Renewable hydrocarbons (kerosene type fraction)

Acute Toxicity: inhalation

Administrative data

Endpoint:

acute toxicity: inhalation

Type of information:

experimental study

Adequacy of study:

key study

Study period:

Experimental Starting Date: 29 January 2013 Experimental Completion Date: 01 July 2013

Reliability:

2 (reliable with restrictions)

Rationale for reliability incl. deficiencies:

other: see 'Remark'

Remarks:

Study on supporting substance (with carbon range C8-26), conducted in compliance with agreed protocols, with no or minor deviations from standard test guidelines and/or minor methodological deficiencies, which do not affect the quality of the relevant results. The study report was conclusive, done to a valid guideline and the study was conducted under GLP conditions. It is appropriate to read this result across to 'Kerosine (Fischer-Tropsch), full range, C8-16 - branched and linear', since the test substance 'Distillates (Fischer-Tropsch), C8-26 - branched and linear' is produced using the same Fischer-Tropsch process and contains all of the same hydrocarbon constituents, but the product fraction additionally contains longer carbon chain lengths, up to C26, compared to C16 in 'Distillates (Fischer-Tropsch), full range, C8-16 - branched and linear'.

Data source

Reference

Reference Type:

study report

Title:

Unnamed

Year:

2013

Report date:

2013

Materials and methods

Test guideline

Qualifier:

according to guideline

Guideline:

OECD Guideline 436 (Acute Inhalation Toxicity: Acute Toxic Class Method)

Deviations:

no

GLP compliance:

yes (incl. QA statement)

Test type:

acute toxic class method

Limit test:

yes

Test material

Test material information

Constituent 1

Reference substance name:

Distillates (Fischer-Tropsch), C8-26-branched and linear

IUPAC Name:

Distillates (Fischer-Tropsch), C8-26-branched and linear

Constituent 2

Reference substance name:

Distillates (Fischer-Tropsch), C8-26, branched and linear

Cas Number:

848301-67-7

Molecular formula:

C8H18 to C26H54

IUPAC Name:

Distillates (Fischer-Tropsch), C8-26, branched and linear

Test material form:

other: liquid

Details on test material:

Description: Clear colorless liquid
Storage conditions: Room temperature, in the dark

Test animals

Species:

rat

Strain:

other: RccHan™ : WIST strain rats

Sex:

male/female

Details on test animals or test system and environmental conditions:

Male and female RccHan™ : WIST strain rats were supplied by reputable supplier. On receipt the animals were randomly allocated to cages. After an acclimatization period of at least five days the animals were given a number unique within the study by ear punching and a number written on a color coded cage card. At the start of the study the animals were approximately eight to twelve weeks old and within the weight range of 200g to 354g*.

The females were nulliparous and non-pregnant.

The animals were housed in groups of up to three by sex in solid-floor polypropylene cages with stainless steel lids, furnished with softwood flakes and provided with environmental enrichment items: wooden chew blocks and cardboard “fun tunnels”.

With the exception of the exposure period, free access to mains drinking water and food was allowed throughout the study. The diet, drinking water, bedding and chew blocks are routinely analyzed and are considered not to contain any contaminants that could reasonably be expected to affect the purpose or integrity of the study.

The environmental controls were set to achieve values of 19 – 25 °C and 30 – 70 % relative humidity. The rate of air exchange was at least fifteen changes per hour and the lighting was controlled to give twelve hours continuous light and twelve hours darkness. The animals were retained in this accommodation at all times except during the exposure period.

* = One male animal from Group 2 was slightly outside the weight range specified in the Study Plan (200g to 350g).
This deviation was considered not to affect the purpose or validity of this study.

Administration / exposure

Route of administration:

inhalation: aerosol

Type of inhalation exposure:

nose only

Vehicle:

clean air

Details on inhalation exposure:

Atmosphere Generation
The test item was aerosolized using a glass concentric jet nebulizer located at the top of the exposure chamber. The nebulizer was connected to a plastic syringe attached to an infusion pump, which provided a continuous supply of test item under pressure, and to a metered compressed air supply.

Compressed air was supplied by means of an oil free compressor and passed through a water trap and respiratory quality filters before it was introduced to the nebulizer.

The cylindrical exposure chamber had a volume of approximately 30 liters (dimensions: 28 cm diameter x 50 cm high). The concentration within the exposure chamber was controlled by adjusting the rate of the infusion pump. The extract from the exposure chamber passed through a ‘scrubber’ trap and was connected with a high efficiency filter to a metered exhaust system. The chamber was maintained under negative pressure. A diagram of the dynamic (continuous flow) system employed is shown in the attached Figure 1.

Homogeneity of the test atmosphere within the chamber was not specifically determined during this study. Chambers of the same design have been fully validated and shown to produce evenly distributed atmospheres in the animals’ breathing zone with a wide variety of test items (Green J D et al, 1984). Prior to the start of the study, test item atmospheres were generated within the exposure chamber.

During this characterization period test item input rates and air flow settings were varied to achieve the required atmospheric conditions.

Exposure Procedure
On the day of exposure (Group 1) and prior to the day of exposure (Group 2) each rat was acclimatized (for approximately 2 hours) to a tapered polycarbonate restraining tube. During the exposure period, each rat was individually held in a tapered, polycarbonate restraining tube fitted onto a single tier of the exposure chamber and sealed by means of a rubber ‘O’ ring. Only the nose of each animal was exposed to the test atmosphere.

Following an appropriate equilibration period two groups, each of six rats (three males and three females), were subjected to a single exposure to the test item for a period of four hours. Based on the expected toxicity of the test item, a target concentration of 5.0 mg/L was used for the first exposure. The second concentration was selected after consideration of the results of the first exposure.

Exposure Chamber Temperature and Relative Humidity
The temperature and relative humidity inside the exposure chamber were measured by an electronic thermometer/humidity meter located in a vacant port in the animals’ breathing zone of the chamber and recorded every thirty minutes throughout the four-hour exposure period. Individual values are given in the attached Table 14 and 15.

Exposure Chamber Oxygen Concentration
Oxygen levels within the exposure chamber were measured by an electronic oxygen analyzer located in a port in the animals breathing zone during the four-hour exposure period. The test atmosphere was generated to contain at least 19% oxygen. Individual values are given in the attached Table 16.

Analytical verification of test atmosphere concentrations:

yes

Duration of exposure:

4 h

Concentrations:

Group 1 mean achieved atmosphere concentration of 5.61 mg/L group 2 mean achieved atmosphere concentration 5.11 mg/L

No. of animals per sex per dose:

Two groups of six RccHan™ : WIST strain rats (three males and three females)

Control animals:

no

Details on study design:

Clinical Signs
All animals were observed for clinical signs at hourly intervals during exposure, immediately on removal from the restraining tubes at the end of exposure, one hour after termination of exposure and subsequently once daily for up to fourteen days. Any evidence of overt toxicity was recorded at each observation.

Body Weight
Individual body weights were recorded on arrival, prior to treatment on the day of exposure and on Days 1, 3, 7 and 14.

Necropsy
At the end of the fourteen day observation period the animals were killed by intravenous overdose of sodium pentobarbitone. All animals were subjected to a full external and internal examination, and any macroscopic abnormalities were recorded. The respiratory tract was subjected to a detailed macroscopic examination for signs of irritancy or local toxicity.

Statistics:

Evaluation of Data
Data evaluations included the relationship, if any, between the animals’ exposure to the test item and the incidence and severity of all abnormalities including behavioral and clinical observations, necropsy findings, body weight changes, mortality and any other toxicological effects. Using the mortality data obtained, an estimate of the acute inhalation median lethal concentration (LC50) of the test item was made.

Results and discussion

Preliminary study:

Dose Group 1 – 5.61 mg/L
During exposure all animals exhibited increased respiratory rate and there were occasional instances of labored respiration. On removal from the chamber all animals exhibited increased respiratory rate and labored respiration. One hour post-exposure, little or no change in the condition of the animals was noted.

One day after exposure, one male and one female animal were found dead. All surviving animals exhibited increased respiratory rate, hunched posture and pilo-erection. Isolated occurrences of labored respiration, splayed gait, tip-toe gait and ptosis were also noted in one female animal. Observations gradually receded over the recovery period such that surviving animals recovered to appear normal from Days 8 to 9 post-exposure.

Effect levels

Sex:

male/female

Dose descriptor:

LC50

Effect level:

> 5 mg/L air

Based on:

test mat.

Exp. duration:

4 h

Remarks on result:

other: 95 % CL not reported.

Mortality:

For full details please see attached tables 6 to 9

Group 1: One male and one female animal were found dead from a group of six rats exposed to a mean achieved atmosphere concentration of 5.61 mg/L one day after exposure

Group 2: One female animal was found dead from a group of six rats exposed to a mean achieved atmosphere concentration of 5.11 mg/L one day after exposure

Clinical signs:

other: Individual clinical observations are given in the attached Tables 6 to 9. Dose Group 1 – 5.61 mg/L During exposure all animals exhibited increased respiratory rate and there were occasional instances of labored respiration. On removal from the chamber a

Body weight:

Individual body weights, together with body weight changes, are given in the attached Tables 10 and 11.

Group 1 – All surviving animals exhibited bodyweight losses on Day 1 post-exposure. With the exception of one female animal which showed no body weight gain from Days 1 to 3 post-exposure, reasonable bodyweight development was noted in all surviving animals during the remainder of the recovery period.
Group 2 – All surviving animals exhibited bodyweight losses on Day 1 post-exposure.
Reasonable bodyweight development was noted in all animals during the remainder of the recovery period.

Gross pathology:

Individual necropsy findings are given in the attached Tables 12 and 13.

Pale or dark patches on the lungs were noted at necropsy amongst four surviving animals from Group 1 and in two out of five surviving animals from Group 2 at the end of the fourteen day recovery period.

The following macroscopic abnormalities were detected in the animals that died during the course of the study at necropsy:
Lungs – abnormally dark or dark patches;
Stomach – gaseous distension;
Large Intestine – gaseous distension.

Due to the observations noted it is considered that the death noted during the study may have been mainly attributable to local toxicity.

Dark patches in the lungs and abnormally dark lungs in a proportion of animals in both treatment groups were associated with congestion and hemorrhage at histopathological examination, and were considered to be treatment-related.

Other findings:

A complete histopathology phase report is presented in the attached Appendix 3.

Histopathology
The following treatment-related findings were present in the lungs of animals surviving to
terminal sacrifice:
- minimal to moderate congestion
- minimal to slight hemorrhage
- minimal to moderate edema
- minimal to slight alveolar macrophages
- minimal acute alveolar inflammation
- minimal acute perivascular inflammation
The following treatment-related findings were present in the lungs of premature decedent animals:
- moderate congestion
- slight hemorrhage
- minimal to slight alveolar macrophages
- slight acute alveolar inflammation
There were no findings in the trachea.

Any other information on results incl. tables

Signs of hunched posture and pilo-erection are commonly seen in animals for short periods on removal from the chamber following 4-hour inhalation studies. Wet fur and red/brown staining around the eyes are commonly recorded both during and for a short period after exposure. These observations are considered to be associated with the restraint procedure and, in isolation, are not indicative of toxicity.

Exposure Chamber Concentration The test atmosphere (Group 1) was sampled seventeen times during the exposure period and the actual concentration of the test item calculated.

The actual concentration (Group 2) of the test item was determined by gas chromatography (GC). The test atmospheres were sampled after theoretical chamber equilibration and then at approximately half-hourly intervals during the exposure period.

The mean values obtained for each group were:

Group number	Atmosphere Concentration
	Mean Achieved (mg/L)	Standard Deviation	Nominal (mg/L)
1	5.61	0.23	15.3
2	5.11	0.09	11.7

The exposure chamber concentrations are given in the attached Tables 1 and 2 and are presented graphically in the attached Figures 2 and 3. The chamber flow rate was maintained at 60 L/min providing 120 air changes per hour. The theoretical chamber equilibration time (T99) was 3 minutes* (Silver, 1946).

Particle Size Distribution

The particle size analysis of the atmosphere drawn from the animals’ breathing zone, was as follows:

Group number	Mean Achieved Atmosphere Concentration (mg/L)	Mean Mass Median Aerodynamic Diameter (µm)	Inhalable Fraction (% <4µm)	Geometric Standard Deviation
1	5.61	2.78	65.7	2.48
2	5.11	2.16	77.5	2.27

The particle size distribution data is presented in the attached Tables 3 and 4 and graphs showing the distribution are presented in the attached Figures 4 to 7.

* = The test atmosphere was generated for 7 minutes and 14 minutes prior to animal insertion (for Groups 1 and 2

respectively) to ensure test item concentration was being achieved.

Applicant's summary and conclusion

Interpretation of results:

not classified

Remarks:

Migrated information Criteria used for interpretation of results: EU

Conclusions:

Two deaths occurred in a group of six rats exposed to a mean achieved atmosphere concentration of 5.61 mg/L, whereas, one death occurred at a mean achieved atmosphere concentration of 5.11 mg/L. It was therefore considered that the acute inhalation median lethal concentration (4 hr LC50) of the closely related substance 'Distillates (Fischer-Tropsch), C8-26-branched and linear', in the RccHanTM : WIST strain rat, was >5 mg/L.

The microscopic findings observed in the lungs (alveolar and perivascular inflammation, edema, congestion and hemorrhage) were all consistent with hydrocarbon aspiration induced inflammation (i.e. a chemical pneumonitis).

Executive summary:

Introduction

A study was performed to assess the acute inhalation toxicity of the test item. The method used was compatible with that described in the OECD Guidelines for Testing of Chemicals (2009) No. 436 “Acute Inhalation Toxicity – Acute Toxic Class Method”.

 

Methods.......

Two groups of six RccHan™ : WIST strain rats (three males and three females) were exposed to an aerosol atmosphere. The animals were exposed for four hours using a nose only exposure system, followed by a fourteen day observation period.

 

Results……..

The mean achieved atmosphere concentration was as follows:

Group number	Atmosphere Concentration
	Mean Achieved (mg/L)	Standard Deviation	Nominal (mg/L)
1	5.61	0.23	15.3
2	5.11	0.09	11.7

 

Group number	Mean Achieved Atmosphere Concentration (mg/L)	Mean Mass Median Aerodynamic Diameter (µm)	Inhalable Fraction (% <4 µm)	Geometric Standard Deviation
1	5.61	2.78	65.7	2.48
2	5.11	2.16	77.5	2.27

 

Group Number	Mean Achieved Atmosphere Concentration (mg/L)	Deaths
		Male	Female	Total
1	5.61	1/3	1/3	2/6
2	5.11	0/3	1/3	1/6

 

Clinical Observations

Common abnormalities noted during the study included increased respiratory rate, labored respiration, hunched posture, pilo-erection and wet fur. There were occasional instances of tiptoe gait and red/brown staining around the eyes, isolated occurrences of splayed gait, ptosis and red/brown staining around the snout were also noted. Surviving Group 1 animals recovered to appear normal from Days 8 to 9 post exposure. Surviving Group 2 animals recovered to appear normal from Days 7 to 8 post-exposure.

 

Body Weight

Group 1 – All surviving animals exhibited bodyweight losses on Day 1 post-exposure. With the exception of one female animal which showed no body weight gain from Days 1 to 3 postexposure, reasonable bodyweight development was noted in all surviving animals during the remainder of the recovery period.

 

Group 2 – All surviving animals exhibited bodyweight losses on Day 1 post-exposure. Reasonable bodyweight development was noted in all animals during the remainder of the recovery period.

 

Necropsy

Pale or dark patches on the lungs were noted at necropsy amongst four surviving animals from Group 1 and in two out of five surviving animals from Group 2 at the end of the fourteen day recovery period.

 

The following macroscopic abnormalities were detected in the animals that died during the course of the study at necropsy:

Lungs – abnormally dark or dark patches;

Stomach – gaseous distension;

Large Intestine – gaseous distension.

Due to the observations noted it is considered that the death noted during the study may have been mainly attributable to local toxicity.

Dark patches in the lungs and abnormally dark lungs in a proportion of animals in both treatment groups were associated with congestion and hemorrhage at histopathological examination, and were considered to be treatment-related.

 

Histopathology

The following treatment-related findings were present in the lungs of animals surviving to terminal sacrifice:

- minimal to moderate congestion

- minimal to slight hemorrhage

- minimal to moderate edema

- minimal to slight alveolar macrophages

- minimal acute alveolar inflammation

- minimal acute perivascular inflammation

The following treatment-related findings were present in the lungs of premature decedent

animals:

- moderate congestion

- slight hemorrhage

- minimal to slight alveolar macrophages

- slight acute alveolar inflammation

There were no findings in the trachea.

 

Conclusion

Two deaths occurred in a group of six rats exposed to a mean achieved atmosphere concentration of 5.61 mg/L, whereas, one death occurred at a mean achieved atmosphere concentration of 5.11 mg/L. It was therefore considered that the acute inhalation median lethal concentration (4 hr LC50) of GTL diesel (Distillates (Fischer-Tropsch), C8-26-branched and linear), in the RccHanTM: WIST strain rat, was >5 mg/L.

The microscopic findings observed in the lungs (alveolar and perivascular inflammation, edema, congestion and hemorrhage) were all consistent with hydrocarbon aspiration induced inflammation (i.e. a chemical pneumonitis).