3-methyl-2-butenal

Repeated dose toxicity: inhalation

Administrative data

Endpoint:

short-term repeated dose toxicity: inhalation

Type of information:

experimental study

Adequacy of study:

key study

Reliability:

1 (reliable without restriction)

Rationale for reliability incl. deficiencies:

other: GLP and guideline study

Data source

Reference

Reference Type:

study report

Title:

Unnamed

Year:

1994

Report date:

1994

Materials and methods

Test guideline

Qualifier:

according to guideline

Guideline:

OECD Guideline 412 (Subacute Inhalation Toxicity: 28-Day Study)

GLP compliance:

yes (incl. QA statement)

Remarks:

testing lab.

Limit test:

no

Test material

Test material information

Constituent 1

Reference substance name:

3-methyl-2-butenal

EC Number:

203-527-6

EC Name:

3-methyl-2-butenal

Cas Number:

107-86-8

Molecular formula:

C5H8O

IUPAC Name:

3-methyl-2-butenal

Details on test material:

- Name of test material (as cited in study report): 3-Methyl-2-butenal, CAS: 107-86-8, Substance No. 90/734-1
- Analytical purity: 98.29%
- Lot/batch No.: 91/ 7
- Physical state: liquid/colorless
- Storage condition of test material: room temperature

Test animals

Species:

rat

Strain:

Wistar

Sex:

male/female

Details on test animals or test system and environmental conditions:

TEST ANIMALS
- Source: Dr . K . Thomae GmbH, D-W7950 Biberach
- Age at study initiation: about 7 weeks
- Fasting period before study: During exposure food and water were withdrawn
- Housing: singly in wire cages (type DK III of Becker, Castrop-Rauxel, Germany)
- Diet: KLIBA rat/mouse/hamster laboratory diet 24-343-4 10 mm pellets ad libitum
- Water: ad libitum
- Acclimation period: yes, not specified


ENVIRONMENTAL CONDITIONS
- Temperature (°C): 20 - 24°C
- Humidity (%): 30 - 70%
- Photoperiod (hrs dark / hrs light): 12/12

Administration / exposure

Route of administration:

inhalation

Type of inhalation exposure:

whole body

Vehicle:

other: unchanged (no vehicle)

Details on inhalation exposure:

GENERATION OF TEST ATMOSPHERE / CHAMBER DESCRIPTION
- Exposure apparatus: glass-steel inhalation chamber, volume V - 1,100 l (manufacturer: BASF Aktiengesellschaft)
- Method of conditioning air:
- piston metering pump (DESAGA )
- continuous infusion pump INFU 362 (INDIGEL, Switzerland)
- glass mixing stage (BASF Aktiengesellschaft )
- glass vaporizer with thermostat (BASF Aktiengesellschaft)
- two-component atomizer (Beckmann )
The test substance was supplied to a vaporizer at a constant rate by means of a metering pump.
- System of generating particulates/aerosols: The generation was carried out using a heated glass evaporator with a downstream mixing unit by means of spraying the substance with a two-component atomizer and compressed air into a counter current of blast air
- Temperature, humidity, pressure in air chamber: A positive pressure was chosen for all test groups during the preflow period .
- Air flow rate: 22 .0 ± 2 m³/hr
- Method of particle size determination:
- exhaust air: 22 .5 ± 2 l/h


TEST ATMOSPHERE
- Brief description of analytical method used: The concentrations of the test groups were analyzed by gas chromatography after absorption of MBA in 2-Propanol
- Samples taken from breathing zone: yes

Analytical verification of doses or concentrations:

yes

Duration of treatment / exposure:

28 days (20 exposures), 6 h/each working day

Frequency of treatment:

5 d/week on working days

Doses / concentrations

Remarks:

Doses / Concentrations:
0, 30, 100, 300 ppm (vapor); corresponding to 0, 0.10, 0.34, and 1.02 mg/l (0, 100, 340, 1020 mg/m3)
Basis:
nominal conc.

No. of animals per sex per dose:

5

Control animals:

yes, concurrent vehicle

Details on study design:

Post-exposure period: none

Examinations

Observations and examinations performed and frequency:

CAGE SIDE OBSERVATIONS: Yes
The behavior and state of health of the test animals were checked on workdays at least 3 times on exposure days and, as a rule, once during the preflow period and the post-exposure observation day.
Mortality: A check for dead animals was made daily .

DETAILED CLINICAL OBSERVATIONS: Yes

BODY WEIGHT: Yes
The body weight of the animals was checked at the beginning of the preflow period, one day before beginning of the exposure period and then, as a rule, once a week . In general, the animals were weighed at the same time of the day . Body weight change was calculated as the difference between body weight on the respective exposure day and body weight on the day before the first exposure . From the individual differences group means were derived .

FOOD CONSUMPTION:
- Food consumption for each animal determined and mean daily diet consumption calculated as g food/kg body weight/day: Yes / No / No data

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: Yes / No / No data

OPHTHALMOSCOPIC EXAMINATION: Yes
At the beginning of the preflow period, the eyes of all animals and at the end of the study the eyes of the animals of test group 0(control group) and of test group 3 (highest concentration) were examined with a focusable hand slit lamp (*) for any changes to the refracting media .

HAEMATOLOGY: Yes
The following parameters were determined in blood with EDTA-K3 as anticoagulant using a particle counter (S Plus model, by Coulter, Krefeld, FRG) :
- leukocytes
- erythrocytes
- hemoglobin
- hematocri t
- mean corpuscular volume
- mean corpuscular hemoglobin
- mean corpuscular hemoglobin concentration
- platelets

CLINICAL CHEMISTRY: Yes
The following parameters were determined:
1 . Enzymes
- alanine aminotransferase
- aspartate aminotransferase
- alkaline phosphatase
- serum-gamma-glutamyltransferase
2 . Blood chemistry
- sodium
- potassium
- chloride
- inorganic phosphate
- calcium
- urea
- creatinine
- glucose
- total bilirubin
- total protein
- albumin
- globulins
- triglycerides
- cholesterol
- magnesium

URINALYSIS: Yes
Blood was taken from the retroorbital venous plexus in the morning from fasted, not anesthetized animals . The blood sampling procedure and the subsequent analysis of the blood and serum samples were carried out in a randomized sequence . The list of randomization instructions was compiled with a computer using a random number generator . For urinalysis the individual animals were transferred to metabolism cages (withdrawal of food and water) and urine was collected overnight . The urine samples were evaluated in a randomized sequence . At each collection period the blood and urine examinations were carried out in the same animals. The assays of blood and serum parameters were performed under internal laboratory quality control conditions with commercial reference controls to assure reliable test results . The results of the clinicochemical and hematological examinations are expressed in units of the International System (SI) .
The specific gravity was determined using a urine refractometer. The sediment was evaluated microscopically.
The following examinations were carried out:
- volume
- color
- turbidity
- nitrite
- pH
- protein
- glucose
- ketones
- urobilinogen
- bilirubin
- blood
- specific gravity
- sediment

Sacrifice and pathology:

GROSS PATHOLOGY: Yes
HISTOPATHOLOGY: Yes

Statistics:

The statistical evaluation of the data was carried out on the computer systems of the Department of Toxicology.

Results and discussion

Results of examinations

Details on results:

(1) Findings in the low dose 30 ppm test group (0.10 mg/l):
Clinical signs were snout wiping, restlessness, eyelid closure at the beginning of the exposure indicating slight, transient sensory irritation. No influence on body weight was noted. No effects on hematological and clinicochemical parameters or urinalysis were noted. No pathological changes noted.

(2) Findings in the intermediate dose 100 ppm test group (0.34 mg/l):
At the beginning of the exposure snout wiping, restlessness, eyelid closure, salivation, ruffled fur, apathy were seen. Animals accustomed to the exposure during the first week and only restlessness and ruffled fur persisted further on. No influence on body weight development was noted. No effects on hematological and clinicochemical parameters or urinalysis were noted. The absolute testes weights were significantly decreased in males (-7.5%). No histopathological changes were observed in the testes. In the respiratory tract, histopathology revealed focal metaplasia of respiratory epithelium into squamous epithelium in the nasal cavity of all males and 3 females, and in the cartilage of the larynx in two males and one female. No other pathological or microscopical changes were noted.

(3) Findings in the high dose 300 ppm test group (1.02 mg/l):
Clear signs of irritation over the entire exposure period such as eyelid closure, salivation, ruffled fur, apathy. The symptoms disappeared rapidly after each exposure. Body weight development was significantly retarded in males (-59.8% compared to controls, days 0 - 28) and slightly impaired in females (-39.7% compared to controls, days 0 - 28). No ophthalmologic changes were detected. Increased number of polymorphonuclear neutrophils accompanied by a decrease in leucocytes in females was interpreted as a sign of inflammation in the respiratory tract. No effects on urinalysis were noted. Gross pathology revealed a significantly decreased terminal body weight in males (-14.1%). The absolute testes weights were significantly decreased (-6.1%) whereas the relative testes weights were significantly increased (+12.1%). No treatment-related histopathological changes were observed in this organ. Histopathology revealed purulent inflammation in the nasal cavity of all males and some females. Focal metaplasia of respiratory epithelium into squamous epithelium in the nasal cavity and in larynx of all males and site specifically in all or some females. In one male and one female atrophy of the olfactory epithelium (level II) was noted.

The clinical findings at low concentrations during the first week of exposure were not interpreted as toxic effects but were attributed to the marked odor of the test substance.

Irritation of the upper respiratory tract was seen after repeated exposure (4 weeks, 5d/wk, 6 hrs/d) at or above 100 ppm 3-methyl-2-butenal (= 0.34 mg/l) as evidenced by the concentration dependend focal metaplasia of respiratory epithelium into squamous epithelium. Additionally, atrophy of the olfactory epithelium was noted in 2/10 animals at 300 ppm.

No ophthalmologic changes were seen up to and including 300 ppm.

No systemic toxicity was seen at concentrations up to 300 ppm (= 1.02 mg/l). The observed decreases in body weights were assigned to the discomfort of the purulent rhinitis found in all high dose male and some female animals.

A substance related effect for the significantly decreased absolute testes weights at 100 ppm and 300 ppm and the significantly increased relative testes weights at 300 ppm was regarded as unlikely as there were no histopathological correlates in this organ. At least for dose group 3 (300 ppm), the weight changes seemed to be a consequence of the reduction of the terminal body weight.

Effect levels

1

Dose descriptor:

NOAEC

Effect level:

300 ppm

Sex:

male/female

Basis for effect level:

other: systemic toxicity

2

Dose descriptor:

NOEC

Effect level:

30 ppm

Sex:

male/female

3

Dose descriptor:

NOAEC

Effect level:

0.1 mg/L air

Sex:

male/female

4

Dose descriptor:

LOAEC

Effect level:

100 ppm

Sex:

male/female

Basis for effect level:

other: for local irritation of the upper respiratory tract

Target system / organ toxicity

Critical effects observed:

not specified

Applicant's summary and conclusion

Conclusions:

Concentrations at or above 100 ppm (= 0.34 mg/l) of MBA vapor inhaled 6 hours per working day for 4 weeks caused irritation of the upper respiratory tract as demonstrated mainly by the concentration dependent focal metaplasia of respiratory epithelium into squamous epithelium. No systemically toxic effects occurred up to the high concentration of 300 ppm (= 1.02 mg/1).