Perhydro-1,3,5-trinitro-1,3,5-triazine

Biodegradation in water and sediment: simulation tests

Administrative data

Endpoint:

biodegradation in water: sewage treatment simulation testing

Type of information:

experimental study

Adequacy of study:

key study

Study period:

May 1981

Reliability:

1 (reliable without restriction)

Data source

Reference

Reference Type:

study report

Title:

Unnamed

Year:

1981

Materials and methods

Test guideline

Qualifier:

equivalent or similar to guideline

Guideline:

EU Method C.10 (Biodegradation: Activated Sludge Simulation Test)

GLP compliance:

not specified

Test material

Test material information

Constituent 1

Reference substance name:

Perhydro-1,3,5-trinitro-1,3,5-triazine

EC Number:

204-500-1

EC Name:

Perhydro-1,3,5-trinitro-1,3,5-triazine

Cas Number:

121-82-4

Molecular formula:

C3H6N6O6

IUPAC Name:

1,3,5-trinitro-1,3,5-triazinane

Study design

Oxygen conditions:

aerobic/anaerobic

Inoculum or test system:

activated sludge, adapted

Details on source and properties of surface water:

For aerobic studies the media were inoculated with activated sludge obtained from the Marlboro Easterly Municipal Sewage Treatment Plant, Marlboro, Mass. The volume of liquid did not exceed 10% of the total volume of the flask; the flasks were incubated at 30°C on a reciprocating shaker (150 strokes/min). For anaerobic studies the vessels were filled to approximately 95% of their capacity, inoculated with anaerobic sewage sludge (obtained from the Nut Island Sewage Treatment Plant, Boston, Mass.), and incubated as stationary cultures at 37°C. The inocula were prepared by diluting the sludge with two volumes of distilled water and filtering through glass wool. A 2% (vol/vol) inoculum was used.

Details on inoculum:

Due to the relative insolubilities of the compounds used as substrates, weighed amounts were added to empty flasks and dissolved in a small amount of acetone. The acetone was evaporated by a stream of N2, leaving a thin film of material on the inside surface of the flask. The culture medium was deoxygenated by boiling, poured into the flask containing the deposit of material, and stirred vigorously until solution was attained. The medium was allowed to cool to 35 to 40°C before inoculation. Samples were removed from the culture vessels after various periods of incubation and centrifuged, and the supernatant solutions were filtered through 0.22-,um membrane filters. The resulting culture medium filtrates (CMF) were used for all analytical procedures.

Parameter followed for biodegradation estimation:

test mat. analysis

Results and discussion

% Degradation

% Degr.:

ca. 100

Parameter:

test mat. analysis

Sampling time:

4 d

Remarks on result:

other: under anaerobic incubation

Transformation products:

yes

Identity of transformation products

Identity of transformation products 1

No.:

#1

Reference

Reference substance name:

Unnamed

IUPAC name:

Hexaydro-1-nitroso-3,5-dinitro-1,3,5-triazine (MNX)

Identity of transformation products 2

No.:

#2

Reference

Reference substance name:

Unnamed

IUPAC name:

hexahydro-1,3-dinitroso-5-nitro-1,3,5-triazine (DNX)

Identity of transformation products 3

No.:

#3

Reference

Reference substance name:

Unnamed

IUPAC name:

Hexahydro-1,3,5-trinitroso-1,3,5-triazine (TNX)

Identity of transformation products 4

No.:

#4

Reference

Reference substance name:

Unnamed

IUPAC name:

methanol

Inventory number:

InventoryMultipleMappingImpl [inventoryEntryValue=EC 200-659-6]

CAS number:

67-56-1

Identity:

Methanol

Molecular formula:

CH4O

Molecular weight:

32.042

SMILES notation:

CO

InChl:

InChI=1/CH4O/c1-2/h2H,1H3

Identity of transformation products 5

No.:

#5

Reference

Reference substance name:

Unnamed

IUPAC name:

hydrazine

Inventory number:

InventoryMultipleMappingImpl [inventoryEntryValue=EC 206-114-9]

CAS number:

302-01-2

Identity:

Hydrazine

Molecular formula:

H4N2

Molecular weight:

32.045

SMILES notation:

NN

InChl:

InChI=1/H4N2/c1-2/h1-2H2

Identity of transformation products 6

No.:

#6

Reference

Reference substance name:

Unnamed

IUPAC name:

formaldehyde

Inventory number:

InventoryMultipleMappingImpl [inventoryEntryValue=EC 200-001-8]

CAS number:

50-00-0

Identity:

Formaldehyde

Molecular formula:

CH2O

Molecular weight:

30.026

SMILES notation:

C=O

InChl:

InChI=1/CH2O/c1-2/h1H2

Details on transformation products:

The biodegradation ofRDX occurs only under anaerobic conditions. Concurrent with the disappearance of RDX is the sequential buildup and disappearance of the mono-, di-, and trinitroso analogs of RDX (Fig. 1). The fact that HCHO formation reaches a maximum in a short time (Fig. 2), whereas that of the nitroso derivatives lags behind, tends to rule out the possibility that HCHO is produced solely by reactions subsequent to the formation of the trinitroso derivative and suggests that formaldehyde is produced early in the reaction sequence from precursors of trinitroso-RDX.

Uninoculated controls containing RDX or TNX and incubated for the same periods of time as the inoculated flasks produced no HCHO. From the accumulated data we propose a pathway for the biodegradation of RDX as illustrated in Fig. 3.
In this scheme RDX is reduced sequentially to the nitroso derivatives, 2 (MNX), 3 (DNX), and 4 (TNX), each of which may undergo further reduction of a nitroso group to form the hypothetical compounds 5 (1-hydroxylamino-3,5-dinitro-1,3,5-triazine), 6 (1-hydroxylamino-3-nitroso-5-nitro-1,3,5-triazine), and 7 (1-hydroxylamino-3,5-dinitroso-1,3,5-triazine). We postulate that the molecule becomes unstable when any one of the nitro groups is reduced beyond the nitroso level. At this point hydrolytic cleavage, followed by rearrangement and further reductions of the fragments, gives rise to the end products observed. Cleavage of 5 via one route yields products 8 (N-hydroxymethyl-methylenedinitramine) and 9 (N-hydroxymethylenehydrazone), and cleavage of 6 via another route yields 10 (N-hydroxylamino-N'-nitromethylenediamine) and 11 (dimethylnitrosamine).
Compound 7 undergoes cleavage via either route. Figure 4 shows the postulated reactions of the fragments arising from the initial cleavage reaction. Cleavage of 8 releases 12 (HCHO) and 13 (methylenedinitramine), which decomposes to yield HCHO and 14 (nitramide), which in turn is reduced to 15 (hydrazine). Compound 9 rearranges and is reduced to 16 (hydroxymethylhydrazine), which yields HCHO and hydrazine. Under the strongly reducing conditions formaldehyde is reduced to methanol.

Applicant's summary and conclusion

Validity criteria

Validity criteria fulfilled:

yes

Conclusions:

RDX at concentrations of 50 or 100 µg/ml disappeared rapidly from nutrient broth cultures inoculated with anaerobic sewage sludge andmincubated anaerobically. RDX disappearance was essentially complete after 4 days. Concentrations of RDX remained unchanged when cultures were inoculated with aerobic activated sewage sludge and incubated aerobically. No RDX disappeared in uninoculated controls.

Executive summary:

Biodegradation of the explosive hexahydro-1,3,5-trinitro-1,3,5-triazine (RDX) occurs under anaerobic conditions, yielding a number of products, including: hexahydro- l-nitroso-3,5-dinitro- 1,3,5-triazine, hexahydro- 1,3-dinitroso-5-nitro- 1,3,5-triazine, hexahydro-1,3,5-trinitroso-1,3,5-triazine, hydrazine, 1,1-dimethylhydrazine, 1,2-dimethylhydrazine, formaldehyde, and methanol. A scheme for the biodegradation of RDX is proposed which proceeds via successive reduction of the nitro groups to a point where destabilization and fragmentation of the ring occurs. The noncycic degradation products arise via subsequent reduction and rearrangement reactions of the fragments. The scheme suggests the presence of several additional compounds, not yet identified. Several of the products are mutagenic or carcinogenic or both. Anaerobic treatment of RDX wastewaters, which also contain high nitrate levels, would permit the denitrification to occur, with concurrent degradation of RDX ultimnately to a mixture of hydrazines and methanol. The feasibility of using an aerobic mode in the further degradation of these products is discussed.