Octahydro-1,3,5,7-tetranitro-1,3,5,7-tetrazocine

Endpoint summary

Description of key information

The LD50 of acute oral toxicity in rats is 6250 mg/kg bw but LD50 in male mice = 1670 mg/kg and 3240 mg/kg for female mice. LD50 for dermal toxicity in rats is >4230 mg/kg bw but LD50 is  634.12 mg/kg for male rabbits and 718.56 mg/kg for female rabbits . No inhalation data is available as the study was waived as being scientifically unjustified due to the large particle sizes of the test substance (>77% is a non-respirable fraction according to the granulometry data).

Key value for chemical safety assessment

Acute toxicity: via oral route

Link to relevant study records

Reference

Endpoint:

acute toxicity: oral

Type of information:

experimental study

Adequacy of study:

key study

Study period:

From 10 to 31 July 1980

Reliability:

2 (reliable with restrictions)

Rationale for reliability incl. deficiencies:

other: Study run to a reliable method but not GLP and no guideline followed.

Qualifier:

no guideline followed

Principles of method if other than guideline:

Five dose levels were selected in which 5 groups of 5 males and 5 females mice were dosed once. An additional group of 5 male and 5 female mice was dosed with vehicle only. The mice were observed in the morning and afternoon for 14 days following administration of the test substance. At death, or at the end of the observation period and at termination, each animal was subjected to a gross post mortem examination.

GLP compliance:

not specified

Test type:

standard acute method

Limit test:

no

Species:

mouse

Strain:

B6C3F1

Sex:

male/female

Details on test animals or test system and environmental conditions:

TEST ANIMALS
- Source: Charles River UK Limited
- Weight at study initiation: The mean body weight on being dosed was 22 g (range 20-22 g) in the dose ranging study for males and 21 g (range 19-22 g) in the dose ranging study for females. The mean body weight in the main study was 22 g (range 18-24 g) for males and 18.5 g (range 16-21 g) for females.
- Fasting period before study: The animals were deprived of food for a 4 h period prior to dosing.
- Housing: They were housed individually in polypropylene cages with stainless steel grid tops and sterilised wood shavings.
- Diet (e.g. ad libitum): The animals were fed on BP Nutrition expanded Mouse Maintenance Diet No. 1.
- Water (e.g. ad libitum): Water was available ad libitum throughout the study

ENVIRONMENTAL CONDITIONS
- Temperature (°C): 22°C (extremes of 19°C-25°C)
- Humidity (%): 57% (extremes of 52%-74%)

IN-LIFE DATES: From 10 to 31 July 1980

No additional data

Route of administration:

oral: gavage

Vehicle:

CMC (carboxymethyl cellulose)

Remarks:

0.5% low viscosity CMC

Details on oral exposure:

VEHICLE
- Amount of vehicle (if gavage): 20 mL/kg

MAXIMUM DOSE VOLUME APPLIED: 20 mL/kg

No additional data

Doses:

Dose ranging study:
300, 700, 1500, 5000 and 15000 mg wet test substance/kg
247, 577, 1236, 4120 and 12360 mg dry test substance/kg

Main study:
1200, 2040, 3468, 5895.6 and 10022.5 mg wet test substance/kg
956, 1626, 2764, 4699 and 7988 mg dry test substance/kg

No. of animals per sex per dose:

Dose ranging study: one male and one female
Main study: 5 males and 5 females

Control animals:

yes

Details on study design:

- Duration of observation period following administration: 14 days
- Frequency of observations and weighing: The mice were observed on the morning and afternoon for 14 days following administration of the test substance.
- Necropsy of survivors performed: yes, At death, or at the end of the observation period and sacrifice, each animal was subjected to a gross post mortem examination.
- Other examinations performed: clinical signs, gross pathology

Statistics:

The LD50 was calculated using a method based on the following:
Finney (1971) "Probit Analysis", Cambridge University Press.

Preliminary study:

In the dose ranging study in pairs of mice, mortality was 0/2, 0/2, 1/2, 1/2 and 1/2 in the dose group 247, 577, 1236, 4120 and 12360 mg dry test substance/day dose groups, respectively.
Clinical signs included piloerection, hypokinesia and ataxia, lasting for up to 3 days after dosing.
Post mortem observations included white fluid in stomach and upper gastro-intestinal tract with kidneys pale and mottled.

Sex:

male

Dose descriptor:

LD50

Effect level:

1 670 mg/kg bw

Based on:

test mat.

95% CL:

>= 1 440 - <= 1 890

Remarks on result:

other: Based on dry test substance

Sex:

female

Dose descriptor:

LD50

Effect level:

3 240 mg/kg bw

Based on:

test mat.

95% CL:

>= 2 920 - <= 3 570

Remarks on result:

other: Based on dry test substance

Sex:

male/female

Dose descriptor:

LD50

Effect level:

2 300 mg/kg bw

Based on:

test mat.

95% CL:

>= 2 110 - <= 2 500

Remarks on result:

other: Based on dry test substance

Mortality:

In the main study, mortality was 0/10, 4/10, 5/10, 10/10 and 10/10 in the 956, 1626, 2764, 4699 and 7988 mg dry test substance/kg dose groups, respectively.

Clinical signs:

other: Clinical signs included piloerection, soiled coat, hyperkinesia, hypokinesia, ataxia, sedation, eyes half shut and penis protruded. In one animal the penis was very swollen and stained with blood with a constant stream of urine.

Gross pathology:

Post mortem observations included stomach and gastro-intestinal tract filled with white fluid, gut contents fluid, blood filled gut, stomach wall white, penis extended and dark red and lungs red.

Other findings:

No information provided

There were no deaths or clinical signs recorded in the control group.

No abnormalities were detected at post mortem observations in the control group.

Interpretation of results:

harmful

Remarks:

Migrated information Criteria used for interpretation of results: EU

Conclusions:

The Oral Median Lethal Doses (LD50s) of dry test substance were calculated to be 1960 mg/kg for male, 3240 mg/kg for female and 2300 mg/kg for male and female B6C3F1 mice.

Endpoint conclusion

Endpoint conclusion:

adverse effect observed

Dose descriptor:

LD50

Value:

1 670 mg/kg bw

Acute toxicity: via inhalation route

Endpoint conclusion

Endpoint conclusion:

no study available

Acute toxicity: via dermal route

Link to relevant study records

Reference

Endpoint:

acute toxicity: dermal

Type of information:

experimental study

Adequacy of study:

key study

Study period:

From 29 July to 29 October 1980

Reliability:

2 (reliable with restrictions)

Rationale for reliability incl. deficiencies:

other: Study run to a reliable method but no GLP and no guideline followed

Qualifier:

no guideline followed

Principles of method if other than guideline:

In the main study, 76 rabbits were randomly allocated to 8 test groups, each of 4 male and 4 female rabbits and 2 control groups, each of 3 male and 3 female rabbits. Four test groups and one control group (vehicle only) were abraded at the treatment site, the remaining 4 groups were non-abraded.

GLP compliance:

not specified

Limit test:

no

Species:

rabbit

Strain:

New Zealand White

Sex:

male/female

Details on test animals or test system and environmental conditions:

TEST ANIMALS
- Source: Cheshire Rabbits Farms
- Weight at study initiation: The mean body weight on being dosed for Phases 1, 2 and 3 was 3.03 kg (range 2.56-3.66 kg) for males and 3.11 kg (range 2.47-3.84 kg) for females. The mean body weight in the main study was 3.13 kg (range 2.35-3.63 kg) for males and 3.08 kg (range 2.45-3.73 kg) for females.
- Housing: They were housed individually in cages with a grid floor, beneath which was a peat moss filled tray.
- Diet (e.g. ad libitum): They were fed Spratt's Rabbit Diet ad libitum throughout the study.
- Water (e.g. ad libitum): Water were available ad libitum throughout the study.

ENVIRONMENTAL CONDITIONS
- Temperature (°C): 18.5°C (extremes of 15°C-23°C) for the dose ranging studies and 17°C (extremes of 14°C-20°C) for the main study.
- Humidity (%): 68% (extremes of 58%-78%) for the dose ranging studies and 64% (extremes of 53%-76%) for the main study.

IN-LIFE DATES: From 29 July to 29 October 1980

No additional data

Type of coverage:

occlusive

Vehicle:

other: 1% high viscosity CMC for Phases 2 and 3, Physiological saline for Phases 1 and 2.

Details on dermal exposure:

TEST SITE
- Area of exposure: The entire trunk of the rabbit between the fore and hind limbs.
- % coverage: Approximately 10% of the body surface.
- Type of wrap if used: The gauze was covered with an impervious covering of Sleek occlusive tape.

REMOVAL OF TEST SUBSTANCE
- Washing (if done): The wrapping was removed and the skin wiped to remove any remaining test material.
- Time after start of exposure: 24 hours

TEST MATERIAL
- Concentration (if solution): 600 mg/mL
- Constant volume or concentration used: yes

No additional data

Duration of exposure:

24 hours

Doses:

Phase 1: 600, 1800 and 3000 mg dry test substance/kg/day bw (in physiological saline)

Phase 2:
600, 1800 and 3000 mg dry test substance/kg/day bw (in physiological saline)
300, 600, 1800 and 3000 mg/kg bw (in 1% CMC)

Phase 3:
Dose ranging study: 600, 1200 and 2400 mg/kg (in CMC)
Main study: 168, 372, 816 and 1788 mg/kg (in 1% CMC)

No. of animals per sex per dose:

Phase 1: one male and one female
Phase 2: one male and one female for test substance in physiological saline, one male and one female for test substance in 1% CMC
Phase 3: one male and one female in Dose ranging study, 4 male and 4 female in Main study, 3 male and 3 female for control group

Control animals:

yes, concurrent vehicle

Details on study design:

- Duration of observation period following administration: 14 days
- Frequency of observations and weighing: Animals were weighed on the day of administration and weekly thereafter, until death or sacrifice. They were observed frequently on the day of administration and in the morning and afternoon for 14 days following administration of the test substance, when surviving animals were sacrificed.
- Necropsy of survivors performed: yes
- Other examinations performed: Histopathology: At death or sacrifice in Phase 2 and 3, selected tissues were taken for histopathological examination. The tissues taken were the liver, kidneys, spleen, heart, lungs, brain, spinal cord and 2 pieces of skin from the dorsal area and these were fixed in 10% neutral buffered formalin.

Statistics:

None stated

Preliminary study:

Phase 1:
Mortality was 1/2, 2/2 and 2/2, respectively. Deaths occurred between 3 and 6 days after dosing.
Clinical signs included hyperkinesia, blood around nose, clonic convulsions, laboured breathing, vocalisation and loss of co-ordination.
At post mortem examination, the following observations were recorded; blood in nasal cavity, subdural haemorrhage between cerebellum and cerebrum, slight pitting of surface of kidneys and pale kidneys.

Phase 2:
Treatment with test substance in physiological saline resulted in 0/2, 2/2 and 2/2 deaths, respectively. Deaths occurred between 1 and 6 days after dosing.
Major clinical signs observed were clonic convulsions, hyperkinesia, miosis, laboured respiration, mydriasis, dyspnoea, cyanosis and hypokinesia.
Observations noted at the post mortem examination included red staining around nose, blood around mouth creamy particulate matter in thoracic cavity and pericardial sac fluid filled.
Treatment with test substance in high viscosity 1% CMC resulted in mortality of 1/2, 1/2, 2/2 and 2/2, respectively.
Clinical signs noted were hyperkinesia, miosis, clonic convulsions, blood around nose, vocalisation, loss of mobility and laboured breathing.
Observations noted at post mortem examination included red staining around mouth and nose, subdural haemorrhage and small blood clot on the brain.

Phase 3 (Dose ranging study):
Mortality was 1/2, 2/2 and 2/2, respectively. Deaths occurred 2 to 4 days after dosing.
Clinical signs noted were hyperkinesia, clonic convulsions, vocalisation and animal unable to move hind legs.
Observations noted at post mortem examination included extensive bruising of hind legs, broken femur, clotted blood in cervical region of spinal cord, clotted blood between cerebellum and cerebrum and dark red lungs.

Sex:

male

Dose descriptor:

LD50

Effect level:

634.12 mg/kg bw

Based on:

test mat.

95% CL:

>= 532.49 - <= 735.75

Remarks on result:

other: Non-abraded skin

Sex:

male

Dose descriptor:

LD50

Effect level:

673.81 mg/kg bw

Based on:

test mat.

95% CL:

>= 562.34 - <= 785.28

Remarks on result:

other: Abraded skin

Sex:

female

Dose descriptor:

LD50

Effect level:

718.56 mg/kg bw

Based on:

test mat.

95% CL:

>= 595.59 - <= 841.53

Remarks on result:

other: Non-adraded skin

Sex:

female

Dose descriptor:

LD50

Effect level:

1 336.7 mg/kg bw

Based on:

test mat.

95% CL:

>= 414.56 - <= 1 758.84

Remarks on result:

other: Abraded skin

Sex:

male/female

Dose descriptor:

LD50

Effect level:

982.03 mg/kg bw

Based on:

test mat.

95% CL:

>= 861.46 - <= 1 102.6

Remarks on result:

other: Non-abraded/abraded skin

Mortality:

Mortality in the non-abraded groups was 0/8, 0/8, 4/8 and 8/8 for each dose group, respectively. Mortality in the abraded groups was 0/8, 0/8, 3/8 and 7/8 for each dose group, respectively. One female non-abraded control animal died 10 days after dosing with vehicle only.

Clinical signs:

other: Clinical signs included hyperkinesia, mydriasis, hypokinesia, clonic convulsion, miosis, aggressive, timid, left hind limb atiff and unable to move, vocalisation, prostration in non-abraded and abraded groups, also noted blood around nose and mouth, diffi

Gross pathology:

There was no evidence of broken back in non-abraded and abraded groups.

Other findings:

Histopathological symptoms included congestion, centrilobular necrosis/degeneration, increased cellularity, eosinophilic cytoplasm in liver; congestion and chronic nephropathy in kidney; congestion, alveolar haemorrhage, alveolar exudation and alveolar degeneration in lung; white pulp depletion and red pulp depletion in spleen.

Interpretation of results:

toxic

Remarks:

Migrated information Criteria used for interpretation of results: EU

Conclusions:

The dermal Median Lethal Doses (LD50) for HMX were calculated as 634.12 and 718.56 mg/kg for male and female New Zealand White rabbits on non abraded skin, respectively. The dermal LD50 was 982.03 mg/kg for male and female New Zealand White rabbits on non-abraded/abraded skin.

Endpoint conclusion

Endpoint conclusion:

adverse effect observed

Dose descriptor:

LD50

Value:

634 mg/kg bw

Additional information

Oral Route

The key study for acute oral toxicity was performed by oral gavage in mice using an HMX suspension in 0.5% carboxymethylcellulose (Inveresk Research International, on behalf of the U.S. Army Medical Research and Development Command, 1985). Five groups of 5 male and 5 female mice were dosed once at 1200, 2040, 3468, 5895.6 and 10022.5 mg wet HMX/kg (equivalent to 956, 1626, 2764, 4699 and 7988 mg dry HMX/kg). Clinical signs included piloerection, soiled coat, hyperkinesia, hypokinesia, ataxia, sedation, eyes half shut and penis protruded. On animal had a very swollen penis stained with blood with a constant stream of urine. Post mortem observations included stomach and gastro-intestinal tract filled with white fluid, gut contents fluid, blood filled gut, stomach wall white, penis extended and dark red and lungs red. The LD50 for male mice was 1670 mg/kg/day and for female mice was 3240 mg/kg/day.

A supporting study for acute oral toxicity in rats was performed by oral gavage using an HMX suspension in 0.5% carboxymethylcellulose (Inveresk Research International, on behalf of the U.S Army Medical Research and Development Command, 1985). The LD50 for male and female rats was calculated to be 6250 mg/kg bw (5510 mg/kg bw for males and 6440 mg/kg bw for females).

Inhalation Route

Study was waived as being scientifically unjustified due to the particle size of HMX consisting of >77% non-respirable fraction.

Dermal Route

The key study for acute dermal toxicity was performed in rabbits using an HMX solution in 1% carboxymethylcellulose (Inveresk Research International, on behalf of the U.S. Army Medical Research and Development Command, 1985). After inital dose ranging studies, HMX in 1% CMC at a constant concentration of 600 mg/mL and at varying dose volumes was administered dermally under occlusion. The dose levels selected were 1268, 372, 816 and 1788 mg/kg. Groups of 4 male and 4 female rabbits were dosed at each level. The application site was the entire trunk of the rabbit between the fore and hind limbs. This was shaved and the skin abraded (where applicable) in such a way as to penetrate the stratum corneum but not the dermis by making abrasions every 2 -3 cm longitudinally over the entire area of exposure, by means of a 'sterilin' blood lancet. The test material was applied to the prepared skin on a piece of gauze of a length to cover approximately 10% of the body surface. The gauze was coevered with an impervious covering of occlusive tape. Test substance was kept in contact with skin for 24 hr. At the end of the exposure period, the wrapping was removed and the skin wiped to remove remaining test material. The percutaneous LD50 in male rabbits was 634.12 mg/kg and in female rabbits was 718.56 mg/kg.

The supporting study in rats was performed using an HMX solution in physiological saline (Inveresk Research International, on behalf of the US Army Medical Research and Development Command, 1985). In the study no deaths occurred. Therefore the percutaneous LD50 in rats of HMX in physiological saline was >4230 mg dry HMX/kg.

Justification for selection of acute toxicity – oral endpoint

Study run to a scientific method similar to modern international guidelines though not to  GLP.

Justification for selection of acute toxicity – dermal endpoint

Study run to a scientific method similar to modern international guidelines though not to  GLP.

Justification for classification or non-classification

HMX is classified as Category 4 for acute oral toxicity and Category 3 for acute dermal toxicity.