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Neurotoxicity

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Description of key information

Data on main constituents of the Other Petroleum Gases category indicate members of the category show low sub-chronic and chronic toxicity and low potential for neurotoxicity. Inhalation exposure is the most relevant route. No significant exposure-related toxicological or neurotoxicological effects have been observed in inhalation studies up to 90 days duration on the C2-C4 alkanes or a category stream.

Key value for chemical safety assessment

Effect on neurotoxicity: via oral route

Link to relevant study records
Reference
Endpoint:
neurotoxicity: inhalation
Remarks:
other: A 13-week whole-body inhalation toxicity study in rats with neurotoxicity assessments and in vivo genotoxicity assessments
Type of information:
experimental study
Adequacy of study:
key study
Reliability:
1 (reliable without restriction)
Rationale for reliability incl. deficiencies:
other: GLP compliant (except for test substance identity, stability and lot number), guideline study, available as unpublished report, no restrictions, fully adequate for assessment.
Qualifier:
according to guideline
Guideline:
other: OECD Guideline 413 (Subchronic Inhalation Toxicity: 90-Day)
Deviations:
yes
Remarks:
The identity and stability was not in compliance with GLP regulations. The lot number was not available.
Qualifier:
according to guideline
Guideline:
other: EPA OPPTS 870.3465 (90-Day Inhalation Toxicity)
Principles of method if other than guideline:
13-week whole-body inhalation toxicity study in rats with neurotoxicity assessments and in vivo genotoxicity assessments
GLP compliance:
yes
Limit test:
no
Species:
rat
Strain:
other: Sprague-Dawley CD
Sex:
male/female
Details on test animals or test system and environmental conditions:
TEST ANIMALS
- Species: Albino rats (Outbred) VAF/Plus®, Sprague-Dawley derived (CD®), Crl:CD®(SD)IGS BR
- Source: Charles River Laboratories, Kingston, New York 12484, USA
- Age at study initiation: Approximately 8 weeks
- Weight at study initiation: Males mean 280 g (range 243-308 g); females mean 209.1 g (range 187-231 g)
- Fasting period before study: None
- Housing: Individually in stainless steel suspended cages with wire mesh floors and fronts.
- Diet: Certified Rodent diet No 5002 (PMI Nutrition International, St Louis, Missouri, USA) ad libitum
- Water: Municipal water ad libitum
- Acclimation period: Approximately 2 weeks

ENVIRONMENTAL CONDITIONS
- Temperature: 17-25°C
- Humidity: 22-99%
- Air changes (per hr): Not reported
- Photoperiod: 12 hrs dark / 12 hrs light

IN-LIFE DATES: From: 20 April 2005 To: 27 July 2005
Route of administration:
inhalation
Vehicle:
other: air
Details on exposure:
GENERATION OF TEST ATMOSPHERE / CHAMBER DESCRIPTION
- Exposure apparatus: Stainless steel and glass whole body exposure chambers with a volume of approximately 1000 L
- Method of holding animals in test chamber: Individually housed in stainless steel, wire mesh cages within the exposure chamber, with the placement of animals in chambers rotated weekly to ensure uniform exposure.
- Generation: Pre-study trials had evaluated the optimal set of conditions and equipment that generated a stable and uniform atmosphere at the target exposure levels. Test substance flowed from cylinder into a copper tubing coil, maintained in a warm water bath. From the coil the test substance flowed through a metering valve to a mass flowmeter and then via tubing to the turret of the exposure chamber, where it was mixed with room air.
- Temperature, humidity in exposure chamber: 19-28°C, 22-61%
- Air flow rate: Operated at a minimum rate of 203 L/min. Final airflow set to provide at least one air change /5 mins (12/hour) and a T99 equilibrium time of at most 23 mins.
- Oxygen level: at least 19%
- Animal loading factor: below 5%
- Method of particle size determination: yes. Samples drawn for 20 secs at a flow rate of 5 L/min. MMAD, GSM and total mass concentration were calculated.
- Treatment of exhaust air: coarse filter, a HEPA filter and activated charcoal

TEST ATMOSPHERE
- Brief description of analytical method used: Infrared spectrophotometer (IR) 4 times per chamber per day. Gas chromatography (GC) used to characterise at least 5 major constituents (comprising at least 90% by weight of test substance) once/week/chamber to show test substance stability and comparison between neat test substance and test atmospheres.
- Samples taken from breathing zone: yes
Analytical verification of doses or concentrations:
yes
Details on analytical verification of doses or concentrations:
Exposure levels were determined using an infrared spectrophotometer 4 times/chamber/day. Particle size distribution measurements were also made once weekly using a TSI Aerodynamic Particle Sizer.
Duration of treatment / exposure:
13 weeks
Frequency of treatment:
6 hours/day, 5 days/week
Remarks:
Doses / Concentrations:
0, 1000, 5000, 10000 ppm
Basis:
other: target conc.
Remarks:
Doses / Concentrations:
0.0 ± 0.0, 1019 ± 58, 5009 ± 174, 9996 ± 261 ppm
Basis:
analytical conc.
No. of animals per sex per dose:
15
Control animals:
yes, sham-exposed
Details on study design:
- Dose selection rationale: Based on results of a range-finding study (HLS Study number 03-6140) which showed no effects at 100, 1000 or 10000 ppm and no more than 50% of the lower explosion limit (LEL = 21000 ppm)
Observations and clinical examinations performed and frequency:
CAGE SIDE OBSERVATIONS: Yes
- Time schedule: twice daily (mortality and severe toxic effects)

DETAILED CLINICAL OBSERVATIONS: Yes
- Time schedule: During exposure, animals were observed at least once as a group for behavioural effects, signs of toxicity, and mortality. All animals except those selected for neuropathology were examined twice pretesting and weekly during the study period for clinical signs of toxicity.

BODY WEIGHT: Yes
- Time schedule for examinations: On assignment to treatment group, on day of treatment began, and weekly thereafter

FOOD CONSUMPTION: Yes
- Time schedule for examinations: weekly

WATER CONSUMPTION: No

OPHTHALMOSCOPIC EXAMINATION: Yes
- Time schedule for examinations: Pre-test and at termination

HAEMATOLOGY: Yes
- Time schedule for collection of blood: at termination
- Anaesthetic used for blood collection: Yes (60% carbon dioxide/40% oxygen)
- Animals fasted: Yes (overnight)
- How many animals: 10 sex/group
- Parameters examined: haemoglobin concentration, haematocrit, erythrocyte count, platelet count, mean platelet volume, mean corpuscular volume, mean corpuscular haemoglobin, mean corpuscular haemoglobin concentration, red cell distribution width, total leukocyte count, reticulocyte count, differential leukocyte count, erythrocyte and platelet morphology, prothrombin time, activated partial thromboplastin time

CLINICAL CHEMISTRY: Yes
- Time schedule for collection of blood: at termination
- Anaesthetic used for blood collection: Yes (60% carbon dioxide/40% oxygen)
- Animals fasted: Yes (overnight)
- How many animals: 10 sex/group
- Parameters examined: aspartate aminotransferase, alanine aminotransferase, alkaline phosphatase, lactate dehydrogenase, blood urea nitrogen, creatinine, glucose, creatinine kinase, cholesterol, total protein, albumin, total bilirubin, direct bilirubin, sodium, potassium, chloride, calcium, inorganic phosphorus, gamma-glutamyl transferase, triglycerides, globulin, albumin/globulin ratio, indirect bilirubin

URINALYSIS: No
Specific biochemical examinations:
NEUROPATHY TARGET ESTERASE (NTE) ACTIVITY: No

CHOLINESTERASE ACTIVITY: No
Neurobehavioural examinations performed and frequency:
FREQUENCY / NUMBER OF ANIMALS: Neurobehavioral studies were conducted on non-exposure days, at least 16 hours post exposure, on 10 rats/sex/group. Testing was staggered over 5 sessions. All animals selected for neuropathology were tested as well as 5 rats/sex/group from the main study. Neurobehavioral testing consisted of functional observational battery and motor activity assessments

FUNCTIONAL OBSERVATIONAL BATTERY: Yes
- Parameters examined: posture, vocalisation, palpebral closure, motor movements, handling evaluations, open field evaluations, reflex assessments, grip strength, hindlimb extensor strength, landing foot splay, air righting ability, bodyweight, body temperature)
- Minimisation of bias: Yes
- Same technicians used throughout testing: No data
- Technicians were blind to treatment status of animals: Yes
- Site of testing: Home cage (posture, vocalisation, palpebral closure, motor movements), outside of home cage (handling evaluations, open field evaluations, reflex assessments, grip strength, hindlimb extensor strength, landing foot splay, air righting ability, bodyweight, body temperature)
- Time schedule for examinations: pre-test and during 2nd, 4th, 8th and 13th week of exposure.
- Environmental conditions:
- Noise level: 60 decibels by a white noise generator
- Other: Temperature 20-23°C, Relative humidity 36-72%, illumination 65-67 footcandles

LOCOMOTOR ACTIVITY: Yes
- Type of equipment used: Automated Photobeam Activity System
- Length of session, number and length of subsessions: 60 minutes subdivided into 12 sessions of 5 mins. Time of testing balanced across treatment groups.
- Parameters measured: Mean motor activity (number of beam breaks per interval).
- Time schedule for examinations: pre-test and during 2nd, 4th, 8th and 13th week of exposure.
- Environmental conditions:
- Noise level: 60-63 decibels
- Other: Temperature 22-24°C, Relative humidity 36-68%, illumination 49-67 footcandles
Sacrifice and (histo)pathology:
SACRIFICE: At end of treatment period, all animals were sacrificed. they were fasted overnight prior to exsanguination following carbon dioxide inhalation.

GROSS PATHOLOGY: Yes (all animals)- included external examinations as well as examination of the brain, spinal cord, organs and tissues of the cranial, thoracic, abdominal, and pelvic cavities and neck.

ORGAN WEIGHTS: Yes (all animals). Adrenal gland, brain, epididymes, heart, kidneys, liver, lungs (with mainstem bronchi), ovaries, pituitary, prostate, seminal vesicles, spleen, testes, thyroids (with parathyroids), Zymbals gland.

HISTOPATHOLOGY: Yes.
- tissues examined: (control and high dose only) adrenal glands, aorta (thoracic), bone (sternum/femur), brain (medulla/pons, cerebrum and cerebellum), epididymes, oesophagus, eye, heart, kidneys, large intestine (caecum, colon and rectum), lachrymal gland, larynx, liver, lungs (with mainstream bronchi), lymph node (mesenteric), lymph node (mediastinal), mammary glands, muscle (biceps femoris), nasopharngeal tissue, nerve (sciatic), optic nerve, ovaries, pancreas, prostate, salivary gland with submandibular lymph node, seminal vesicles, small intestine (duodenum, ileum and jejunum), spinal cord (cervical, thoracic and lumbar), spleen, stomach, testes, thymic region, thyroids with parathyroids, trachea, urinary bladder, uterus (body/horns with cervix), Zymbals glands, all macroscopic lesions and tissue masses.

NEUROPATHOLOGY: 5/sex/group. Animals were perfused and tissues dissected. Microscopic examination of tissues was as listed above. Measurement of the size (length and width) and weight of whole brain (cerebrum, cerebellum and pons-medulla) were made. In the control and high dose group, the brain (forebrain, central cerebellum, hippocampus, basal ganglia, midbrain, cerebellum, pons and medulla), eye with optic nerve, spinal cord (cervical, thoracic, lumbar cross and longitudinal sections), sciatic nerve (cross and longitudinal sections), tibial nerve (cross and longitudinal sections), sural nerve (cross and longitudinal sections), trigeminal ganglia, dorsal root ganglia (from C3-C6 and L4-L6), dorsal root fibres (from C3-C6 and L4-L6), ventral root fibres (from C3-C6 and L4-L6), lungs and trachea and tissues with macroscopic findings were examined microscopically.
Statistics:
Group mean values of parameters for all the exposure groups were compared to the control group mean values at each time interval, using appropriate statistical methods.
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
Ophthalmological findings:
no effects observed
Behaviour (functional findings):
no effects observed
Gross pathological findings:
no effects observed
Neuropathological findings:
no effects observed
Details on results:
There were no treatment related effects on functional observational battery, motor activity parameters or neuropathology.
Dose descriptor:
NOAEC
Remarks:
neurotoxicity
Effect level:
10 000 ppm
Sex:
male/female
Basis for effect level:
other: highest concentration tested
Remarks on result:
other:
Conclusions:
There were no treatment related effects on functional observational battery, motor activity parameters or neuropathology.

The no observed adverse effect concentration for neurotoxicity for liquified petroleum gas (LPG) was 10000 ppm in this study.
Executive summary:

This study was designed to assess the potential inhalation toxicity of liquefied petroleum gas (LPG) when administered via whole-body exposures to rats for 13 weeks. The assessment included routine toxicology parameters as well as detailed evaluations of neurotoxicity and genotoxicity parameters.

Sprague-Dawley CD® rats (15/sex/group) were exposed for six hours per day to 0 (air control), 1000, 5000 or 10000 ppm of LPG for 5 days per week for 13 consecutive weeks (highest exposure concentration was selected for safety reasons and approximated 50% of the lower explosive limit).

At the end of the treatment period, all animals were euthanized and necropsied. The following parameters were evaluated: viability, clinical observations, body weights, feed consumption, functional observation battery and motor activity, oestrus cycles, sperm assessments, micronucleus assessment, clinical pathology, organ weights, macroscopic and microscopic observations.

All animals (except one female animal exposed at the 10000 ppm level) survived to termination. There were no exposure-related differences in the test substance exposed animals compared to the air control animals. There were no treatment related effects on functional observational battery, motor activity parameters or neuropathology.

In conclusion,13 weeks of exposure of rats to liquified petroleum gas resulted in no adverse effects of exposure. Therefore, the 10000 ppm exposure level was considered a no observed adverse effect level concentration.

Endpoint conclusion
Endpoint conclusion:
no study available

Effect on neurotoxicity: via inhalation route

Endpoint conclusion
Endpoint conclusion:
no adverse effect observed
Dose descriptor:
NOAEC
10 257 mg/m³
Study duration:
subchronic
Species:
rat
Quality of whole database:
Adequate for assessment

Effect on neurotoxicity: via dermal route

Endpoint conclusion
Endpoint conclusion:
no study available

Additional information

Repeat dose toxicity data with neurotoxicity assessments are available for the C2-C4 alkanes and one category stream material. Inhalation exposure is the most relevant route.

 

No systemic toxicity (i.e., no affect on survival, haematological or clinical chemistry parameters, food consumption, body weight, organ weight, and histopathology) or neurological effects (as measured by clinical observations, functional observational battery, and motor activity) were observed in 6-week studies to GLP-compliant guidelines in which ethane, propane, isobutane, butane and were administered to rats by inhalation. The NOAEC in all cases were the maximum dose levels tested (16000, 12000, 9000 and 9000 ppm respectively (equivalent to 19678, 21641, 21394 and 21394 mg/m3 respectively).

 

HLS (2009) report an OECD 414 study designed to assess the potential inhalation toxicity of liquefied petroleum gas (LPG, major constituents propane and propene) when administered via whole-body exposures to rats for 13 weeks. The assessment included routine toxicology parameters as well as detailed evaluations of neurotoxicity parameters. Rats were exposed to 0 (air control), 1000, 5000 or 10000 ppm of LPG, the highest exposure concentration was selected for safety reasons and approximated 50% of the lower explosive limit.

All animals (except one female animal exposed at the 10000 ppm level) survived to termination. There were no exposure-related differences in the test substance exposed animals compared to the air control animals. There were no treatment related effects on functional observational battery, motor activity parameters or neuropathology. The NOAEC was 10000 ppm. 

Justification for classification or non-classification

Members of the Other Petroleum Gases category are flammable gases at room temperature and therefore dermal and oral exposure is unlikely. They have low sub-chronic inhalation toxicity and studies which included evaluation of neurotoxicity parameters indicate low potential for neurotoxicity. Classification under CLP is not warranted.

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