2,2'-azobis[4-methoxy-2,4-dimethylvaleronitrile]

Acute Toxicity: inhalation

Administrative data

Endpoint:

acute toxicity: inhalation

Type of information:

experimental study

Adequacy of study:

key study

Study period:

The study was preformed: 08-Jan-2016 until 14 Mar. 2016. The review of the final report was completed on 08- Aug. 2016.

Reliability:

1 (reliable without restriction)

Rationale for reliability incl. deficiencies:

guideline study

Data source

Reference

Reference Type:

study report

Title:

Unnamed

Report date:

2016

Materials and methods

Test guideline

Qualifier:

according to guideline

Guideline:

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

GLP compliance:

yes (incl. QA statement)

Test type:

acute toxic class method

Test material

Test material information

Constituent 1

Reference substance name:

2,2'-azobis[4-methoxy-2,4-dimethylvaleronitrile]

EC Number:

239-593-8

EC Name:

2,2'-azobis[4-methoxy-2,4-dimethylvaleronitrile]

Cas Number:

15545-97-8

Molecular formula:

C16H28N4O2

IUPAC Name:

2,2'-azobis[4-methoxy-2,4-dimethylvaleronitrile]

impurity 1

Reference substance name:

IUPAC-name not available, the reference substance may consist of one or more impurities

Molecular formula:

not applicable

IUPAC Name:

IUPAC-name not available, the reference substance may consist of one or more impurities

Test material form:

solid: particulate/powder

Test animals

Species:

rat

Strain:

Wistar

Sex:

male/female

Details on test animals or test system and environmental conditions:

Animals:
Young adult animals were selected (approximately 9 weeks old).
Animals used within the study were of approximately the same age and body weight variation did not exceed +/- 20% of the sex mean.
Environment:
Environmental controls for the animal room were set to maintain 18 to 24°C, a relative humidity of 40 to 70%, at least 10 air changes/hour, and a 12-hour light/12-hour dark cycle. Any variations to these conditions were maintained in the raw data and had no effect on the outcome of the study

Administration / exposure

Route of administration:

inhalation: aerosol

Type of inhalation exposure:

nose only

Vehicle:

air

Mass median aerodynamic diameter (MMAD):

5.8 µm

Geometric standard deviation (GSD):

1.9

Remark on MMAD/GSD:

The Mass Median Aerodynamic Diameter (MMAD) and geometric standard deviation (gsd) were determined twice during the exposure period. The MMAD was 5.8 μm (gsd 1.9) at both occasions.
Multiple attempts were made to reduce the particle size during method development, either by lowering the concentration, using particle separators into the set-up or applying different grinding techniques, so the size obtained was considered the smallest that could be technically be achieved.

Details on inhalation exposure:

The test atmosphere generation was based on the method developed during extensive trial generations. Trial generation results showed that it was not possible to reduce the particle size by grinding or milling, since the test item turned into a paste. Therefore the test item was sieved prior to use. The generation was performed at the technically maximum attainable concentration.
The test item was passed through a steel mesh sieve (grid size 750 um) prior to use. Due to technical difficulties, part of the study (from approximately 90 minutes after the start onwards) had to be performed with test item stored at room temperature (protected from light) for six days

Analytical verification of test atmosphere concentrations:

not specified

Remarks:

Due to the technical difficulties, the nominal concentration (amount of test item used divided by the volume of pressurized air used) and the generation efficiency (ratio of actual and nominal concentration) could not be determined.

Duration of exposure:

4 h

Concentrations:

Nominal Concentration
Due to the technical complications, the weight of the amount of test item could not be determined. As a consequence, the nominal concentration (amount of test item used divided by the volume of pressurized air used) and the generation efficiency (ratio of actual and nominal concentration) could not be calculated. Since these values are not so meaningful for dust generations, it was considered that the study integrity was not affected by this. The volume of air was calculated from the average air flow (measured by means of thermal mass flow meters and was recorded regularly, preferably in 30 minute intervals) and the exposure time.
Actual Concentration
The actual concentration was determined twenty-six times during the exposure period. Samples were drawn from the test atmosphere through a tube mounted in one of the free animal ports of the exposure chamber. Samples were drawn through a glass fiber filter. The collected amount of the test item in the air sample was measured gravimetrically. Sample volumes were measured by means of a dry gas meter Subsequently the time-weighted mean concentration with the standard deviation was calculated. The actual time-weighted mean concentration was 3.19 ± 0.08 mg/L.
Particle Size Characterization
The particle size distribution was characterized twice during the exposure period. The samples were drawn (2 L/min) from the test atmosphere through a tube mounted in one of the free animal ports of one of the middle section of the exposure chamber. The samples were collected with an 8 stage Marple personal cascade impactor containing fiber glass filters and a fiber glass back-up filter. Amounts of test item collected were measured gravimetrically. Subsequently the Mass Median Aerodynamic Diameter (MMAD) and the Geometric Standard Deviation (GSD) were determined.

No. of animals per sex per dose:

3 per sex

Control animals:

no

Results and discussion

Effect levels

Key result

Sex:

male/female

Dose descriptor:

LC50

Effect level:

3.19 mg/L air

Exp. duration:

4 h

Mortality:

No mortality occurred.

Clinical signs:

other: During exposure, slow breathing was seen (not presented in the table). After exposure, hunched posture was noted for all animals on Day 1 only.

Body weight:

Overall body weight gain in males and females was within the range expected for rats of this strain and age used in this type of study and were therefore considered not indicative of toxicity.

Gross pathology:

No abnormalities were found at macroscopic post mortem examination of the animals.

Applicant's summary and conclusion

Interpretation of results:

GHS criteria not met

Conclusions:

Based on this result, the inhalatory LC50, 4h value of 2,2’-Azobis (4-methoxy-2,4-dimethyl valeronitrile) in Wistar rats exceeds the technically maximum obtainable concentration of 3.2 mg/L. According to the OECD 436 test guideline, the LC50, 4h cut-off value was considered to exceed the technically maximum obtainable concentration of 3.2 mg/L.
Based on these results 2,2’-Azobis (4-methoxy-2,4-dimethyl valeronitrile) does not have to be classified and has no obligatory labelling requirement for acute inhalation toxicity according to the Globally Harmonized System of Classification and Labelling of Chemicals (GHS) of the United Nations (2015) (including all amendments) and Regulation (EC) No 1272/2008 on classification, labelling and packaging of substances and mixtures (including all amendments).

Executive summary:

Assessment of acute inhalation toxicity with 2,2’-Azobis (4-methoxy-2,4-dimethyl valeronitrile) in the rat (acute toxic class method) (nose-only) The study was carried out based on the guideline described in Organisation for Economic Co-operation and Development (OECD), OECD Guidelines for Testing of Chemicals, Section 4, Health Effects. No.436, "Acute Inhalation Toxicity - Acute Toxic Class Method", September 2009. The test atmosphere generation was based on the method developed during extensive trial generations. The generation was performed at the technically maximum attainable concentration. 2,2’-Azobis (4-methoxy-2,4-dimethyl valeronitrile) was administered as a dust by nose-only inhalation for 4 hours to one group of three male and three female Wistar rats. Mortality and clinical signs were observed daily during the observation period and body weights were determined on Days 1, 2, 4, 8 and 15. Macroscopic examination was performed after terminal sacrifice (day 15). The actual time-weighted mean concentration was 3.19 ± 0.08 mg/L. The concentration measurements equally distributed over time showed that it was difficult to maintain the concentration at a constant level and that the generation had to be interrupted at several occasions. The generation was initiated with test item that was stored two days in the freezer after passing through the sieve. During the first hour the concentration remained below 2.5 mg/L, which was considered too low to provide relevant results. It was therefore decided to replace the test item with test item that remained from the trial generations, which was stored at room temperature for 6 days (protected from light). After passing the test item through the sieve, the generation was restarted and a higher concentration level could be maintained without interruption. Overall, the generation was interrupted at two occasions and the generation time was elongated with 15 minutes in order to achieve an actual exposure time of 240 minutes. By calculating the time-weighted mean concentration, the effects of the interruptions and variation in concentration level were taken into account resulting in an actual reflection of the mean exposure concentration over time. Due to the technical difficulties, the nominal concentration (amount of test item used divided by the volume of pressurized air used) and the generation efficiency (ratio of actual and nominal concentration) could not be determined. Since these values are not so meaningful for dust generations, it was considered that the study integrity was not affected by this. The Mass Median Aerodynamic Diameter (MMAD) and geometric standard deviation (gsd) were determined twice during the exposure period. The MMAD was 5.8 μm (gsd 1.9) at both occasions. Multiple attempts were made to reduce the particle size during method development, either by lowering the concentration, using particle separators into the set-up or applying different grinding techniques, so the size obtained was considered the smallest that could be technically be achieved. No mortality occurred. During exposure, slow breathing was seen. After exposure, hunched posture was noted for all animals on Day 1 only. Overall body weight gain in males and females was within the range expected for rats of this strain and age used in this type of study. No abnormalities were found at macroscopic post mortem examination of the animals.

Based on this result, the inhalatory LC50, 4h value of 2,2’-Azobis (4-methoxy-2,4-dimethyl valeronitrile) in Wistar rats exceeds the technically maximum obtainable concentration of 3.2 mg/L. According to the OECD 436 test guideline, the LC50, 4h cut-off value was considered to exceed the technically maximum obtainable concentration of 3.2 mg/L. Based on these results 2,2’-Azobis (4-methoxy-2,4-dimethyl valeronitrile) does not have to be classified and has no obligatory labelling requirement for acute inhalation toxicity according to the Globally Harmonized System of Classification and Labelling of Chemicals (GHS) of the United Nations (2015) (including all amendments) and Regulation (EC) No 1272/2008 on classification, labelling and packaging of substances and mixtures (including all amendments).