N-nitro-N-(3-methyl-3,6-dihydro-2H-1,3,5-oxadiazin-4-yl)amine

Acute Toxicity: inhalation

Administrative data

Endpoint:

acute toxicity: inhalation

Type of information:

experimental study

Adequacy of study:

key study

Study period:

December 1995 - March 1996

Reliability:

1 (reliable without restriction)

Rationale for reliability incl. deficiencies:

other: GLP guideline study; documentation sufficient for assessment

Data source

Reference

Reference Type:

study report

Title:

Unnamed

Year:

1996

Report date:

1996

Materials and methods

Test guideline

Test guideline 1

Qualifier:

according to guideline

Guideline:

OECD Guideline 403 (Acute Inhalation Toxicity)

Deviations:

yes

Remarks:

relative humidity oft the test atmosphere (12 %) below the stipulated range of 40 - 60 %; lowest temperature (19.8 °C) slightly below the stipulated rabge (22 +/- 2 °C);

Test guideline 2

Qualifier:

according to guideline

Guideline:

EU Method B.2 (Acute Toxicity (Inhalation))

Deviations:

yes

Remarks:

particle size distribution: measurement carried out with a 10-stage cascade impactor instead of a 11-stage cascade

Test guideline 3

Qualifier:

according to guideline

Guideline:

other: USEPA FIFRA 81-3

Deviations:

yes

Remarks:

see above

GLP compliance:

yes (incl. QA statement)

Remarks:

This study was performed in compliance with the OECD Principles of Good Laboratory Practice.

Test type:

standard acute method

Limit test:

yes

Test material

Test material information

Constituent 1

Reference substance name:

-

EC Number:

431-060-1

EC Name:

-

Cas Number:

153719-38-1

Molecular formula:

C4H8N4O3

IUPAC Name:

3-methyl-N-nitro-3,6-dihydro-2H-1,3,5-oxadiazin-4-amine

Details on test material:

- Name of test material (as cited in study report): CA 2343 A (Intermediate of CGA 293343); N-Nitro-5-methyl-tetrahydro-4H-1,3,5-oxadiazin-4-imin
- Physical state: fine white powder
- Analytical purity: 96.7 %
- Batch No.: P.503005
- Storage condition of test material: 4 °C
- melting point: 138 - 140 °C

Test animals

Species:

rat

Strain:

Wistar

Sex:

male/female

Details on test animals or test system and environmental conditions:

TEST ANIMALS
- Source: Charles River Wiga, Sulzfeld, Germany
- Age at study initiation: 5-6 weeks
- Mean body weight at study initiation: 292 g (males) nad 193 g (females)
- Housing during exposure: housed individually
- Housing after exposure: 5 males or 5 females to a cage
- Cages: stainless steel cages fitted with wire-mesh floor and front
- Diet: cereal-based rodent diet
- Water: tap water
- Acclimation period: 25 days
- Quarantine after arrival at test laboratory: 5 days
- Identification: earmark (tattoo)


ENVIRONMENTAL CONDITIONS
- Temperature (°C): 21.0 - 23.5
- Humidity (%): 45 - 70
- Air changes (per hr): 10
- Photoperiod (hrs dark / hrs light): 12 / 12

Administration / exposure

Route of administration:

inhalation

Type of inhalation exposure:

nose only

Remarks:

nose-only inhalation chamber; modification of the chmaber manufactured by ADG Developments Ltd, UK

Vehicle:

other: unchanged (no vehicle)

Details on inhalation exposure:

GENERATION OF TEST ATMOSPHERE / CHAMBER DESCRIPTION
- Exposure apparatus: nose-only inhalation chamber
- Exposure chamber volume: ca. 50 L
- Description of chamber: cylindrical aluminium column, surrounded by a transparent cylinder
- Dexscription of column: column consisted of a top assembly with two mixing chambers, underneath a rodent tube section and the exhaust section at the bottom
- rodent tube section: 20 ports for animal exposure
- empty ports were used for test atmosphere sampling, particle size analysis, measurements of oxygen concentration, temperature and relative humidity
- Method of holding animals in test chamber: animals were secured in plastic animal holders (Battelle), positioned radially through the outer cylinder around the central column (animals were placed in altering order, male, female, etc.; remaining ports were closed); only the nose of the rats protruded into the interior of the column
- Source and rate of air: test atmosphere was generated by passing the ground test material using a dry material helix feeder to a jet mill, which was operated with dry pressurized air (less than 1 % humidity)
- System of generating particulates/aerosols: delivery of test material: slip stream of airconditioned room air and pressurized air (stream: 33 % of total air); generated aerosol was passed to the inlet of the exposure unit and directed downward through the mixing chambers towards the animals
- Method of particle size determination: 10-stage Andersen cascade impactor with the largest cut-off size of 32 µm; last stage (viz. with cut-off size 0.1 µm) was not used
- Temperature, humidity, pressure in air chamber: T = 20.6 +/- 0.4 °C; rel. humidity = 12 +/- 1 % ; c(O2) = 21.9 %; positive pressure


TEST ATMOSPHERE
Brief description of analytical method used:
- continuous supply of fresh test atmosphere
- the test atmosphere was exhauted at the bottom of the unit
- animal exposure: 30 minutes after start of the generation of the test atmosphere
- mean amount of air passed through the exposure unit: 105.8 l/min

Samples taken from breathing zone: yes




TEST ATMOSPHERE (if not tabulated)
- actual concentration was determined by gravimetric analysis (twice each hour)
- Particle size distribution: 67 % of the particles present in the breathing zone are smaller or equal 8.2 µm
- MMAD (Mass median aerodynamic diameter) / GSD (Geometric st. dev.): 6.0 µm / 2. 5 µm

Analytical verification of test atmosphere concentrations:

yes

Remarks:

determination of actual (twice each hour) and nominal concentration

Duration of exposure:

4 h

Concentrations:

nominal: 13.1 g/m³;
actual: 6.92 +/- 0.50 g/m³

No. of animals per sex per dose:

5

Control animals:

no

Details on study design:

- Duration of observation period following administration: 14 days
- Frequency of observations (behaviour, clinical signs, mortality): just before exposure, during and shortly after the exposure, and at least once during the observation period
- Frequency of weighing: just prior to exposure (day 0), and on days 7 and 14.
- Necropsy of survivors performed: yes (on day 14, exsanguination from the abdominal aorta under ether anaesthesia)

Statistics:

Calculation of the Mass Median Aerodynamic Diameter (MMAD) and the mean geometric standard deviation (gsd).

Results and discussion

Effect levels

Sex:

male/female

Dose descriptor:

LC50

Effect level:

> 6.92 other: g/m³

Exp. duration:

4 h

Mortality:

- none of the animals died during the exposure or during the observation period

Clinical signs:

other: During exposure: - shallow breathing and visually decdreased breathing rate in all animals - second, third and fourth exposure hour: slight shollw breathing - slight visually decreased breathing during the last two hours of exposure Signs shortly after e

Body weight:

The overall body weight gain was considered to be normal for male rats and slightly reduced for female rats of this strain and age.

Gross pathology:

Abnormalities:
- grey discoloured lungs in one male and two female rats
- small lung haemorrhages in two males which were not considered to be treatment related

Other findings:

none

Any other information on results incl. tables

Aerodynamic particle size distribution in test atmosphere

Particle size (µm)	Mass (mg)	Cumulative (%)
≤ 32.0	0.08	97.9
≤ 20.5	0.14	94.6
≤ 8.2	1.18	67.1
≤ 5.0	1.34	35.7
≤ 3.1	0.57	22.4
≤ 1.9	0.52	10.3
≤ 1.1	0.27	4.0
≤ 0.7	0.12	1.2
≤ 0.4 *	0.05	0.0
≤ 0.1 **	--	--
total	4.28

 * The total amount of the test material sampled included back up filter last stage

** This stage was not used

 The gas volume used for cascade sample was 2.3 L sample flow was 2.3 L/min;

 Mass Median Aerodynamic Diameter (MMAD): 6.0 µm

 Mean geometric standard deviation (gsd): 2.5 µm

Applicant's summary and conclusion

Interpretation of results:

not classified

Remarks:

Migrated information LC50 is higher than 6.92 g/m³ Criteria used for interpretation of results: not specified

Conclusions:

The aim of this study was to determine the 4 -hour LC50 value of CA 2343 A (Intermediate of CGA 293343). Hereto one group of 5 male and 5 female rats was exposed to a limit concentration of 6.92 +/- 0.50 g/m³ CA 2343 A (Intermediate of CGA 293343) during a single period of four hours. Since none of the rats died at this concentration level, the 4 -hour LC50 value of CA 2343 A (Intermediate of CGA 293343) was higher than 6.92 g/m³

Executive summary:

The aim of this study was to determine the 4 -hour LC₅₀ value of CA 2343 A (Intermediate of CGA 293343). Each five male and female rats were exposed to the test atmosphere generated by passing the ground test article using a dry material helix feeder to a jet mill for four hours (nose-only exposure) at an actual concentration of 6.92 +/- 0.50 g/m³.

Clinical signs were recorded during exposure and daily thereafter for 14 days. Body weights were measured prior to exposure and on days 7 and 14. At termination of the study, all surviving animals were subjected to gross necropsy.

67 % of the particles present at the animals breathing zone had an aerodynamic diameter of ≤ 8.2 µm. The mass median aerodynamic diameter (MMAD) was 6.0 µm

None of the test animals died. Shallow breathing and decreased breathing rates were observed during the exposure period and shortly thereafter. Lethargy, piloerection, dirty heads, wet fur in the abdominal region, ovally shaped eyes, encrustations around eyes or nose and salivation were further clinical signs observed soon after exposure, some of them still present on day 3. Overall body weight gain was normal for males and slightly reduced for females. At necropsy, grey discoloured lungs were observed in one male and two females.

The acute inhalation (4 hour) LC₅₀ for CA 2343 A (Intermediate of CGA 293343) in the rat is higher than 6920 mg/m³.