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Repeated dose toxicity: inhalation

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Administrative data

Endpoint:
short-term repeated dose toxicity: inhalation
Type of information:
experimental study
Adequacy of study:
key study
Study period:
In life phase: 28 August 2007 to 24 October 2007
Reliability:
1 (reliable without restriction)
Rationale for reliability incl. deficiencies:
other: GLP compliant, guideline study, available as unpublished report, no restrictions, fully adequate for assessment

Data source

Reference
Reference Type:
study report
Title:
Unnamed
Year:
2008
Report Date:
2008

Materials and methods

Test guidelineopen allclose all
Qualifier:
according to
Guideline:
OECD Guideline 412 (Subacute Inhalation Toxicity: 28-Day Study)
Deviations:
no
Qualifier:
according to
Guideline:
EPA OPPTS 870.3465 (90-Day Inhalation Toxicity)
Deviations:
no
GLP compliance:
yes (incl. certificate)
Limit test:
no

Test material

Reference
Name:
Unnamed
Type:
Constituent
Type:
Constituent
Details on test material:
- Analytical purity: 99.68%
- Physical state: Volatile colourless to brown liquid
- Lot/batch No.: 7883
- Expiration date of the lot/batch: 18 June 2008
- Storage conditions: In tightly sealed container (not plastic), in cool, dark, well-ventilated area

Test animals

Species:
rat
Strain:
Sprague-Dawley
Sex:
male/female
Details on test animals and environmental conditions:
TEST ANIMALS
- Source: Charles River, Deutschland, Sulzfeld, Germany
- Age at study initiation: Approximately 6 weeks
- Weight at study initiation: 315.7 g (males) and 214.3 g (females)
- Fasting period before study: No
- Housing: 5 per sex in macrolon cages
- Diet: Rat and mouse No. 3 breeding diet RM3 (Special Diet Services, Witham, England) ad libitum except during exposure
- Water: ad libitum except during exposure
- Acclimation period: 10 days for males, 11 days for females

ENVIRONMENTAL CONDITIONS
- Temperature: 20-24°C
- Humidity: 40-70%
- Air changes (per hr): Not reported
- Photoperiod: 12 hrs dark / 12 hrs light

IN-LIFE DATES: From: 28 August 2007 To: 24 October 2007

Administration / exposure

Route of administration:
inhalation: vapour
Type of inhalation exposure:
nose only
Vehicle:
air
Details on inhalation exposure:
GENERATION OF TEST ATMOSPHERE / CHAMBER DESCRIPTION
- Exposure apparatus: A cylindrical column, surrounded by a transparent cylinder with a volume of 50 L. The test atmosphere was introduced at the top of the central column and exhausted at the bottom. A similar unit was used for the control group exposures.
- Method of holding animals in test chamber: In plastic holders (Battelle), postioned radially through the outer hood around the central column (males and females alternately)
- Source and rate of air: For each exposure level a small flow of dry nitrogen, controlled by a mass flow controller, was bubbled through a small glass bottle with liquid furfural. The bottle was maintained at 18.5°C. The flow was diluted with a controlled amount of humidified air and directed to the inlets of the exposure units at the top and and exhausted at the bottom of the units
- Temperature, humidity, pressure in air chamber: Temperature: 20.3 and 24.2°C. Relative humidity: 34 and 56%. Oxygen concentrations were 20.7%, 20.7%, 20.7% and 20.6% for the low, mid, high and top concentration, respectively.

TEST ATMOSPHERE
- Brief description of analytical method used: The concentrations of the test material in the test atmospheres was measured by off-line chemical analysis. Test atmosphere samples were passed through single silica-DNPH cartridges containing 2,4-dinitrophenylhydrazine. The DNPH-furfural complex in the cartridge was eluted with acetonitrile and its concentration was measured by HPLC analysis.
- Samples taken from breathing zone: yes
- The nominal concentration was determined once a week by measuring the amount of test material used (by weight) divided by the volume of test atmosphere generated. The generation efficiency was calculated from the actual concentration and the nominal concentration.
Analytical verification of doses or concentrations:
yes
Details on analytical verification of doses or concentrations:
The mean actual concentrations as measured three times during each exposure by offline chemical analysis, were 1.93 (± 0.13), 3.70 (± 0.31), 7.38 (±0.60) and 17.05 (±1.25) mg/m3 for the low, mid, high and top concentration respectively.
The nominal concentration was on average 2.3 (± 0.3), 4.9 (± 0.6), 10.0 (± 0.4), and 24.1 (± 0.6) mg/m3 for the low, mid, high and top concentration, respectively. This indicates generation efficiencies of 84.3%, 75.5%, 73.7% and 70.7% for the low, mid, high and top concentration respectively.
Duration of treatment / exposure:
28 day (20 exposures)
Frequency of treatment:
6 hours/day, 5 days/week
Doses / concentrationsopen allclose all
Remarks:
Doses / Concentrations:
0, 2, 4, 8 or 20 mg/m3
Basis:
nominal conc.
Remarks:
Doses / Concentrations:
0, 1.93, 3.70, 7.38 and 17.05 mg/m3
Basis:
analytical conc.
No. of animals per sex per dose:
10 for main study plus 10/sex/group for 4 week recovery.
Control animals:
other: exposed to clean dry air
Details on study design:
- Post-exposure recovery period in satellite groups: 28 days

Examinations

Observations and examinations performed and frequency:
CAGE SIDE OBSERVATIONS: Yes
- Time schedule: At least once daily

DETAILED CLINICAL OBSERVATIONS: Yes
- Time schedule: Daily

BODY WEIGHT: Yes
- Time schedule for examinations: On the day prior to exposure and weekly thereafter

FOOD CONSUMPTION:
- Food consumption for each animal determined and mean daily diet consumption calculated as g food/kg body weight/day: Yes
- Food consumption was measured for 3 successive periods of 7 days, followed by one period of 6 days. For the recovery groups this was followed by another 3 successive periods of 7 days and one period of 6 days.

FOOD EFFICIENCY:
- Body weight gain in kg/food consumption in kg per unit time X 100 calculated as time-weighted averages from the consumption and body weight gain data: No

WATER CONSUMPTION: No

OPHTHALMOSCOPIC EXAMINATION: Yes
- Time schedule for examinations: Prior to the start of the first exposure in all animals of the recovery groups and after the exposure period in animals of the control and top concentration of the recovery groups (groups 1 and 5). Eye examinations were carried out using an ophthalmoscope after induction of mydriasis by a solution of atropine sulphate.

HAEMATOLOGY: Yes
- Time schedule for collection of blood: Blood samples were taken at necropsy from the abdominal aorta of rats whilst under Nembutal® anaesthesia. K2-EDTA was used as anticoagulant
- Animals fasted: Yes
- How many animals: All animals of the main study
- Parameters checked: Haemoglobin, packed cell volume, red blood cell count, reticulocytes, total white blood cell count, differential white blood cell count - prothrombin time, thrombocyte count, mean corpuscular volume, mean corpuscular haemoglobin, mean corpuscular haemoglobin concentration
- Haematology was not conducted in the recovery groups because no significant effects in these parameters were found in the main study

CLINICAL CHEMISTRY: Yes
- Time schedule for collection of blood: Blood samples were taken at necropsy from the abdominal aorta of rats whilst under Nembutal® anaesthesia. Heparin was used as anticoagulant
- Animals fasted: Yes
- How many animals: All animals of the main study
- Parameters checked: Alkaline phosphatase activity, aspartate aminotransferase activity, alaninc aminotransferase activity, gamma glutamyl transferase activity, albumin, bilirubin, calcium, cholesterol, chloride, creatinine, phospholipids, potassium, sodium, total protein, triglyceride, ratio albumin to globulin, urea, inorganic phosphate and fasting glucose
- Total protein, albumin and the albumin/globulin ratio were also determined for the female recovery groups

URINALYSIS: No

NEUROBEHAVIOURAL EXAMINATION: No
Sacrifice and pathology:
GROSS PATHOLOGY: Yes
- All animals of the main study after 28 days, and the recovery animals after a 28 day post-exposure period.

Organ WEIGHTS: Yes
- The following organs were weighed (paired organs together) as soon as possible after dissection: Adrenals, brain, heart, kidneys, liver, spleen, testes, thymus, thyroid (with parathyroids), lungs with trachea and larynx, ovaries, uterus and epididymides

HISTOPATHOLOGY: Yes
- Adrenals, aorta, * axillary lymph nodes, brain (brain stem, cerebrum and cerebellum), caecum, colon, epididymides, eyes (with optic nerve), exorbital lachrymal glands, * femur with joint, heart, kidneys, liver, lungs/trachea/larynx, mammary glands (females), mandibular (cervical) lymph nodes, nasal passages (with teeth), nerve-peripheral (sciatic nerve), oesophagus, ovaries, pancreas, parathyroids, pharynx, * parotid salivary glands, pituitary, prostate, rectum, seminal vesicles with coagulating glands, * skeletal muscle (thigh), skin (flank), small intestines (duodenum, ileum, jejunum), spinal cord (three levels), spleen, sternum with bone marrow, stomach (glandular, non-glandular), sublingual salivary glands, submaxillary salivary glands, testes, thymus, thyroid, tracheobronchial (mediastinal) lymph nodes, urinary bladder, uterus (with cervix), and all relevant gross lesions
* The tissues marked with an asterisk were preserved but not processed for histopathological examination, because histological examination was considered not to be necessary on the basis of the results of gross observations.
Statistics:
Body weights: one-way analysis of covariance (covariate: body weight on day 0) followed by Dunnett's multiple comparison test; food consumption, red blood cell and coagulation variables, total white blood cell counts, absolute differential white blood cell counts, clinical chemistry values and organ weights: one-way analysis of variance (Anova) followed by Dunnett's multiple comparison test; reticulocytes and relative differential white blood cell counts: Kruskal-Wallis non-parametric Anova followed by Mann-Whitney U-tests; histopathological changes: Fisher's exact probability test.

Results and discussion

Results of examinations

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
Haematological findings:
no effects observed
Clinical biochemistry 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:
effects observed, treatment-related
Details on results:
HISTOPATHOLOGY: NON-NEOPLASTIC
- Microscopic examination of organs and tissues of animals sacrificed at the end of the treatment period revealed exposure-related inflammatory changes in the nasal cavity of 3 males and 5 females of the top concentration group. These changes were localized at level 3 of the nasal cavity at the inner side of the nasoturbinates and characterized by (very) slight transitional respiratory epithelial cell hyperplasia associated with a (very) slight mixed inflammatory cell infiltrate. In one male and one female similar changes were observed at the same location at a more anterior level, level 2.
- Microscopic examination of level 3 of the nasal cavity of control and top concentration rats of the recovery group revealed no abnormalities.

Effect levels

open allclose all
Dose descriptor:
NOAEC
Remarks:
systemic toxicity
Effect level:
20 mg/m³ air
Sex:
male/female
Basis for effect level:
other: no systemic effects at highest dose tested
Dose descriptor:
NOAEC
Remarks:
local irritant effects
Effect level:
8 mg/m³ air
Sex:
male/female
Basis for effect level:
other: microscopic pathology of nasal cavity (hyperplasia and inflammatory cell infiltration) at 20 mg/m3 which recovered within 4 weeks

Target system / organ toxicity

Critical effects observed:
not specified

Applicant's summary and conclusion

Conclusions:
Inhalation exposure to furfural 6 hours/day, 5 days/week for 4 weeks resulted in no evidence of systemic toxicity at concentrations up to 20 mg/m3. Local irritant effects (hyperplasia of transitional respiratory epithelia and inflammatory cell infiltration) was seen in both sexes at 20 mg/m3. The NOAEC for local effects was 8 mg/m3.
Executive summary:

In a 28 day nose-only inhalation study, male and female Sprague Dawley rats were exposed levels of furfural at 0, 2, 4, 8 or 20 mg/m3 for 6 hours/day, 5 days/week. Clinical observations, food consumption, body weight, haematological and clinical chemistry parameters were measured. At study termination, the animals were examined post mortem and selected organs weighed. Selected tissues were examined histopathologically. Additional animals were similarly exposed and kept for an observation period of 28 days.

No evidence of systemic toxicity was seen at concentrations up to 20 mg/m3. Local irritant effects (hyperplasia of transitional respiratory epithelia and inflammatory cell infiltration) was seen in both sexes at 20 mg/m3.

The No-Observed-Adverse-Effect-Concentration (NOAEC) in rats exposed to furfural for 6 hours/day, 5 days/week during a period of 28 days was 8 mg/m3.