Phosphorodithioic acid, mixed O,O-bis(2-ethylhexyl and iso-Bu and pentyl) esters, zinc salts

Biodegradation in water: screening tests

Administrative data

Endpoint:

biodegradation in water: ready biodegradability

Type of information:

migrated information: read-across from supporting substance (structural analogue or surrogate)

Adequacy of study:

key study

Study period:

26 March 1998 - 12 November 1998

Reliability:

2 (reliable with restrictions)

Rationale for reliability incl. deficiencies:

other: Guideline study; well documented study report

Data source

Reference

Reference Type:

study report

Title:

Unnamed

Year:

1998

Report date:

1998

Materials and methods

Test guideline

Test guideline 1

Qualifier:

according to guideline

Guideline:

OECD Guideline 301 B (Ready Biodegradability: CO2 Evolution Test)

Deviations:

no

Test guideline 2

Qualifier:

according to guideline

Guideline:

EU Method C.4-C (Determination of the "Ready" Biodegradability - Carbon Dioxide Evolution Test)

Deviations:

no

GLP compliance:

yes

Test material

Test material information

Constituent 1

Reference substance name:

Phosphorodithioic acid, mixed O,O-bis(1,3-dimethylbutyl and iso-Pr) esters, zinc salts

EC Number:

283-392-8

EC Name:

Phosphorodithioic acid, mixed O,O-bis(1,3-dimethylbutyl and iso-Pr) esters, zinc salts

Cas Number:

84605-29-8

Molecular formula:

Not applicable

IUPAC Name:

Phosphorodithioic acid, mixed O,O-bis(1,3-dimethylbutyl and iso-Pr) esters, zinc salts

Details on test material:

The analogue substance CAS 84605-29-8 is structurally similar to this substance CAS 68988-45-4 which is also a zinc dialkyl dithiophosphate.

Study design

Oxygen conditions:

aerobic

Inoculum or test system:

activated sludge, non-adapted

Details on inoculum:

Activated sludge was collected from the Prospect Bay Wastewater Treatment Facility in April 1998. The sludge was sieved using a 2 mm screen and then aerated for approximately 4 hours. The sludge was homogenized in a blender at medium speed for approximately 2 minutes and then was allowed to settle for approximately 30 minutes. The supernatant was used as the inoculum for this study and was used the same data that it was prepared. A total suspended solids measurement and standard plate count were performed on the inoculum using procedures based on Standard Methods. The average measured total suspended solids concentration of the inoculum was 78.6 mg/L. The plates were incubated at 20 degrees C for approximately 48 hours and then the number of colony forming units was enumerated. The result of the standard plate count performed on the inoculum was 1.37x10^6 CFU/mL. The standard plate count result indicated that the inoculum used in this study contained an acceptable number of viable microbes according to the guideline.

Duration of test (contact time):

28 d

Initial test substance concentration

Initial conc.:

other: 10 mg carbon/liter Carbon determined by elemental analysis

Parameter followed for biodegradation estimation

Parameter followed for biodegradation estimation:

CO2 evolution

Details on study design:

TEST CONDITIONS
- Composition of medium: The test medium was a modified biochemical oxygen demand test dilution water. The test medium contained the following standard reagent solutions per liter of high quality Nanopure water: 1mL ammonium sulfate solution 4%; 1 mL of calcium chloride solution (2.75%); 4 mL of ferric chloride solution (0.025%); 1 mL of magnesium sulfate solution (2.25%); 2 ml of phosphate buffer solution (pH 7.2).

- Test temperature: 20C
- pH: The pH values ranged from 5.2 to 6.9.
- pH adjusted: yes/no

- Suspended solids concentration: The solids concentration of the test solution was 0.786 mg/L


TEST SYSTEM
- Culturing apparatus: The test chambers were ambered 4 liter bottles. The air entering the chambers was passed through Drierite to remove ambient moisture and then through Ascarite to produce CO2 free air. The air exiting the test chambers was passed through a series of three gas washing bottles each containing KOH to trap the CO2 that had evolved within the chamber. All chambers were aerated with CO2 free air for 24 hours at a rate of 50 to 100 mL per minute to purge the system of CO2. After the aeration period, the flow of CO2 was stopped and three CO2 traps each containing KOH were connected to the exit air lines of each chamber. An amount of test substance necessary to deliver 10 mg C/L was added to the treatment group test chambers by direct weight addition. An amount of reference substance necessary to deliver 10 mg C/L was added to the reference group test chambers by direct weight addition. The biodegradation was started by bubbling CO2 free air through each of the test chambers at a rate of 50 to 100 mL per minute. The CO2 produced from the degradation of organic carbon sources within the test chamber was trapped as K2CO3 in the KOH solution and the amount of inorganic carbon in the trapping solution was measured at various intervals during the study using a Shimadzu TOC 5000 carbon analyzer.

- Number of culture flasks/concentration: 3 Replicates


SAMPLING
The CO2 traps were removed for analysis on days 1, 4, 8, 11, 14, 19, 21, 25, and 29. The CO2 trap nearest the chamber was removed and analyzed for inorganic carbon. The two remaining traps were placed one position closer to the test chamber and a new trap was placed on the end of the series. On the 28h day of the test, an aliquot of the contents of each test chamber was removed and the pH determined. The contents of each chamber then were acidified by the addition of 3 mL of concentrated hydrochloric acid to drive off inorganic carbonate. The chambers were aerated overnight after which the trapping solutions closest to the test chambers were analyzed for inorganic carbon.

CONTROL AND BLANK SYSTEM
The test contained a blank control group, a reference group, and a treatment group. The blank control, reference, and treatment groups each contained three replicate test chambers

Reference substance

Reference substance:

benzoic acid, sodium salt

Results and discussion

% Degradation

Parameter:

% degradation (CO2 evolution)

Value:

1.5

Sampling time:

28 d

Details on results:

The mean percent theoretical CO2 evolved for the test substance was 1.5% (range 0.6 to 2.0%). The control chambers evolved an average of 6.5 milligrams of CO2 over the test period. The mean percent theoretical CO2 evolved for the reference substances was 99%. The temperature range recorded during the test was 18 to 20C. The average measured total suspended solids concentration of the inoculum was 78.6 mg/L. The result of the standard plate count performed on the inoculum was 1.37 x 10^6 CFU/mL. The measured pH values ranged from 5.2 to 6.9. The control chambers evolved an average of 6.5 milligrams of CO2 over the test period.

BOD5 / COD results

Results with reference substance:

The mean percent theoretical carbon dioxide evolved for sodium benzoate was 99%

Any other information on results incl. tables

Cumulative Percent of Theoretical Carbon Dioxide Evolved

Day	Benzoate Replicate values	OS13961BG Replicate values
1	0.0; 0.0; 0.0	0.0; 0.1; 0.0
4	13.1; 34.8; 50.0	1.2; 0.5; 0.0
8	64.4; 46.8; 73.2	1.1; 0.5; 0.0
11	85.6; 94.9; 94.5	1.3; 0.5; 0.1
14	91.1; 100.4; 98.4	1.2; 1.0; 1.8
19	92.9; 101.5; 100.4	1.8; 0.6; 1.9
21	92.7; 102.1; 100.9	2.0; 0.8; 2.0
25	94.1; 103.4; 101.2	2.2; 0.8; 2.2
29	94.6; 103.0; 100.4	2.0; 0.6; 1.9

Justification for Read Across from Analogue EC 283-392-8

Common Manufacturing Process:The submission substance (EC 273-527-9) and the analogue (EC 283-392-8) are produced under a common manufacturing process in which a phosphorodithioic acid ester intermediate, (RO)₂PS₂H, is produced by the reaction of phosphorus pentasulfide with an alcohol or a mixture of two alcohols of a similar class - branched alcohol containing C8, C5 and C4 carbons (submission substance) and C3 and C6 carbons (analogue). The intermediate is neutralized with zinc oxide to produce the final multicomponent substance. The reaction is performed in the presence of a highly refined base oil which accounts for 8 – 10.3 % of the final products.

Impurities:The level of impurities in the submission substance and the analogue (data source) is minimal. Impurities have been identified as residual, unreacted alcohols from the production of the phosphorodithioic acid ester intermediates (isobutanol, pentanol and 2-ethylhexanol in the submission substance and isopropyl and 1,3-dimethylbutyl alcohols in the analogue).

Same Chemical Category:The submission substance (EC 273-527-9) and the analogue (EC 283-392-8), generically referred to as ZDDPs, have been shown to have sufficient structural similarities to be included in the Zinc Dialkydithiophosphate Category (ZDDPs) in the United States Environmental Protection Agency High Production Volume (HPV) Chemical Challenge Program.

Structural Similarity:The primary feature accounting for the similarity of the submission substance (EC 273-527-9) and the analogue (EC 283-392-8) is the common organometallic core structure consisting of a central zinc metal bonded to four alkyldithiophosphate esters (ligands) by coordinate covalent bonds -Zn[(S₂P(OR)₂]_2.Structural variations between the submission substance and the analogue are related to the alkyl (R) groups of the alkyldithiophosphate ligands.

The analogue/data source (EC 283-392-8) is a multicomponent mixture of ZDDP monomers and dimers containing isopropyl dithiophosphate ligands, 1,3-dimethylbutyl dithiophosphate ligands and mixtures of isopropyl and 1,3-dimethylbutyl dithiophosphate ligands with a molecular weight range of 492 – 660 (monomer).

The submission substance (EC 273-527-9) is a multicomponent mixture of ZDDP monomers and dimers containing isobutyl dithiophosphate ligands, pentyl dithiophosphate ligands, 2-ethylhexyl dithiophosphates ligands and mixtures of isobutyl, pentyl and 2-ethylhexyl dithiophosphate ligands with a molecular weight range of 548 – 772 (monomer).

Tanimoto Fingerprint (ToxMatch Version 1.06 software) gives a similarity index greater than 0.8 (values range from 0, no similarity to 1, identical). Peer reviewed literature indicates that values greater than 0.6 are significantly similar.DSSTox similarity was 80% between the submission substance and the analogue.

Applicant's summary and conclusion

Validity criteria fulfilled:

yes

Interpretation of results:

other: Not readily biodegradable under test conditions

Conclusions:

The mean percent theoretical CO2 evolved for the test substance was 1.5% (range 0.6 to 2.0%). The control chambers evolved an average of 6.5 milligrams of CO2 over the test period. The mean percent theoretical CO2 evolved for the reference substances was 99%.

Executive summary:

The objective of the study was to measure the amount of carbon dioxide produced from the biodegradation of the test substance under the conditions tested. The test contained a blank control group, a reference group, and a treatment group, and each contained three replicate test chambers. The study was conducted according to procedures specified by the OECD Guideline for Testing of Chemicals, Method 301B and EEC Method C.4-C. The test substance was administered to the treatment group test chambers by direct weight addition at a concentration of 10 mg Carbon per liter. After 28 days the cumulative percent of theoretical carbon dioxide evolved for the test material was 1.5%. The solids concentration of the test solution was within the acceptable limit specified I the test guideline. The standard plate count result indicated that the inoculum used in this study contained an acceptable number of viable microbes according to the guidelines