The only significant difference in auditory response time was in the 500 ppm group at week 9. Since this finding was not dose-related, it was considered to not be biologically important. A significant difference in the thermal response in the mid-dose groups, 100 and 500 ppm, was seen in the pre-test. However, this was most likely due to an unusually low response time for the control group at this time-point.

NEUROPATHOLOGY
No exposure related neuropathological lesions, or degenerative changes were seen.
Key result
Dose descriptor:
NOAEC
Effect level:
1 500 ppm
Sex:
male
Basis for effect level:
other: The test substance had no neurotoxic effects at any of the test concentrations.
Remarks on result:
other:
Key result
Dose descriptor:
LOAEC
Effect level:
1 500 ppm
Sex:
male
Basis for effect level:
other: The test substance was slightly toxic at 1500 ppm as evidenced by depressed weight gain.
Remarks on result:
other:
Conclusions:
The test substance was not neurotoxic at any concentration in the test, therefore the NOAEC = 1500 ppm for neurotoxicity. There was a slight depression in weight gain at 1500 ppm, therefore the LOAEC = 1500 ppm for sub-chronic inhalation toxicity.
Executive summary:

This study was conducted to determine the neurotoxicity of high flash aromatic naphtha. Groups of 20 male rats were exposed via inhalation to 100, 500, or 1500 ppm high flash aromatic naphtha for 6 hrs per day, 5 days per week, for 13 weeks. Neurotoxicity testing was performed at 5, 9, and 13 weeks of exposure. Animals were evaluated for motor activity, fore and hind limb grip strength, audio startle response, thermal response, and hind limb foot splay. Animals were also observed weekly for clinical and behavioural signs, and pharmacotoxic signs. Results of the neurotoxicity testing showed no neurotoxic effects giving an NOAEC of 1500 ppm for neurotoxicity. The 1500 ppm group showed a 12% reduction in body weight gain. Based on the reduction in weight gain, the LOAEC for the test substance is 1500 ppm for sub-chronic inhalation toxicity.

Endpoint:
neurotoxicity: acute inhalation
Type of information:
experimental study
Adequacy of study:
weight of evidence
Study period:
2010
Reliability:
1 (reliable without restriction)
Rationale for reliability incl. deficiencies:
other: Conducted according to sound scientific principles.
Justification for type of information:
A discussion and report on the read across strategy is given as an attachment in IUCLID Section 13.
Reason / purpose for cross-reference:
read-across: supporting information
Qualifier:
no guideline available
Principles of method if other than guideline:
Acute neurobehavior tests.
GLP compliance:
not specified
Species:
rat
Strain:
Crj: CD(SD)
Sex:
male
Details on test animals or test system and environmental conditions:
TEST ANIMALS
- Source:Charles River Wiga, Sulzfeld, Germany.
- Age at study initiation: 14 weeks
- Weight at study initiation: normal
Route of administration:
inhalation: vapour
Vehicle:
unchanged (no vehicle)
Details on exposure:
Rats were exposed by inhalation for periods of up to 8 hours/d for 3 consecutive days in modified H1000 inhalation chambers. The exposures were repeated over several days to assess whether there was accumulation of hydrocarbon solvent constituents in the CNS with an exacerbation of effects over time. Test atmospheres were created by pumping liquid material through heated water baths to create vapors. The vapors were transported with an air stream and added to the main airflow systems for the inhalation chambers. The test atmospheres were continuously monitored as total hydrocarbons using a total carbon analysis method and converted to exposure levels by comparison to concentrations in Tedlar bags of known size and content.
Analytical verification of doses or concentrations:
yes
Duration of treatment / exposure:
8 hours
Frequency of treatment:
3 consecutive days
Remarks:
Doses / Concentrations:
200, 1000, and 5000 mg/m3
Basis:
nominal conc.
Remarks:
Doses / Concentrations:
200, 1010, and 5008 mg/m3
Basis:
analytical conc.
No. of animals per sex per dose:
8 males/ dose
Control animals:
yes, concurrent no treatment
Neurobehavioural examinations performed and frequency:

Evaluation of CNS Effects
The animals were evaluated for viability and other indicators of well-being, functional observations, motor activity, and 2-choice visual discrimination performance.

Functional observations and motor activity. Neurobehavioral functioning was evaluated using a standardized functional observational battery (FOB) that consisted of standardized observations and simple tests designed to evaluate gross changes in neurological and behavioral functioning. Spontaneous motor activity was measured in sessions of 30 minutes using an automated video image analysis system. Rats were tested before exposure and after the first and third 8-hour exposure periods.

Visual discrimination performance. Separate groups of rats were evaluated for 2-choice visual discrimination performance. The apparatus consisted of 16 operant chambers and programming and recording equipment programmed with the MedState notation system. Each operant chamber was equipped with 2 levers, 2 stimulus lights, and a water dipper for delivering water reinforcement. In addition, a photocell assembly was mounted in the water trough to detect the entry of each rat’s head when obtaining reinforcement. Each operant chamber was located in a ventilated, sound-attenuated cubicle. Prior to treatment, water-deprived rats were first trained to obtain water reinforcements and to lever press using auto-shaping techniques. The rats subsequently received 4 weeks of training on a discrete-trial light—dark visual discrimination task to stabilize baseline responding. Animals were trained 5 d/w, from Monday to Friday.
Test sessions consisted of 100 trials or 60 minutes, whichever came first, and were conducted at approximately the same time each day. Dose groups were counterbalanced across time of testing and testing device. Trials were initiated by the illumination of either the left or right stimulus light, and the rat’s task was to depress the lever under the illuminated light to obtain a water reward. Illumination of right and left stimulus lights was counterbalanced and occurred in a predetermined semi-random order. If the rat pressed the correct lever, the stimulus light was extinguished and a water reward was delivered. If the initial response during a trial was on the incorrect lever, the rat was allowed to correct its mistake by pressing the lever under the illuminated stimulus light. A given trial remained in effect until the correct lever had been pressed. Trials were separated by an inter-trial interval (ITT) of 10 seconds. A response during the ITT reset the ITT timer, and the rat was required to wait a further 10 seconds before initiation of the following trial. Rats were tested on the day prior to the first exposure day and on each day of exposure immediately after the exposure period. A post-exposure test was performed the day after the last exposure period to evaluate the persistence of effects.

For each rat, the accuracy of the initial response in each trial was recorded. If the initial trial response was correct, the latency of the lever press was also recorded. If the initial response was incorrect, the number of incorrect lever responses made by the rat before switching to the correct lever was recorded. Following a correct lever response, the water dipper was raised. The system recorded whether the rat positioned itself above the dipper to obtain the water reward, providing a measure of the number of reinforcements obtained. The latency to obtain the reinforcement on each trial was also recorded. During the inter-trial period, lever responses were recorded to determine the number of ITI periods in which 1 or more lever presses occurred and the number of repetitive ITI lever responses.
Statistics:
All data were analyzed using the Statistical Analysis System (SAS) statistical software package (v. 6.12). For each test measure, probability values of P < .05 were considered significant.

Body weights and body temperatures were analyzed using 1-way analysis of variance (ANOVA) conducted at each time point followed by Dunnett multiple comparison tests.

Continuous variables from the FOB were analyzed using ANOVA for pre and post-exposure performance. Treatment effects were analyzed using repeated measures ANOVAEach session consisted of 5 time blocks of 6 minutes each. Rank data were analyzed by Kruskal-Wallis 1-way ANOVA on each test day followed by planned multiple comparisons in case of a significant result.

Baseline visual discrimination performance prior to exposure was examined in 2 ways: (1) by examining the mean performance averaged across the 5 days in the week prior to exposure (preweek responding) and (2) by examining the performance on the day preceding exposure (preday responding). Treatment effects were analyzed using repeated measures ANOVA of the data recorded during the 3-day exposure period. Huynh-Feldt adjustment of P values of the repeated measures factor was applied in case the assumption of sphericity of observations was violated. When a significant treatment effect was demonstrated, pairwise group comparisons were performed to determine which solvent-treated group differed significantly from the control group. When a significant treatment-by-time interaction was demonstrated, 1-way ANOVA was performed at each test time point followed by Dunnett multiple comparison tests. Persistence of effects was evaluated by ANOVA of post- exposure data.
Clinical signs:
effects observed, treatment-related
Description (incidence and severity):
red-colored exudate around the mouth (at 5000 mg/m3)
Mortality:
mortality observed, treatment-related
Description (incidence):
red-colored exudate around the mouth (at 5000 mg/m3)
Body weight and weight changes:
effects observed, treatment-related
Description (incidence and severity):
(approximately 10%; at 5000 mg/m3)
Food consumption and compound intake (if feeding study):
not examined
Food efficiency:
not examined
Water consumption and compound intake (if drinking water study):
not examined
Ophthalmological findings:
not examined
Clinical biochemistry findings:
not examined
Behaviour (functional findings):
effects observed, treatment-related
Description (incidence and severity):
(at 5000 mg/m3)
Gross pathological findings:
no effects observed
Neuropathological findings:
no effects observed
Details on results:
Viability and Other Indicators of Well-Beings
Mixed isomer C9 aromatic solvent After the first 8-hour exposure period, 5 of 8 rats from the high exposure group (5000 mg/rn3) had red-colored exudate around the mouth and nose. Some of these also had wet lower jaws due to salivation; piloerection; and wet anogenital areas due to urination. Similar observations involving 4 of 8 rats from the high exposure group were made after the third 8-hour exposure period. There was a significant difference in body weight (approximately 10%; P < .05) between the high exposure group and the corresponding controls after the third 8-hour exposure, and the body temperatures in the high exposure group were reduced by approximately 1.5°C after the first and third 8-hour exposures (P < .05; data not shown).

Functional Observations
Mixed isomer C9 aromatic so/vent There were effects on gait in the high exposure group—all animals walked on tiptoes, 6 of 8 animals had hunched body positions, and I was slightly ataxic after the first 8-hour exposure. Similar observations were made after the third 8-hour exposure although at this time point, only I animal had a hunched body position. The severity of these gait abnormalities was low to moderate, and a statistically significant (P < .05) difference between the high exposure group and the control group was indicated. There were no significant differences in arousal or approach, click response or tail pinch, foot splay or grip strength. Response to touch was significantly (P < .05) reduced in the high exposure group after the third 8-hour exposure. Motor activity was also affected in the high exposure group with both total distance and number of movements significantly (P < .05) reduced in comparison to controls after the third 8-hour exposure. Although these differences in motor activity were small, they did appear to have been treatment-related.

Visual Discrimination Performance
Mixed isomer C9 aromatic solvent. The results of the visual discrimination tests of the mixed isomer C9 aromatic solvent were very similar to those of TMB, described above. During the 3-day exposure period, there were no differences in number of trials completed, discrimination ratios, or responses during the ITI. There was a significant (P < .05) increase in trial response latency in the mid and high exposure groups, together with increased variability of response latencies in the high exposure group. Significant (P < .05) reductions in the number of short latencies and increases in the number of long latencies were recorded during the 3-day exposure period in the mid and high exposure groups. There was also a significant (P < .05) increase in drink response latency in the high exposure group. No differences were found in the post-exposure assessments.
Conclusions:
Statistically significant effects in these domains were apparent in the high exposure groups (5000 mg/rn3). There were no statistically significant effects on visual discrimination in the intermediate exposure groups (1000 mg/rn3). Post-exposure studies demonstrated the reversibility of all effects. These data, as well as evidence from previous studies, suggested that exposures to levels below approximately 1000 mg/m3 are unlikely to produce profound acute CNS effects or to produce irreversible systemic toxicity.
Executive summary:

Statistically significant effects in these domains were apparent in the high exposure groups (5000 mg/rn3). There were no statistically significant effects on visual discrimination in the intermediate exposure groups (1000 mg/rn3). Post-exposure studies demonstrated the reversibility of all effects. These data, as well as evidence from previous studies, suggested that exposures to levels below approximately 1000 rng/m3 are unlikely to produce profound acute CNS effects or to produce irreversible systemic toxicity.

Endpoint:
neurotoxicity: acute inhalation
Type of information:
experimental study
Adequacy of study:
weight of evidence
Study period:
2010
Reliability:
1 (reliable without restriction)
Rationale for reliability incl. deficiencies:
other: Conducted according to sound scientific principles.
Justification for type of information:
A discussion and report on the read across strategy is given as an attachment in IUCLID Section 13.
Reason / purpose for cross-reference:
read-across: supporting information
Qualifier:
no guideline available
Principles of method if other than guideline:
Acute neurobehavior tests.
GLP compliance:
not specified
Species:
rat
Strain:
Crj: CD(SD)
Sex:
male
Details on test animals or test system and environmental conditions:
TEST ANIMALS
- Source:Charles River Wiga, Sulzfeld, Germany.
- Age at study initiation: 14 weeks
- Weight at study initiation: normal
Route of administration:
inhalation: vapour
Vehicle:
unchanged (no vehicle)
Details on exposure:
Rats were exposed by inhalation for periods of up to 8 hours/d for 3 consecutive days in modified H1000 inhalation chambers. The exposures were repeated over several days to assess whether there was accumulation of hydrocarbon solvent constituents in the CNS with an exacerbation of effects over time. Target exposure concentrations were 125, 1250, or 5000 mg/rn3 for TMB. The highest concentration tested was near a maximally attainable vapor concentration at 20°C. Test atmospheres were created by pumping liquid material through heated water baths to create vapors. The vapors were transported with an air stream and added to the main airflow systems for the inhalation chambers. The test atmospheres were continuously monitored as total hydrocarbons using a total carbon analysis method and converted to exposure levels by comparison to concentrations in Tedlar bags of known size and content.
Analytical verification of doses or concentrations:
yes
Duration of treatment / exposure:
8 hours
Frequency of treatment:
3 consecutive days
Remarks:
Doses / Concentrations:
125, 1250, or 5000 mg/m3
Basis:
nominal conc.
Remarks:
Doses / Concentrations:
128, 1255, and 4980 mg/m3
Basis:
analytical conc.
No. of animals per sex per dose:
8 males/ dose
Control animals:
yes, concurrent no treatment
Neurobehavioural examinations performed and frequency:

Evaluation of CNS Effects
The animals were evaluated for viability and other indicators of well-being, functional observations, motor activity, and 2-choice visual discrimination performance.

Functional observations and motor activity. Neurobehavioral functioning was evaluated using a standardized functional observational battery (FOB) that consisted of standardized observations and simple tests designed to evaluate gross changes in neurological and behavioral functioning. Spontaneous motor activity was measured in sessions of 30 minutes using an automated video image analysis system. Rats were tested before exposure and after the first and third 8-hour exposure periods.

Visual discrimination performance. Separate groups of rats were evaluated for 2-choice visual discrimination performance. The apparatus consisted of 16 operant chambers and programming and recording equipment programmed with the MedState notation system. Each operant chamber was equipped with 2 levers, 2 stimulus lights, and a water dipper for delivering water reinforcement. In addition, a photocell assembly was mounted in the water trough to detect the entry of each rat’s head when obtaining reinforcement. Each operant chamber was located in a ventilated, sound-attenuated cubicle. Prior to treatment, water-deprived rats were first trained to obtain water reinforcements and to lever press using auto-shaping techniques. The rats subsequently received 4 weeks of training on a discrete-trial light—dark visual discrimination task to stabilize baseline responding. Animals were trained 5 d/w, from Monday to Friday.
Test sessions consisted of 100 trials or 60 minutes, whichever came first, and were conducted at approximately the same time each day. Dose groups were counterbalanced across time of testing and testing device. Trials were initiated by the illumination of either the left or right stimulus light, and the rat’s task was to depress the lever under the illuminated light to obtain a water reward. Illumination of right and left stimulus lights was counterbalanced and occurred in a predetermined semi-random order. If the rat pressed the correct lever, the stimulus light was extinguished and a water reward was delivered. If the initial response during a trial was on the incorrect lever, the rat was allowed to correct its mistake by pressing the lever under the illuminated stimulus light. A given trial remained in effect until the correct lever had been pressed. Trials were separated by an inter-trial interval (ITT) of 10 seconds. A response during the ITT reset the ITT timer, and the rat was required to wait a further 10 seconds before initiation of the following trial. Rats were tested on the day prior to the first exposure day and on each day of exposure immediately after the exposure period. A post-exposure test was performed the day after the last exposure period to evaluate the persistence of effects.

For each rat, the accuracy of the initial response in each trial was recorded. If the initial trial response was correct, the latency of the lever press was also recorded. If the initial response was incorrect, the number of incorrect lever responses made by the rat before switching to the correct lever was recorded. Following a correct lever response, the water dipper was raised. The system recorded whether the rat positioned itself above the dipper to obtain the water reward, providing a measure of the number of reinforcements obtained. The latency to obtain the reinforcement on each trial was also recorded. During the inter-trial period, lever responses were recorded to determine the number of ITI periods in which 1 or more lever presses occurred and the number of repetitive ITI lever responses.
Statistics:
All data were analyzed using the Statistical Analysis System (SAS) statistical software package (v. 6.12). For each test measure, probability values of P < .05 were considered significant.

Body weights and body temperatures were analyzed using 1-way analysis of variance (ANOVA) conducted at each time point followed by Dunnett multiple comparison tests.

Continuous variables from the FOB were analyzed using ANOVA for pre and post-exposure performance. Treatment effects were analyzed using repeated measures ANOVAEach session consisted of 5 time blocks of 6 minutes each. Rank data were analyzed by Kruskal-Wallis 1-way ANOVA on each test day followed by planned multiple comparisons in case of a significant result.

Baseline visual discrimination performance prior to exposure was examined in 2 ways: (1) by examining the mean performance averaged across the 5 days in the week prior to exposure (preweek responding) and (2) by examining the performance on the day preceding exposure (preday responding). Treatment effects were analyzed using repeated measures ANOVA of the data recorded during the 3-day exposure period. Huynh-Feldt adjustment of P values of the repeated measures factor was applied in case the assumption of sphericity of observations was violated. When a significant treatment effect was demonstrated, pairwise group comparisons were performed to determine which solvent-treated group differed significantly from the control group. When a significant treatment-by-time interaction was demonstrated, 1-way ANOVA was performed at each test time point followed by Dunnett multiple comparison tests. Persistence of effects was evaluated by ANOVA of post- exposure data.
Clinical signs:
no effects observed
Mortality:
no mortality observed
Body weight and weight changes:
effects observed, treatment-related
Description (incidence and severity):
(5% reduction at 5000 mg/m3)
Food consumption and compound intake (if feeding study):
not examined
Food efficiency:
not examined
Water consumption and compound intake (if drinking water study):
not examined
Ophthalmological findings:
not examined
Clinical biochemistry findings:
not examined
Behaviour (functional findings):
effects observed, treatment-related
Description (incidence and severity):
(at 5000 mg/m3)
Gross pathological findings:
no effects observed
Neuropathological findings:
no effects observed
Details on results:
Viability and Other Indicators of Well-Beings
There were no noteworthy observations. There were small (approximately 5%) but statistically significant (P < .05) body weight differences in the 5000 mg/ m3 groups from both the FOB and visual discrimination studies at the end of the 3-day exposure period.

Functional Observations
There was 1 observation of abnormal gait; 1 rat walked on tiptoes after the first 8-hour exposure period but did not show any evidence of impaired gait after the third 8-hour exposure in the high exposure group. There were also some changes in forelimb grip strength. A significantly (P < .05) reduced mean grip strength was observed in the low exposure group after the first 8-hour exposure, but due to the absence of dose response, this observation is not considered related to exposure. The only effect in the high exposure group was a statistically significant (P < .05) increase in forelimb grip strength after the third exposure when compared to the control group. This is considered a spurious finding as a relationship between TMB exposure and an improvement in grip strength seems implausible. There were no statistically significant differences in any of the other functional observations.

There were some statistically significant findings in the motor activity assessment, specifically significant reductions in total distance traveled and number of movements. These findings may have been related to treatment; however, differences became significant (P < .05) only after the third day of treatment, and not on the first, although brain concentrations in the high exposure group were higher on the first day of treatment than the third; the relationship of this observation to treatment is not straightforward.

Visual Discrimination Performance
Visual discrimination performance testing revealed some treatment-related effects, particularly on response latency in the high exposure group. There were no differences in number of trials completed. The discrimination ratio was significantly (P < .05) better in the high exposure group (5000 mg/m3) than in controls after a single 8-hour exposure, but a relationship to treatment did not seem plausible. There were no differences in frequency of response during the ITIs. The frequency of repetitive ITI responses was significantly (P < .05) reduced in comparison to control after either 2 or 3 exposures, but again, it seems unlikely that treatment would have improved performance. There was a small but statistically significant (P < .05) increase in trial response latency in the high exposure group after a single 8-hour exposure, accompanied by a reduction in short (ie, <2 seconds) latency responses and an increase in long (ie, >6 seconds) latency responses. In addition, a significant (P < .05) increase in drink response latency was observed after a single 8-hour exposure in the 1250 and 5000 mg/rn3 groups. It should be noted, however, that the mean drink response latency in the 1250 mg/rn3 group was increased also on the preexposure day, and, therefore, the significance of the effect of exposure in this group is questionable. The effects on response latency did seem to have been treatment related in that there was a relationship to exposure level. It was apparent that the CNS effects were transient because there were no differences between groups in the post- exposure measurements.
Conclusions:
Statistically significant effects in these domains were apparent in the high exposure groups (5000 mg/rn3). There were no statistically significant effects on visual discrimination in the intermediate exposure groups (1250 mg/rn3). Post-exposure studies demonstrated the reversibility of all effects. These data, as well as evidence from previous studies, suggested that exposures to levels below approximately 1000 rng/m3 are unlikely to produce profound acute CNS effects or to produce irreversible systemic toxicity.
Executive summary:

Statistically significant effects in these domains were apparent in the high exposure groups (5000 mg/rn3). There were no statistically significant effects on visual discrimination in the intermediate exposure groups (1250 mg/rn3). Post-exposure studies demonstrated the reversibility of all effects. These data, as well as evidence from previous studies, suggested that exposures to levels below approximately 1000 rng/m3 are unlikely to produce profound acute CNS effects or to produce irreversible systemic toxicity.

Effect on neurotoxicity: via dermal route

Endpoint conclusion
Endpoint conclusion:
no study available

Additional information

Hydrocarbons, C9 Aromatics:

A study was conducted to determine the neurotoxicity of high flash aromatic naphtha. Groups of 20 male rats were exposed via inhalation to 100, 500, or 1500 ppm high flash aromatic naphtha for 6 hrs per day, 5 days per week, for 13 weeks. Neurotoxicity testing was performed at 5, 9, and 13 weeks of exposure. Animals were evaluated for motor activity, fore and hind limb grip strength, audio startle response, thermal response, and hind limb foot splay. Animals were also observed weekly for clinical and behavioural signs, and pharmacotoxic signs. Results of the neurotoxicity testing showed no neurotoxic effects giving an NOAEC of 1500 ppm for neurotoxicity. The 1500 ppm group showed a 12% reduction in body weight gain. Based on the reduction in weight gain, the LOAEC for the test substance is 1500 ppm for sub-chronic inhalation toxicity.

 

In an acute inhalation neurotoxicity study (McKee et al. 2010) that tested Commercial, C9 hydrocarbon solvents, statistically significant effects were apparent in the high exposure groups (5000 mg/m3). There were no statistically significant effects on visual discrimination in the intermediate exposure groups (1000 mg/m3). Post-exposure studies demonstrated the reversibility of all effects. These data, as well as evidence from previous studies, suggested that exposures to levels below approximately 1000 mg/m3 are unlikely to produce profound acute CNS effects or to produce irreversible systemic toxicity.

 

In an acute inhalation neurotoxicity study (McKee et al. 2010) that tested 1,2,4-Trimethyl benzene, statistically significant effects were apparent in the high exposure groups (5000 mg/m3). There were no statistically significant effects on visual discrimination in the intermediate exposure groups (1250 mg/m3). Post-exposure studies demonstrated the reversibility of all effects.These data, as well as evidence from previous studies, suggested that exposures to levels below approximately 1000 mg/m3 are unlikely to produce profound acute CNS effects or to produce irreversible systemic toxicity.

 

Hydrocarbons, C10 Aromatics:

A study was conducted to evaluate the behavioral effects of rats exposed to hydrocarbons, C10, aromatics. Three test groups (with one control) comprising of 8 rats each were exposed to hydrocarbons, C10, aromatics at different concentrations including: 0 (air), 200 mg/m3 (35 ppm), 600 mg/m3 (110 ppm), 2000 mg/m3 (365 ppm). Animals were exposed to the test atmosphere 8 hours/day for 3 consecutive days. Test methods included selected functional observational measures, automated motor activity assessment and visual discrimination performance.

 

Results of the behavioral tests indicated hydrocarbons, C10, aromatics induced disturbances in measures from different functional domains including gait abnormalities and visual discrimination performance. Some gait abnormalities were observed throughout the 3-day exposure period in rats exposed to the highest concentration of hydrocarbons, C10, aromatics (2000 mg/m3). The severity of these abnormalities was low to moderate. Effects were also observed on measures of learned performance. Exposure to the highest concentration of hydrocarbons, C10, aromatics (2000 mg/m3) induced increased latencies to make a correct choice and latencies to obtain water reinforcement, and also increased the variability in the speed of responding. The effects of exposure to hydrocarbons, C10, aromatics on performance speed were most clearly observed after the first 8-hour exposure period. Also, a small but significant decrease in the number of collected reinforcements was observed in the highest exposure group (2000 mg/m3).

 

Short-term, high-level exposure to hydrocarbons, C10, aromatics induced some mild, reversible neurobehavioral effects on functional observations and measurements of learned performance. Effects were observed during or after 3 consecutive 8 hour exposures at the highest tested concentration of 2000 mg/m3 of hydrocarbons, C10, aromatics. Exposure to 200 mg/m3 or 600 mg/m3 of hydrocarbons, C10, aromatics did not induce exposure-related neurobehavioral effects.

 

No chronic neurotoxicity specific studies for C10 aromatic fluids were located. However, in a 13 week sub-chronic inhalation study, the toxicity of C10 aromatic fluids was examined in both rats and dogs (Carpenter, 1977). There were no neurological effects noted by the researchers in either species. There were no abnormalities noted in the histopathological examination of the brain or in the peripherial nerves for either species. The NOAEC for rats and for dogs was determined to be > 0.38 mg/L, which was the highest concentration tested. Therefore, C10 aromatic hydrocarbon fluids are not likely to cause neurotoxicity. 

Justification for classification or non-classification

Based on the results from available studies, hydrocarbons, C9, aromatics, warrant classification as STOT Single Exp. 3 (H336: May cause drowsiness or dizziness) under Regulation (EC) 1272/2008 on classification, labelling and packaging of substances and mixtures (CLP).