A mixture of: triphenylthiophosphate and tertiary butylated phenyl derivatives

Biodegradation in water and sediment: simulation tests

Administrative data

Endpoint:

biodegradation in water: simulation testing on ultimate degradation in surface water

Type of information:

other: Study with main component of the substance

Adequacy of study:

key study

Reliability:

1 (reliable without restriction)

Rationale for reliability incl. deficiencies:

guideline study

Data source

Reference

Reference Type:

study report

Title:

Unnamed

Year:

2021

Report date:

2021

Materials and methods

Test guideline

Qualifier:

according to guideline

Guideline:

OECD Guideline 309 (Aerobic Mineralisation in Surface Water - Simulation Biodegradation Test)

Version / remarks:

adopted in April 2004

Deviations:

no

GLP compliance:

yes (incl. QA statement)

Test material

Test material information

Constituent 1

Reference substance name:

O,O,O-triphenyl phosphorothioate

EC Number:

209-909-9

EC Name:

O,O,O-triphenyl phosphorothioate

Cas Number:

597-82-0

Molecular formula:

C18H15O3PS

IUPAC Name:

O,O,O-triphenyl thiophosphate

Specific details on test material used for the study:

SOURCE OF TEST MATERIAL
- Source (i.e. manufacturer or supplier) and lot/batch number of test material: BASF SE, 1302-1004
- Purity, including information on contaminants, isomers, etc.: 100%

RADIOLABELLING INFORMATION
- Radiochemical purity: 97%
- Specific activity: 19.4 MBq/mg
- Locations of the label: phenyl ring ([phenyl-U-C14](O,O,O-tris-phenyl)phosphorothionate)
- Expiration date of radiochemical substance: 21 Feb 2021

STABILITY AND STORAGE CONDITIONS OF TEST MATERIAL
- Storage condition of test material: deep frozen (≤ -18 °C), dark, dry

Radiolabelling:

yes

Study design

Oxygen conditions:

aerobic

Inoculum or test system:

natural water / sediment: freshwater

Details on source and properties of surface water:

Water was sampled from a natural aerobic surface water source in Rhineland-Palatinate (67374 Hanhofen, Germany, 49°18'43.9"N, 8°19'10.3"E). Water was sampled from the top 5 cm to 10 cm of each compartment of the natural resources. The sampling was performed on 10 Aug 2020. The water/sediment system was transported in polyethylene containers to the laboratory. Since the transport duration did not exceed 2 to 3 hours, the water samples didn’t have to be cooled. Prior to use, coarse particles were removed from the water by filtration through a filter with 100 µm mesh size. The test water was directly used in the test and did not have to be stored.
Water properties: TOC: 11 mg/L; Total nitrogen: 1.6 mg/L; Total ammonium: 0.33 mg/L; Total phosphorus: < 0.02 mg/L; BOD = 3.8 mg/L O2; pH = 8.26

Details on source and properties of sediment:

Sediment was sampled from a natural aerobic surface water source in Rhineland-Palatinate (67374 Hanhofen, Germany, 49°18'43.9"N, 8°19'10.3"E). Sediment was sampled from the top 5 cm to 10 cm of each compartment of the natural resources. The sampling was performed on 10 Aug 2020. The water/sediment system was transported in polyethylene containers to the laboratory. Since the transport duration did not exceed 2 to 3 hours, the water samples didn’t have to be cooled. The sediment was sieved through a 2.0 mm sieve.
Sediment properties: loamy sand (clay: 2.5%, silt: 20%, sand: 77.5%), TOC: 0.35%, CEC: 2.9 mmol/100g, density: 1443 g/L

Duration of test (contact time):

61 d

Initial test substance concentration

Initial test substance concentration 1

Initial conc.:

10 µg/L

Based on:

test mat.

Initial test substance concentration 2

Initial conc.:

2 µg/L

Based on:

test mat.

Parameter followed for biodegradation estimation:

CO2 evolution

radiochem. meas.

test mat. analysis

Details on study design:

TEST CONDITIONS
- Volume of test solution/treatment: 500 mL
- Composition of medium: surface water
- Additional substrate: between 5 mg and 10 mg dispersed sediment
- Solubilising agent (type and concentration if used): acetonitrile (The concentration of the organic solvent was below 1% of the amount of test water present in each test vessel.)
- Test temperature: 12.1 °C with a range from 11.3 – 12.9 °C
- pH: 8.1
- pH adjusted: no
- Suspended solids concentration: 11 mg DOC/L
- Continuous darkness: yes
- Any indication of the test material adsorbing to the walls of the test apparatus: no

TEST SYSTEM
- Culturing apparatus: 1000 mL all-glass metabolism flasks (inner diameter: * 10.1 cm; surface: * 80 cm²)
- Number of culture flasks/concentration: 26 flasks, treated with 2 µg/L [14C]TPPT. 16 were analysed (10 flasks as reserve); 26 flasks, treated with 10 µg/L [14C]TPPT. 16 were analysed (10 flasks as reserve); 6 sterile samples, treated with 10 µg/L [14C]TPPT. 2 were analysed (2 flasks as reserve); 8 reference samples (autoclaved test system), treated with 5 µg/L [14C] sodium benzoate and additional the highest amount of solvent, 4 were analysed (4 flasks as reserve); 2 blank controls, treated with the highest amount of solvent
- Method used to create aerobic conditions: The aeration was performed using slightly constant orbital movement and constant air flow
- Test performed in open system: yes
- Details of trap for CO2 and volatile organics if used: The radioactive carbon dioxide evolved in the test flasks was trapped by two series-connected flasks containing sodium hydroxide (60 mL each; 2 M)

SAMPLING
Duplicate samples of each system were worked up at the following sampling time points: 0, 2, 7, 14, 21, 29, 44 and 61 days after treatment (DAT). The flasks for the reference item were sampled on day 0 and 14.
At every sampling time point both flasks were worked up immediately. The samples were analysed on the same day as sampling or stored at ≤-18°C for maximal 36 days prior to analysis.
After analysis, samples were stored in a freezer at ≤-18°.

Reference substance

Reference substance:

benzoic acid, sodium salt

Remarks:

radiolabeled (Benzoic acid [ring-14C(U)]sodium salt)

Results and discussion

Mean total recovery

Mean total recovery 1

Compartment:

natural water: freshwater

Sampling date:

2020

% Total extractable:

87.1

% Non extractable:

4.5

% CO2:

2

% Other volatiles:

< 0.1

% Recovery:

93.7

Remarks on result:

other: 10 µg/L treatment, day 61

Mean total recovery 2

Compartment:

natural water: freshwater

Sampling date:

2020

% Total extractable:

90.5

% Non extractable:

5.6

% CO2:

0

% Other volatiles:

0

% Recovery:

96.1

Remarks on result:

other: 10 µg/L treatment, day 0

Mean total recovery 3

Compartment:

natural water: freshwater

Sampling date:

2020

% Total extractable:

81.1

% Non extractable:

12.6

% CO2:

3.1

% Other volatiles:

<= 0.1

% Recovery:

96.9

Remarks on result:

other: 2 µg/L treatment, day 61

Mean total recovery 4

Compartment:

natural water: freshwater

Sampling date:

2020

% Total extractable:

90.5

% Non extractable:

5.6

% CO2:

0

% Other volatiles:

0

% Recovery:

96.1

Remarks on result:

other: 2 µg/L treatment, day 0

% Degradation

Parent/product:

parent

Compartment:

water

Key result

% Degr.:

>= 3.1 - <= 3.7

Parameter:

CO2 evolution

Sampling date:

2020

Sampling time:

61 d

Remarks on result:

other: 2 and 10 µg/L treatment

Half-life of parent compound / 50% disappearance time (DT50)

Key result

Compartment:

natural water: freshwater

DT50:

> 61 d

Type:

(pseudo-)first order (= half-life)

Temp.:

12 °C

Remarks on result:

other: 2 µg/L and 10 µg/L treatment

Transformation products:

yes

Identity of transformation products

Identity of transformation products 1

No.:

#4

Identity of transformation products 2

No.:

#3

Reference

Reference substance name:

Unnamed

IUPAC name:

phenyl dihydrogen phosphate

Inventory number:

InventoryMultipleMappingImpl [inventoryEntryValue=EC 211-857-7]

CAS number:

701-64-4

Molecular formula:

C6H7O4P

Molecular weight:

174.091

SMILES notation:

OP(=O)(O)Oc1ccccc1

InChl:

InChI=1/C6H7O4P/c7-11(8,9)10-6-4-2-1-3-5-6/h1-5H,(H2,7,8,9)

Identity of transformation products 3

No.:

#2

Reference

Reference substance name:

Unnamed

IUPAC name:

diphenyl hydrogen phosphate

Inventory number:

InventoryMultipleMappingImpl [inventoryEntryValue=EC 212-657-2]

CAS number:

838-85-7

Molecular formula:

C12H11O4P

Molecular weight:

250.187

SMILES notation:

OP(=O)(Oc1ccccc1)Oc2ccccc2

InChl:

InChI=1/C12H11O4P/c13-17(14,15-11-7-3-1-4-8-11)16-12-9-5-2-6-10-12/h1-10H,(H,13,14)

Identity of transformation products 4

No.:

#1

Reference

Reference substance name:

Unnamed

IUPAC name:

triphenyl phosphate

Inventory number:

InventoryMultipleMappingImpl [inventoryEntryValue=EC 204-112-2]

CAS number:

115-86-6

Molecular formula:

C18H15O4P

Molecular weight:

326.283

SMILES notation:

O=P(Oc1ccccc1)(Oc2ccccc2)Oc3ccccc3

InChl:

InChI=1/C18H15O4P/c19-23(20-16-10-4-1-5-11-16,21-17-12-6-2-7-13-17)22-18-14-8-3-9-15-18/h1-15H

Details on transformation products:

- Formation and decline of each transformation product during test: see documents attached
- Pathways for transformation: cleavage of phenol (which is considered to be mineralized to CO2 since phenol was not detected)
- Maximum occurrence of each transformation product: see documents attached
- Other: Two minor metabolites could not be identified (M2, M6, see documents attached)

Evaporation of parent compound:

no

Volatile metabolites:

no

Residues:

no

Details on results:

TEST CONDITIONS
- Aerobicity (or anaerobicity), moisture, temperature and other experimental conditions maintained throughout the study: Yes
- Anomalies or problems encountered (if yes): no

MINOR TRANSFORMATION PRODUCTS
- Range of maximum concentrations in % of the applied amount and day(s) of incubation when observed: see documents attached
- Range of maximum concentrations in % of the applied amount at end of study period: see documents attached

MINERALISATION
- % of applied radioactivity present as CO2 at end of study: 3.1 - 3.7%

VOLATILIZATION
- % of the applied radioactivity present as volatile organics at end of study: 0 - 0.1 %

STERILE TREATMENTS (if used)
The sterile flasks were sampled and analysed after 61 days of incubation. The recovery of radioactivity was 93.8 % AR. 14CO2 was detected in amounts of 0.8 % AR. No organic volatiles were detected (≤0.1 % AR). The sterile sample showed a similar degradation pattern than the biologically active test flasks. The parent substance was found in the total system at an amount of 88.1 % AR. Four metabolites/impurities were observed (M1, M3, M4 and M5) with maximum amounts of 2.7 % AR.

Results with reference substance:

After 14 days, 83.4 % AR of the reference item was mineralised. Thus, the test system was biologically active and the test was valid.

Any other information on results incl. tables

Sample Concentration prior to Analysis

Prior to analysis by HPLC, all samples were concentrated as described in chapter 2.9.4. Procedural recoveries for concentrating the acetonitrile extract lay always between 90 and 110 %. For concentration of the extracted water phase recoveries were below 90 %. Since the remaining radioactivity in the water phase is very small (below 15 % AR, for almost all samples below 6 %AR), there was no significant loss referred to the total amount of radioactivity applied.

Applicant's summary and conclusion

Validity criteria

Validity criteria 1

Validity criteria:

The total recovery (mass balance) at the end of the experiment should be between 90% and 110%
for radiolabelled substances, whereas the initial recovery at the beginning of the experiment should be between 70% and 110% for non-labelled substances.

Observed value:

Mean recoveries of radioactivity in the water system of the tests with the low (2 µg/L) and high (10 µg/L) test concentrations were within the range of 91.5 % to 96.9 % AR and 93.0 % to 101.1 % AR, respectively.

Validity criteria fulfilled:

yes

Validity criteria 2

Validity criteria:

The reference substance must be degraded with two weeks.

Observed value:

The mean recovery of radioactivity in the water system of the reference samples was 99.3 % and 91.1 % AR at 0 and 14 DAT, respectively. After 14 days 83.4 % AR of the reference item was mineralised.

Validity criteria fulfilled:

yes

Conclusions:

Based on the test results the test substance can be considered very persistent

Executive summary:

The study of biodegradation of the test item with two different concentrations in natural aerobic surface water was performed with [¹⁴C]TPPT (O,O,O-Triphenylphosphothionate) under aerobic conditions in the dark using natural aerobic surface water from a large water body and dispersed sediment from the respective source. The test water has a dissolved organic carbon content of 11 mg/L and a BOD of 3.8 mg/L. Each vessel was filled with 500 mL water and between 5 mg and 10 mg dispersed sediment. The study was conducted in accordance with the OECD Guideline for Testing of Chemicals No. 309 (April 2004). To be able to determine the degradation rate and to follow the transformation products and due to the low water solubility of the test item, two different application rates (2 µg/L and 10 µg/L) of radio-labelled test item were applied. Based on the specific activity of 19.4 MBq/mg, this corresponds to an applied radioactivity of about 0.019 MBq and 0.097 MBq per vessel, respectively. The test water was incubated in the dark at 12 ± 2°C under constant bubbling of air through the water. The incubation period after treatment was 61 days. Traps for organic volatiles and carbon dioxide were used. Duplicate samples were taken for analysis at specified intervals up to 61 days after application. The water phase was extracted by adding C18 adsorber and extracting it with acetonitrile. The radioactivity (in extract and extracted water phase) was quantified by liquid scintillation counting and characterised by high performance liquid chromatography (HPLC). Thin layer chromatography (TLC) was used for confirmation of metabolites in selected samples.

Mean recoveries of radioactivity in the water system of the tests with the low (2 µg/L) and high (10 µg/L ) test concentrations were within the range of 91.5 % to 96.9 % AR and 93.0 % to 101.1 % AR, respectively. CO₂ was observed in amounts of 3.1 % AR for the low and 3.7 % AR for the high test concentration, respectively. No organic volatiles were detected (≤ 0.1 % AR). The amount of [¹⁴C]TPPT fluctuated between 80.2 % AR and 90.5 % AR for the low test concentration and between 84.2 % AR and 89.6 % for the high test concentration.

The calculated DT₅₀ values are above the 61 days test period. Thus, the test substance can be considered very persistent in water.