Gases (petroleum), C3-4

Endpoint summary

Description of key information

Only one stream (Liquified Petroleum Gases CAS 68476 -85 -7) of the Other Petroleum Gases category has been tested but this and the main constituents of the category (C1-C4 alkanes and propene) indicate low sub-chronic toxicity by the inhalation route of exposure, the most relevant route. No significant exposure-related toxicological effects or target organ toxicity have been observed in inhalation studies up to 90 days duration on the C2-C4 alkanes, as well as Liquefied Petroleum Gas, the composition of which is mainly propane and propene. Propene has also been thoroughly tested for repeated exposure toxicity up to very high exposure concentrations in sub-chronic and chronic studies. Overall, only minimal local irritation effects to the nasal cavity (mild rhinitis) were observed and only following chronic lifetime exposure to high doses in rats and mice. After repeated dose exposure via oral or inhalation routes, benzene causes adverse effects on the haematopoietic system of animals and humans. The mammalian toxicity effects of this category will be driven by the content of benzene if the latter is present at levels of >10%. Additionally, the category may contain carbon monoxide which could trigger classification.

Key value for chemical safety assessment

Repeated dose toxicity: via oral route - systemic effects

Endpoint conclusion

Endpoint conclusion:

no adverse effect observed

Repeated dose toxicity: inhalation - systemic effects

Link to relevant study records

Reference

Endpoint:

short-term repeated dose toxicity: inhalation

Remarks:

combined repeated dose and reproduction / developmental screening

Type of information:

experimental study

Adequacy of study:

key study

Reliability:

1 (reliable without restriction)

Rationale for reliability incl. deficiencies:

other: GLP compliant (minor exception listed below), guideline study, available as unpublished report, no restrictions, fully adequate for assessment.

Qualifier:

according to guideline

Guideline:

OECD Guideline 422 (Combined Repeated Dose Toxicity Study with the Reproduction / Developmental Toxicity Screening Test)

Deviations:

yes

Remarks:

no lot number available of test material

Qualifier:

according to guideline

Guideline:

other: OPPTS 870.3650 (Combined Repeated Dose Toxicity Study with the Reproduction / Developmental Toxicity Screening Test)

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, Raleigh, North Carolina 27610, USA
- Age at study initiation: Approximately 8 weeks
- Weight at study initiation: Males mean 269 g (range 243-297 g); females mean 200 g (range 180-220 g)
- Fasting period before study: None
- Housing: Individually in stainless steel suspended cages with wire mesh floors and fronts (except for mating period when 1 male and 1 female were housed together)
- Diet: Certified Rodent diet No 5002 (PMI Nutrition International, St Louis, Missouri, USA) ad libitum except during exposure
- Water: Municipal water ad libitum except during exposure
- Acclimation period: Approximately 2 weeks

ENVIRONMENTAL CONDITIONS
- Temperature: 20.1 - 25.7°C
- Humidity: 18.22-92.93%
- Air changes (per hr): Not reported
- Photoperiod: 12 hrs dark / 12 hrs light

IN-LIFE DATES: From: 19 January 2004 To: 18 February 2004

Route of administration:

inhalation

Type of inhalation exposure:

whole body

Vehicle:

other: air

Remarks on MMAD:

MMAD / GSD: MMAD: 2.273, 2.541, 4.061, 6.560 µm; GSD: 1.994, 2.015, 1.981, 2.187; total mass concentration: 3.53 x 10E-3, 4.32 x10E-3, 9.96xE10-3, 1.58x10E-3 mg/m3 for target exposure concentrations of 0, 1200, 4000 and 12000 ppm respectively.

Details on inhalation exposure:

GENERATION OF TEST ATMOSPHERE / CHAMBER DESCRIPTION
- Exposure apparatus: 1000 L glass and stainless steel whole-body exposure chamber
- Method of holding animals in test chamber: housed individually, the placement of animals in the chamber was rotated daily to ensure uniform exposure
- System of generating particulates/aerosols: the test substance was delivered from a single cylinder, through a regulator and two backpressure gauges via a flowmeter into the exposure chambers
- Time to T99: no data
- Airflow rate: no data
- Temperature and humidity in chamber: no data
- Oxygen level: no data
- Air flow rate: no data
- Air change rate: no data
- Method of particle size determination: determined weekly using a TSI Aerodynamic Particle Sizer
- Treatment of exhaust air: filtered through a system which consisted of a coarse filter, a HEPA filter and an activated charcoal bed

TEST ATMOSPHERE
- Brief description of analytical method used: Infrared spectrophotometer (IR) 4 times per chamber per day
- 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. The test substance was evenly distributed within each chamber. The mean (± SD) analaytical concentrations were 0.0 ± 0.0, 1230 ± 34, 3990 ± 156, and 12168 ± 415 ppm.

Duration of treatment / exposure:

Males: 2 weeks prior to mating and after post-mating to give a minimum 28 day exposure period.
Females: 2 weeks prior to mating and post-mating for an additional 4 weeks.

Frequency of treatment:

6 hours/day, 7 days/week

Remarks:

Doses / Concentrations:
0, 1200, 4000 and 12000 ppm
Basis:
other: target concentration

Remarks:

Doses / Concentrations:
0.0 ± 0.0, 1230 ± 34, 3990 ± 156, 12168 ± 415 ppm
Basis:
analytical conc.

No. of animals per sex per dose:

12/sex/group

Control animals:

yes, sham-exposed

Details on study design:

- Dose selection rationale: Based on results of a 2-week range-finding study (HLS Study No. 03-6146) which showed no toxicity at 120, 1200 and 12000 ppm). The high level was established at 12000 ppm since it is 50% of the lower explosion limit for the test substance.

Observations and examinations performed and frequency:

CAGE SIDE OBSERVATIONS: Yes
- Time schedule: twice daily (mortality and clinical condition)

DETAILED CLINICAL OBSERVATIONS: Yes
- Time schedule: Weekly

BODY WEIGHT: Yes
- Time schedule for examinations: At randomisation, first day of exposure and weekly thereafter.

FOOD CONSUMPTION: Yes
- Time schedule for examinations: Pre-test and weekly thereafter.

WATER CONSUMPTION: No

OPHTHALMOSCOPIC EXAMINATION: No

HAEMATOLOGY: Yes
- Time schedule for collection of blood: Study termination
- Anaesthetic used for blood collection: Yes (CO2/O2)
- Animals fasted: Yes (overnight)
- How many animals: 12/sex/group
- Parameters examined: haemoglobin concentration, haematocrit, erythrocyte count, platelet count, mean corpuscular volume, mean corpuscular haemoglobin, mean corpuscular haemoglobin concentration, total leukocyte count, reticulocyte count, differential leukocyte count, erythrocyte and platelet morphology (from peripheral blood smear), prothrombin time, activated partial thromboplastin time..

CLINICAL CHEMISTRY: Yes
- Time schedule for collection of blood: Study termination
- Anaesthetic used for blood collection: Yes (CO2/O2)
- Animals fasted: Yes (overnight)
- How many animals: 12/sex/group
- Parameters examined: aspartate aminotransferase, alanine aminotransferase, alkaline phosphatase, blood urea nitrogen, creatinine, glucose, cholesterol, total protein, triglycerides, albumin, total bilirubin, sodium, potassium, chloride, calcium, inorganic phosphorus, gamma-glutamyl transpeptidase, globulin, albumin/globulin ratio

URINALYSIS: No

NEUROBEHAVIOURAL EXAMINATION: Yes
- Time schedule for examinations: During last week of exposure
- Dose groups that were examined: All
- Battery of functions tested: sensory observations (startle response to auditory stimuli, tail pinch response), neuromuscular observations (grip strength - hindlimb and forelimb), physical observations (rectal temperature) and motor activity assessments.

Sacrifice and pathology:

GROSS PATHOLOGY: Yes (all animals)
- examined: external surfaces, all orifices, cranial cavity, nasal cavity, neck and its associated tissues and organs, thoracic, abdominal and pelvic cavities and their associated tissues and organs and external surfaces of the brain.
ORGAN WEIGHTS: Yes (all animals)
- organs weighed: adrenal glands, brain, epididymes, heart, kidneys, liver, lungs (with mainstem bronchi), ovaries (with oviducts), spleen, testes, thymus, uterus with vagina.
HISTOPATHOLOGY: Yes (control and high dose only).
- tissues examined: adrenal glands, bone (sternum/femur), brain (cerebellum, cerebrum and cerebellum), epididymes, heart, kidneys, large intestine (caecum, colon and rectum), liver, lungs (with mainstream bronchi), lymph node (mesenteric), lymph node (mediastinal), mammary glands (with adjacent skin), ovaries (with oviducts), prostate, seminal vesicles, small intestine (duodenum, ileum and jejunum), spinal cord (cervical, thoracic and lumbar), spleen, stomach, testes, thymus, thyroids with parathyroids, tibial nerve, trachea, urinary bladder, uterus with vagina, all macroscopic lesions and tissue masses.

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:

effects observed, treatment-related

Description (incidence and severity):

Exposure-related 25% decrease in weight gain during the first week of exposures and this difference persisted for the remainder of the 4 weeks of exposure.

Food consumption and compound intake (if feeding study):

no effects observed

Haematological findings:

no effects observed

Clinical biochemistry findings:

no effects observed

Behaviour (functional findings):

no effects observed

Organ weight findings including organ / body weight ratios:

no effects observed

Gross pathological findings:

no effects observed

Histopathological findings: non-neoplastic:

no effects observed

Details on results:

No neurological, haematological, or clinical chemistry effects were observed. There was a 25% decrease in body weight gain in males exposed to 12000 ppm during the first week of exposure which persisted for the remainder of the 4 weeks exposure.

Dose descriptor:

NOAEC

Remarks:

overall systemic toxicity

Effect level:

4 000 ppm

Sex:

male/female

Basis for effect level:

other: based on a reduction of bodyweight gain in males at 12000 ppm.

Dose descriptor:

NOAEC

Remarks:

overall systemic toxicity

Effect level:

7 214 mg/m³ air

Sex:

male/female

Basis for effect level:

other: based on a reduction of bodyweight gain in males at 12000 ppm.

Dose descriptor:

LOAEC

Effect level:

12 000 ppm

Sex:

male/female

Basis for effect level:

other: based on a reduction of bodyweight gain in males at 12000 ppm.

Dose descriptor:

LOAEC

Effect level:

21 641 mg/m³ air

Sex:

male/female

Basis for effect level:

other: based on a reduction of bodyweight gain in males at 12000 ppm.

Critical effects observed:

not specified

Conclusions:

No general systemic or neurological, haematological, or clinical chemistry effects were observed except for a 25% decrease in body weight gain in males exposed to 12000 ppm during the first week of exposure which persisted for the remainder of the 4 weeks exposure.

The overall NOAEC of propane was 4000 ppm and the LOAEC was 12000 ppm in this study.

Executive summary:

The study assessed the potential toxicity, including neurotoxicity and reproductive performance in male and female rats following butane exposure at 1200, 4000 and 12000 ppm (highest exposure level is 50% of the lower explosive limit). It also was designed to investigate effects in both sexes on mating behaviour and on gonadal function, as well as effects on conception, development, parturition and pup survival to lactation day 4.

Male and female rats were exposed for 6 hours/day, 7 days/week for 2 weeks prior to mating initiation. Main study males and females were then evaluated for subchronic effects and were exposed once daily (6 hours/day), seven days/week for 4 weeks (28 days).

There was no effect of treatment on survival. There were no exposure-related systemic effects or effects on body weight, except the 12000 ppm exposed male animals showed an exposure-related 25% decrease in weight gain during the first week of exposures and this difference persisted for the remainder of the 4 weeks of exposure. There were no exposure-related effects on food consumption, functional observation battery (FOB) or motor activity parameters for either sex. Furthermore there were no exposure-related differences in haematology, clinical chemistry and no macroscopic or microscopic changes at post-mortem.

Exposure of male and female rats to target concentrations of 1200, 4000 or 12000 ppm of propane by whole-body inhalation for 4-6 weeks resulted in no general systemic/neurotoxic effects, apart from a reduction of body weight gain in the males resulting from the exposures at 12000 ppm.

The authors report an overall no-observed-adverse effect concentration (NOAEC) of 4000 ppm for general systemic/neurotoxic endpoints in this study, this was based on reduced bodyweight gain in males during the first week. No bodyweight effects were seen in females.

Therefore:

Overall NOAEC both sexes 4000 ppm (equivalent to 7214 mg/m3 (mw 44.094g/mol)) for general systemic/neurotoxic endpoints in this study, based on reduced bodyweight gain in males during the first week.

NOAEC males 4000 ppm (equivalent to 7214 mg/m3) based on reduced bodyweight gain at 12000 ppm during the first week.

NOAEC females 120000 ppm (equivalent to 21641 mg/m3).

Endpoint conclusion

Endpoint conclusion:

no adverse effect observed

Dose descriptor:

NOAEC

7 214 mg/m³

Study duration:

chronic

Species:

rat

Quality of whole database:

Adequate for assessment.

Repeated dose toxicity: inhalation - local effects

Link to relevant study records

Reference

Endpoint:

chronic toxicity: inhalation

Remarks:

combined repeated dose and carcinogenicity

Type of information:

experimental study

Adequacy of study:

key study

Reliability:

2 (reliable with restrictions)

Rationale for reliability incl. deficiencies:

other: Non-GLP; predates implementation of GLP and/or development of study guidelines but otherwise acceptable for assessment

Reason / purpose for cross-reference:

reference to other study

Qualifier:

equivalent or similar to guideline

Guideline:

OECD Guideline 453 (Combined Chronic Toxicity / Carcinogenicity Studies)

Deviations:

yes

Remarks:

only mortality/morbidity, clinical observations, body weight and histopathology were assessed

GLP compliance:

no

Limit test:

no

Species:

rat

Strain:

Fischer 344

Sex:

male/female

Details on test animals or test system and environmental conditions:

TEST ANIMALS
- Source: Charles River Breeding Laboratories (Portage, MI, USA)
- Age at study initiation: 9-10 weeks
- Weight at study initiation: mean bw per group for males 159-169 g; mean bw per group for females 115-121 g
- Fasting period before study: none
- Housing: Individually housed in stainless steel wire cages (Lab Products, Rochelle Pk, NJ, USA)
- Diet: Wayne Lab-Blox® (Allied Mills, Inc., Chicago, IL, USA); freely available except during inhalation exposure
- Water: tap water available ad libitum
- Acclimation period: 5 weeks

ENVIRONMENTAL CONDITIONS
- Temperature: Average of 70°F (equivalent to 21.1 C) (during exposure 75 ± 2°F) (equivalent to 23.9 C)
- Humidity: During non-exposure 54-57 % (during exposure 57 ± 7 %)
- Air changes: 20/hour
- Photoperiod: 12 hrs dark /12 hrs light

IN-LIFE DATES: From: 29 October 1979 To: 28 October 1981

Route of administration:

inhalation: gas

Type of inhalation exposure:

whole body

Vehicle:

other: air

Details on inhalation exposure:

GENERATION OF TEST ATMOSPHERE / CHAMBER DESCRIPTION
Propene gas, at an operating pressure of 54 psi, was metered to the exposure chambers and diluted in the chamber fresh-air inlets. The animals were individually housed in mesh cages (6 cages/exposure chamber). Since the exposure chambers were being operated with concentrations of propene close to the lower explosive limit (LEL) of the gas (25% and 50% of the LEL), safety devices were incorporated in the polyethylene vapour hood (vented to the room exhaust) to minimize the hazard to animals and personnel in the event of a leak. The gas was then piped to a second hood containing four double-pattern metering valves. Since the upstream pressure to these valves was well regulated, these valves provided stable control of the gas flow rate and ultimately of the concentration in the chambers. To provide the proper chamber concentration, the valves were set and periodically checked, by matching the calculated with the actual flow measured by a bubble meter. From the double-pattern metering valves, the gas was piped to each exposure chamber. A shut-off valve at the entrance to the chamber permitted easy, rapid termination of gas flow. All materials in the gas distribution system were stainless steel, Teflon®, viton, or brass.

TEST ATMOSPHERE
The vapour concentration uniformity in the chamber was measured with a portable photoionization detector at 12 positions (2 positions, one at the front and one at the back, for each of the six animal cage units per chamber). The sample point was just above and about 10 cm in from the front or back centre of each cage unit. Propene concentrations in the exposure chambers, control chambers, and exposure room were automatically monitored approximately 10 times during each exposure day by gas chromatography.

Analytical verification of doses or concentrations:

yes

Details on analytical verification of doses or concentrations:

Throughout the studies, samples taken from the chambers several times each day indicated that average daily chamber concentrations were usually within 5 %-6 % of the target concentrations. However, wider variations in exposures were observed during the first 40 weeks of the studies as compared with the remainder of the studies.
Atmospheric samples were obtained from the control and 10000 ppm chambers during an exposure period during week 30 and were analyzed by gas chromatography. No peaks were observed in the air from the control chamber. Only those impurities present in the bulk propene at the pretest analysis were observed in the air from the 10000 ppm chamber

Duration of treatment / exposure:

103 weeks

Frequency of treatment:

6 hours per day, 5 days per week

Remarks:

Doses / Concentrations:
0, 5000, 10000 ppm
Basis:
nominal conc.

Remarks:

Doses / Concentrations:
0, 4985±274, 9891±515 ppm
Basis:
analytical conc.

No. of animals per sex per dose:

50

Control animals:

yes, sham-exposed

Details on study design:

- Dose selection rationale: No compound-related effects were seen in a 14 week inhalation study following exposure at 0, 625, 1250, 2500, 5000, or 10000 ppm. Based on these results even though no propene-related toxicity was observed, concentrations of 5000 and 10000 ppm propene were selected for rats in the 2-year studies. Concentrations higher than 10000 ppm propene could not be selected because of the risk of explosion

Observations and examinations performed and frequency:

CAGE SIDE OBSERVATIONS: Yes
- Time schedule: Twice per day for signs of moribundity and mortality

DETAILED CLINICAL OBSERVATIONS: Yes
- Time schedule: once per month

BODY WEIGHT: Yes
- Time schedule for examinations: Once per week for 14 weeks, once per month for 76 weeks and then biweekly thereafter.

FOOD CONSUMPTION: No

WATER CONSUMPTION: No

OPHTHALMOSCOPIC EXAMINATION: No

HAEMATOLOGY: No

CLINICAL CHEMISTRY: No

URINALYSIS: No

NEUROBEHAVIOURAL EXAMINATION: No

Sacrifice and pathology:

GROSS PATHOLOGY: Yes. Complete necropsy exam performed on all animals.

HISTOPATHOLOGY: Yes. Complete histopathological examination performed on all animals. The following tissues were examined: gross lesions, skin, mandibular lymph node, mammary gland, sternebrae, vertebrae or femur including marrow, thymus, trachea (2 sections), lungs and bronchi, heart, thyroid gland, parathyroids, oesophagus, stomach, colon, small intestine, liver (2 sections), pancreas, spleen, kidneys and adrenal glands (2 sections), urinary bladder, prostate/testes (2 sections) or ovaries/uterus (2 sections), nasal cavity and nasal turbinates (3 sections), brain (3 sections), pituitary gland, and (if abnormal) spinal cord, eyes, and pharynx.

Statistics:

The probability of survival was estimated by the product-limit procedure of Kaplan and Meier (1958). Statistical analyses for a possible dose-related effect on survival used the method of Cox (1972) for testing two groups for equality and Tarone's (1975) life table test for a dose-related trend. All reported P values for the survival analysis are two-sided.
The incidence of neoplastic or non-neoplastic lesions is given as the ratio of the number of animals bearing such lesions at a specific anatomic site to the number of animals in which that site was examined.
Three statistical methods are used to analyze tumour incidence data (Life table analysis, incidental tumour analysis and unadjusted analyses). The two that adjust for intercurrent mortality employ the classical method for combining contingency tables developed by Mantel and Haenszel (1959). Tests of significance included pairwise comparisons of high dose and low dose groups with chamber controls and tests for overall dose-response trends.

Clinical signs:

effects observed, treatment-related

Mortality:

mortality observed, treatment-related

Body weight and weight changes:

effects observed, treatment-related

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

Haematological findings:

not examined

Clinical biochemistry findings:

not examined

Urinalysis findings:

not examined

Behaviour (functional findings):

not examined

Organ weight findings including organ / body weight ratios:

not examined

Gross pathological findings:

not specified

Histopathological findings: non-neoplastic:

effects observed, treatment-related

Histopathological findings: neoplastic:

effects observed, treatment-related

Details on results:

CLINICAL SIGNS AND MORTALITY; No clinical signs were recorded. No significant differences in survival were observed between any groups of either sex.

BODY WEIGHT AND WEIGHT GAIN: The mean bodyweights of exposed male and female rats were comparable to those of the controls throughout the study. The fluctuations in weight gain were not dose-related.

HISTOPATHOLOGY: NON-NEOPLASTIC: Several non-neoplastic effects were observed. Squamous metaplasia of the respiratory epithelium of the nasal cavity occurred in females exposed to propene at both concentrations and in males exposed at the high concentration. Inflammatory changes of the nasal cavity occurred in both males. The changes were characterized as unspecified inflammation and suppurative inflammation. The former lesion, consisting of a mild submucosal influx of lymphocytes, macrophages, and a few granulocytes, occurred in males exposed at the low concentration. The latter lesion was more severe and contained macrophages that migrated through the epithelium and accumulated in the lumen, occurring in male and females exposed at the high concentration. The combined incidence of these nasal cavity lesions was higher in male rats exposed at both
concentrations and in female rats exposed at the high concentration than in controls. These changes may reflect local tissue responses to long-term inhalation exposure to propene.

HISTOPATHOLOGY: NEOPLASTIC: No evidence was found for a carcinogenic effect of propene in rats. C-cell adenomas and C-cell adenomas or carcinomas (combined) of the thyroid gland occurred in female rats with a negative trend, and the incidence of C-cell adenomas in the high concentration group was significantly lower than that in the controls. The incidences in the controls (13%-15%) were higher than those observed in unexposed chamber control F344/N female rats in a propene oxide inhalation study (NTP, 1985) and in untreated control F344/N female rats in other studies. The incidences of C-cell hyperplasia occurred with a positive trend. In rats, C-cell hyperplasia, C-cell adenoma, and C-cell carcinoma appear to represent a continuous spectrum of progressive lesions. When hyperplasia, adenoma, and carcinoma are combined, the negative trend disappears. These comparisons suggest that the lower incidence of thyroid gland neoplasms is not related to administration of propene.

Dose descriptor:

LOAEC

Effect level:

5 000 ppm (nominal)

Sex:

male/female

Basis for effect level:

other: (8,600 mg/m3). No NOAEC identified for local effects. Mild rhinitis (nasal inflammation) and associated epithelial alterations without obvious dose response relationship - refer to expert report of Harkema (2002)

Critical effects observed:

not specified

Inflammation of the nasal cavity, characterized by an influx of lymphocytes, macrophages, and granulocytes into the submucosa and by granulocytes into the lumen, occurred at increased incidences in low concentration and high concentration male rats and in high concentration female rats.

Incidences of nasal inflammatory changes in rats in the two-year inhalation studies of propene

	Control	5000 ppm	10000 ppm
Males
Inflammation, unspecified	4/60 (8%)	14/50 (28%)	5/60 (10%)
Inflammation, suppurative	7/60 (14%)	7/50 (14%)	14/60 (28%)
Inflammation, unspecified or suppurative	11/60 (22%)	21/50 (42%)	19/50 (38%)
Females
Inflammation, unspecified	0/49 (0%)	2/60 (4%)	2/60 (4%)
Inflammation, suppurative	8/49 (16%)	7/60 (14%)	11/60 (22%)
Inflammation, unspecified or suppurative	8/49 (16%)	10/60 (20%)	13/60 (26%)

Conclusions:

A NOAEC was not achieved. The LOAEC was 5000 ppm (lowest dose tested) for males and females based on mild rhinitis (nasal inflammation) and associated epithelial alteration suggesting chronic, low-grade irritation in these rodents. There was no obvious dose response relationship for this effect.

Executive summary:

Toxicology and carcinogenesis studies of propene (greater than 99% pure) were conducted by exposing groups of 50 F344/N rats of each sex to propene in air by inhalation at concentrations of 5000 or 10000 ppm, 6 hours per day, 5 days per week, for 103 weeks. Other groups of 50 rats of each sex received air only on the same schedule and served as chamber controls. The highest concentration of propene that was considered safe for these studies was 10000 ppm because of the risk of explosion that can occur at higher concentrations.

A NOAEC was not reported. The LOAEC reported was 5000 ppm (lowest dose tested) for males and females based on mild rhinitis.

The survival of exposed and control rats was comparable. Throughout most of the studies, mean body weights of exposed male and female rats were slightly lower (0%-5%) than those of the controls, but the decrements were not concentration-related. No compound-related adverse clinical signs were observed. An increased incidence of squamous metaplasia of the nasal cavity was observed in female rats exposed at 5000 and 10000 ppm (control, 0/49; low, 15/50; high, 6/50) and in male rats exposed at 5000 ppm (2/50; 19/50; 7/50). Epithelial hyperplasia of the nasal cavity was increased in female rats exposed at the 10000 ppm concentration (0/49; 4/50; 9/50); the incidences in male rats were 2/50, 2/50, and 5/50. Inflammation of the nasal cavity, characterized by an influx of lymphocytes, macrophages, and granulocytes into the submucosa and by granulocytes into the lumen, occurred at increased incidences in low concentration and high concentration male rats and in high concentration female rats. In the nasal cavity, propene induced squamous metaplasia of the respiratory epithelium in male and female rats and epithelial hyperplasia in female rats.

The nasal findings reported in this study were further investigated by re-evaluation of the archived tissue specimens (Harkema, 2002).

Endpoint conclusion

Endpoint conclusion:

adverse effect observed

Dose descriptor:

LOAEC

8 600 mg/m³

Study duration:

chronic

Species:

rat

Quality of whole database:

Adequate for assessment

Repeated dose toxicity: dermal - systemic effects

Link to relevant study records

Reference

Endpoint:

short-term repeated dose toxicity: dermal

Data waiving:

study technically not feasible

Justification for data waiving:

other:

Critical effects observed:

not specified

Endpoint conclusion

Endpoint conclusion:

no study available

Repeated dose toxicity: dermal - local effects

Link to relevant study records

Reference

Endpoint:

short-term repeated dose toxicity: dermal

Data waiving:

study technically not feasible

Justification for data waiving:

other:

Critical effects observed:

not specified

Endpoint conclusion

Endpoint conclusion:

no study available

Additional information

Repeat dose toxicity data are available for the main constituents of this category and Liquefied Petroleum Gas; also data are available on C1-C4 alkane mixtures.

 

Other Petroleum Gases are flammable at room temperature and therefore exposure via the dermal or oral routes is unlikely and the requirement to test is waived in accordance with REACH Annex XI.

 

Non-human studies

Methane CAS Number 74-82-8

No quantitative repeat dose toxicity data are available specifically for methane.

 

Ethane CAS Number 74-84-0

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 a 6-week study to modern guidelines and GLP in which ethane was administered to rats by inhalation. The experimentally defined NOAEC is 16,000 ppm (19678 mg/m3), the highest exposure level tested and 50% of the lower explosive limit (HLS 2010).

 

Propane CAS Number 74-98-6

No neurological, haematological, or clinical chemistry effects were observed in a 6-week study to modern guidelines and GLP in which propane was administered to male and female rats by inhalation. There was no effect of treatment on survival and there were no exposure-related systemic effects or effects on body weight, except the 12000 ppm exposed male animals showed an exposure-related 25% decrease in weight gain during the first week of exposures and this difference persisted for the remainder of the 4 weeks of exposure. The lowest observed adverse effect concentration (LOAEC) in this study is 12,000 ppm (equivalent to 21641 mg/m³), the highest exposure level tested and 50% of the lower explosive limit, based on the reduced bodyweight gain in males. The NOAEC is 4,000 ppm or 7214 mg/m³(HLS 2009).

 

 

Isobutane CAS Number 75-28-5

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 a 6-week study to modern guidelines and GLP in which isobutane was administered to rats by inhalation. The experimentally defined NOAEC is 9,000 ppm (21394 mg/m3), the highest exposure level tested and 50% of the lower explosive limit (HLS 2010).

A 90 day inhalation study on a 50:50 wt% mixture of isobutane:isopentane exposed male and female rats to nominal 1000 and 4500 ppm daily for 13 weeks, with an interim kill after 28 days. There were no deaths and transient clinical signs were considered treatment but were not dose related. There were no treatment related gross lesions or kidney/liver weight changes. The rats were not significantly affected by the exposures and there was no evidence of hydrocarbon-induced nephropathy in either sex at study termination. At the 28 -day interim kill mild, transient treatment-related kidney effects were observed in the male rats, statistically significant at 1000 ppm only, however there was no evidence of a dose response and the effect disappeared by 90 days. The NOAEC for this study was 4458 ppm, the highest dose tested (Aranyi 1986).

 

Butane CAS Number 106-97-8

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 a 6-week study to modern guidelines and GLP in which butane was administered to rats by inhalation. The experimentally defined NOAEC is 9,000 ppm (21394 mg/m3), the highest exposure level tested and 50% of the lower explosive limit (HLS 2008).

A 90 day inhalation study on a 50:50 wt% mixture of n-butane:n-pentane exposed male and female rats to nominal 1000 and 4500 ppm daily for 13 weeks, with an interim kill after 28 days. There were no deaths and transient clinical signs were considered treatment but was not dose related. Statistically significant decreases in body weights of both sexes were observed by test weeks 3 and 4, with the males, but not the females, recovering towards the end of the exposure period. There were no treatment related gross lesions or kidney/liver weight changes. Rats were not significantly affected by the exposures and there was no evidence of hydrocarbon-induced nephropathy in either sex at study termination. At the 28-day interim kill mild, transient treatment-related but not exposure-related kidney effects were observed in the male rats, this effect disappeared by 90 days. The NOAEC for this study is 4489 ppm, the highest dose tested (Aranyi 1986).

 

In a mixture study, male rats were exposed by inhalation to concentrations up to 11.8 mg/L (11800 mg/m3 or 4437 ppm) of a mixture containing 25% (by weight) each of isobutane, n-butane, n-pentane, and isopentane for 6 hours per day, 5 days per week, for 3 weeks. There were no signs of systemic toxicity, no effects on bodyweight, organ weights, haematology or serum chemistry and no treatment-related gross or microscopic lesions. The NOAEC for systemic effects and kidney effects is 11.8 mg/L (11800 mg/m3 or 4437 ppm), the highest dose tested (Halder, 1986).

 

Butene isomers (butenes):

Although oral administration is an unusual and non-relevant route for a gaseous substance, no toxicologically significant effects occurred when 2-methylpropene was administered orally to rats at 148.6 mg/kg for 28 days (Hazleton 1986b). No significant exposure-related toxicological effects or target organ toxicity have been observed in 6 week inhalation studies on 1-butene and 2-butene in rats at concentrations up to 8000 ppm (18,359 mg/m3) (TNO 1992, Huntingdon 2003).Carcinogenicity studies on 2-methylpropene were conducted by the NTP. Rats and mice were exposed to 2-methylpropene at concentrations of up to 8,000 ppm, (18,359 mg/m3) for 105 weeks (NTP, 1998). The non-neoplastic findings from these studies were confined to effects on nasal tissues. In mice, hyaline degeneration of the olfactory and respiratory epithelium was increased in both sexes. The severities of hyaline degeneration increased with increasing exposure concentration. However, this was considered by the NTP to be a nonspecific adaptive response that had no adverse effect on affected animals. The NOAEC for toxicity in mice was therefore 8000ppm (18,359 mg/m3). Similar findings were observed in rats although the lesions were more severe. An additional finding in rats was that hypertrophy of goblet cells lining the nasopharyngeal duct was marginally increased with 100% incidence in males at 8000 ppm. The NOAEC for toxicity in the rat study was therefore 2000 ppm (4589 mg/m3), lower than that in mice (OECD SIDS Report for Isobutylene, 2003).

 

Propene CAS Number 115-07-1

Key information for the assessment of the repeat-dose toxicity of propene has been reported in several near-guideline sub-chronic and chronic studies via the inhalation route (NTP, 1985). Propene has been thoroughly tested for repeated exposure toxicity up to very high exposure concentrations (10,000 ppm (17,200mg/m3), half of its lower explosive limit value) in sub-chronic and chronic studies. Overall, only minimal local irritation effects to the nasal cavity were observed and only following chronic lifetime exposure to high doses in rats and mice. The mild classification of the lesion and lack of dose-response relationship is consistent with results of shorter term studies, where nasal effects have not been reported in any studies, including 28 day and 90 day NTP studies on propene.The LOAEC for mild rhinitis, only reported following lifetime exposure, is 5000 ppm (8600 mg/m³) in rats and mice. Although there was no clear dose-response relationship or NOAEL for this effect, the weight of evidence clearly indicates that any irritant properties of propene must be extremely weak.

 

Ethylene

Non-human data

The key study is considered to be a GLP-conducted 13-week inhalation toxicity study in rats at ethylene concentrationsof 300, 1000, 3000 and 10,000 ppm (344.2, 1147, 3442 and 11,473 mg/m3) (Dow, 2010). There was no evidence of systemic toxicity based on the lack of toxicologically significant differences in clinical signs (including functional tests, grip performance, and motor activity), body weight, food consumption, body temperature, clinical pathology or organ weights. Evidence of minor local irritation was indicated by histopathological changes in the upper respiratory tract (bilateral, eosinophilic rhinitis with mucous cell hyperplasia/hypertrophy (MCH) and occasional epithelial hyalinosis). The changes, seen at all dose concentrations, were described as very slight or slight, with only minor increase in severity at the highest exposure of 10,000 ppm. Quantitative morphometric analyses substantiated the qualitative histopathologic findings by demonstrating quantifiable increases of both intraepithelial mucosubstances (IM; a quantitative estimate of epithelial remodelling) and eosinophil numeric density (quantitative estimate of nasal inflammation – eosinophilic rhinitis) in the nasal mucosal tissue of ethylene-exposed rats. The NOAEC for systemic toxicity was 10,000ppm (11,473 mg/m3). A NOAEC for local effects was not established, with 300 ppm (344.2 mg/m3) a LOEC for nasal pathology.

 

The local effects described above were investigated further by The Dow Chemical Company (2012). Time- and concentration-dependent changes in histopathology, cellular inflammation, and mucosal gene expression were assessed in nasal tissue from female F344 rats exposed to 0, 10, 50, 300 or 10,000 ppm ethylene vapour for up to 4 weeks. Additional recovery groups (exposed to 300 or 10,000 ppm ethylene for 4 weeks) were maintained for a further 13 weeks to establish the reversibility of any effects seen. Repeated exposure to ethylene was associated with a generally concentration and time dependent increase in minimal to mild nasal inflammation and increased storage of mucosubstances in the rat nasal respiratory mucosa, but was without effect on cell death (apoptosis or necrosis) or epithelial cell regeneration (BrdU incorporation). Systemic IgE or IgG isotypes were unaffected by exposure. Where present, the nasal changes were site specific, generally co-located and limited to the upper respiratory tract; i.e the eosinophilic inflammatory response was greater in nasal respiratory mucosa of the proximal nasal airways rather than in the distal airways, and increases in intraepithelial mucosusbstances were more prominent in the respiratory mucosa lining the distal nasal airways than in the proximal airways.Expression of Th2 cytokines (e.g., IL-5, IL-13) and YM1/2 gene expression in the nasal mucosa was much greater than that of Th1 cytokines after exposure to ethylene. Lesions observed after 4 weeks exposure to 300 or 10,000 ppm ethylene were not present following a 13 week recovery period. The results indicate that sub-acute exposure to ethylene vapour at concentrations up to 10,000 ppm is associated with reversible inflammatory changes in nasal epithelium from F344 rats. An overall LOEC of 10 ppm (11.47 mg/m3) is obtained from this investigation.

A further study assessed whether comparable nasal changes were present in Fischer 344 and Wistar Han rats exposed to target ethylene concentrations of either 0 or 10,000 ppm for 4 or 13 weeks (Hamner Institute 2009, Harkema 2010). There was no mortality, no systemic toxicity and no statistically significant differences in body weights or food consumption during the study. Histopathological findings were described as similar to those seen previously (in Dow 2010). F344 rats had nasal lesions consisting of multifocal subacute rhinitis with accompanying airway epithelial remodelling. The exposure-related inflammatory cell infiltrate consisted mainly of eosinophils and mononuclear leukocytes with lesser numbers of neutrophils. The principal nasal epithelial changes included MCH and eosinophilic hyalinosis. This study included a 4-week exposure period at 10,000 ppm; ethylene-induced inflammatory and epithelial lesions in F344 rats were similar in character, but less severe, to those in F344 rats exposed for 13 weeks. This study also demonstrated a strain-dependent nasal response to ethylene after 13 weeks of exposure, the response being weaker in the Wistar rats.

In two previous studies (Rhudy 1978 and Hamm 1984) groups of rats (strain not specified and Fischer 344) were exposed to ethylene at concentrations of 0, 300, 1000, 3000 and 10,000 ppm for 13 weeks or 0, 300, 1000, or 3000 ppm for 24 months (with groups of animals necropsied and examined after 6, 12 or 18 months). There was no exposure-related mortality and no effects on body weight, weight gain or food consumption. Haematology, clinical chemistry, and urinalyses results also failed to reveal any differences between control and test animals. There were no gross or histopathological alterations attributed to ethylene exposure.

 

Liquefied Petroleum Gases

The major constituents are identified as propane and propene (93.5%).

The repeated-dose inhalation toxicity of petroleum gas products in laboratory animals was investigated in a 90 day study to modern guidelines and GLP. Groups of rats were exposed to target concentrations of 0; 1,000; 5,000; or 10,000 ppm liquefied petroleum gas (LPG) for 13 weeks (HLS, 2008). The highest exposure concentration was approximated 50% of the lower explosive limit. There was no treatment-related effect on survival, terminal body weight, food consumption, functional observational battery, motor activity parameters, haematological parameters, clinical chemistry values, macroscopic or microscopic evaluations, or on organ weights at any exposure concentration. A no observed adverse effect concentration (NOAEC) of 10,000 ppm is reported for the repeated-dose toxicity of the LPG tested.

 

Human studies

Little quantitative data on Other Petroleum Gases were identified.

In a controlled exposure study, Stewart et al (1977, 1978) exposed adult volunteers to isobutane at 500 ppm (1189 mg/m3) 1, 2 or 8 hours/day, five days/week for 2 weeks. During the investigation, all volunteers were kept under comprehensive medical surveillance which included cardiac and pulmonary responses. Repetitive exposures to isobutane were without any measurable untoward physiological effect.

Streams containing >1% benzene:

Oral

Benzene toxicity following sub-chronic and chronic oral (gavage) exposure was investigated in studies using F344/N rats and B6C3F1 mice conducted as part of the National Toxicology Program (NTP, 1986). Animals were dosed 5 days per week for 17 or up to 103 weeks. The most significant findings in rats were a dose-related leukopenia and lymphocytopenia observed in males at ≥ 200 mg/kg and in females at ≥ 25 mg/kg in the 17 week study and in all groups dosed for one and two years. No NOAEL couldbe determined in the 2-year study. The LOAEL was 50 mg/kg/day for male rats and 25 mg/kg/day for female rats, i.e. the lowest doses administered.

In mice, tremors were observed intermittently at 400 and 600 mg/kg throughout the 17 week study. White blood cell and lymphocyte counts were decreased in males at 50 mg/kg bw or more and in females white blood cells were decreased at 600 mg/kg and lymphocytes were decreased at 400 mg/kg or more. In the chronic toxicity study weight gain reductions occurred in male and female mice at 100 mg/kg. Haematotoxic effects were limited to lymphocytopenia and associated leukocytopenia in all dose groups (males from 3 to 18 months, female mice from 12 to 18 months). Benzene increased the frequency of micronucleated normochromatic peripheral erythrocytes in male and female mice of all dose groups, males were more sensitive than females. Haematopoietic hyperplasia in the bone marrow and splenic haematopoiesis was observed in all dosed mice groups. The LOAEL was 25 mg/kg bw/day for male and female mice. A NOAEL was, therefore, not achieved.

Inhalation

In the rat, the key study is considered to be that of Ward et al, 1985. Animals were exposed to concentrations of 0, 1, 10, 30 or 300 ppm (0, 3.2, 9.6, 960 mg/m³) benzene vapour, 6 h/day, 5 days/week, for 13 weeks. Decreased blood lymphocyte counts, relative increase in neutrophil percentages and slightly decreased femoral marrow cellularity were the only significant treatment-related parameters noted in animals exposed to 300 ppm. The NOAEC for toxicity at 28 and 90 days was 30 ppm (96 mg/m³) for both male and female rats.

In mice haematotoxic effects following repeated inhalation exposure to benzene include: decreases in haematocrit, total haemoglobin, erythrocyte count, leukocyte count, platelet count, myeloid/erythroid ratios, and percentage of lymphocytes at 300 ppm (960 mg/m³) (Ward et al, 1985); depressed bone marrow and splenic Multipotential Haematopoietic Stem cells (CFU-S) and Granulocyte/Macrophage Progenitor cells (GM-CFU-C) at benzene concentrations ≥103 ppm (Green et al, 1981a, b); bone marrow erythroid progenitor cell numbers were depressed 1 day after exposure to concentrations ≥ 10 ppm (32 mg/m³) (Dempster and Snyder, 1990); significant depression in femoral lipopolysaccharide (LPS) -induced B-colony-forming ability and splenic phytohaemagglutinin (PHA) -induced blastogenesis at 31 ppm (Rozen et al, 1984); a reduction in bone marrow cellularity and the number of pluripotent stem cells in the bone marrow at ≥ 100 ppm for 10 exposures (Cronkite et al, 1985).

On the basis of these studies the LOAEC for haematotoxicity in mice is 10 ppm (32 mg/m³). A NOAEC could not be defined.

Dermal

No published data are available.

Human data

In a review bySchnatter et al2020, repeat dose studies in workers (seventy-seven genotoxicity and thirty-six haematotoxicity) were scored for study quality with an adapted tool based on that of Vlaanderen et al., 2008 (Environ Health. Perspect. 116 1700−5). These endpoints were selected as they are the most sensitive and relevant to the proposed mode of action (MOA) and protecting against these will protect against benzene carcinogenicity. Lowest and No- Adverse Effect Concentrations (LOAECs and NOAECs) were derived from the highest quality studies (i.e. those ranked in the top tertile or top half) and further assessed as being “more certain” or “less certain”. Several sensitivity analyses were conducted to assess whether alternative “high quality” constructs affected conclusions. The lowest haematotoxicity LOAECs showed effects near 2 ppm (8 h TWA), and no effects at 0.59 ppm. For genotoxicity, studies also showed effects near 2 ppm and showed no effects at about 0.69 ppm. Several sensitivity analyses supported these observations. These data define a benzene LOAEC of 2 ppm (8 h TWA) and a NOAEC of 0.5 ppm (8 h TWA). Allowing for possible subclinical effects in bone marrow not apparent in studies of peripheral blood endpoints, an OEL of 0.25 ppm (8 h TWA) is proposed.

Streams contining >1% 1,3 -Butadiene:

Human information

No chronic non-neoplastic effects have been seen in humans, although data is limited (EU RAR 2002). Long-term exposure of humans to 1,3-butadiene may result in an increased risk of lymphohaematopoietic cancer (see Section on Carcinogenicity). Studies of the causes of mortality of workers exposed to 1,3-butadiene have shown no increases in mortality due to lung cancer, cardiovascular disease and digestive system cancer indicating that 1,3-butadiene has no chronic effects on these organ systems in humans (Delzell et al 2006). A more recent study (Tsai et al, 2003) also showed no evidence of adverse haematological findings associated with exposure to 1,3-butadiene when workers from 2 plants were compared with a non-1,3-butadiene exposed group.

 

Summary

Simple short chain alkanes (i.e methane, ethane, propane, butane, isobutane) can be considered in a similar manner, inhalation exposure is the most relevant route, and current GLP-compliant guideline study data are available for ethane, propane, butane and isobutane which demonstrate low repeat dose toxicity (up to six weeks in duration). These data are supported by studies up to 90 days in duration on C4-C5 mixtures and a 90 day study on liquefied petroleum gas (LPG, main constituent propane and propene), which gave a no observed adverse effect level (NOAEC) of 10,000 ppm, the maximum dose level tested. A consideration of the data available for the alkene, propene, similarly supports a conclusion of low sub-chronic toxicity.

 

The mammalian toxicity effects of this category will be not driven by the content of benzene as the latter is present at levels of <1%.However, Petroleum Gas streams may contain carbon monoxide, levels of which could trigger classification.

 

Carbon Monoxide CAS number 630-08-0

(Classification: CLP - STOT-RE Category 1, H372)

The World Health Organisation published an extensive review of carbon monoxide in 1999 (WHO, 1999, updated 2004). In the human body, the gas reacts readily with haemoglobin to form carboxyhaemoglobin (COHb). Its toxic effects on humans are due to hypoxia, which becomes evident in organs and tissues with high oxygen consumption such as the brain, the heart, exercising skeletal muscle (and the developing foetus).Chronic exposure to low concentrations of carbon monoxide may lead to cardiovascular effects, tiredness, lethargy, headaches, nausea, dizziness, personality changes, memory problems, as well as impairment of visual, auditory or cognitive function.

 

Reference

World Health Organisation, 1999

Environmental Health Criteria 213 (Carbon Monoxide, second edition)

1999, updated 2004

 

Justification for selection of repeated dose toxicity via oral route - systemic effects endpoint:
These streams are gases at room temperature, hence repeated dose toxicity testing via the oral route is not technically feasible.

Justification for selection of repeated dose toxicity inhalation - systemic effects endpoint:
Information available for the short chain alkanes ethane, propane, propene, butane and isobutene, supported by results obtained when testing C4-C5 mixtures and liquefied petroleum gas (main constituent propane and propene), are consistent with a NOAEC of up to 10,000 ppm.

Justification for selection of repeated dose toxicity inhalation - local effects endpoint:
Mild rhinitis (nasal inflammation) and associated epithelial alteration suggesting chronic, low-grade irritation in these rodents was observed following chronic exposure to 5000 (8600 mg/m3) and 10000 ppm (17200 mg/m3) propene.

Justification for selection of repeated dose toxicity dermal - systemic effects endpoint:
These streams are gases at room temperature, hence repeated dose toxicity testing via the dermal route is not technically feasible.

Justification for selection of repeated dose toxicity dermal - local effects endpoint:
These streams are flammable gases at room temperature, hence repeated dose toxicity testing via the dermal route is not technically feasible.

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.

The health effects of ethylene have been evaluated in a number of repeated-dose toxicity studies conducted in rats and no evidence of systemic toxicity has been seen at any concentration. The only treatment-related effects noted in these studies were changes to the upper respiratory tract following repeated exposures of at least 4 weeks duration, which were considered to be a local adaptive response and showed no dose relationship. The findings are considered not to justify classification under CLP.

Levels of benzene and/or carbon monoxide in excess of 1% will trigger classification under CLP. After repeated dose exposure via oral or inhalation routes, benzene causes adverse effects on the haematopoietic system of animals and in humans. Consequently, benzene and carbon monoxide at levels >1% is classified as STOT RE, Cat 2 (H373) or if above >10% as Cat 1 (H372) according to Regulation (EC) No 1272/2008 of the European Parliament.