Propane-1,2,3-triyl 3,5,5-trimethylhexanoate

Biodegradation in water: screening tests

Link to relevant study record(s)

Reference

Endpoint:

biodegradation in water: ready biodegradability

Type of information:

experimental study

Adequacy of study:

key study

Study period:

31 Jul-29 Aug 2012

Reliability:

1 (reliable without restriction)

Rationale for reliability incl. deficiencies:

guideline study

Qualifier:

according to guideline

Guideline:

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

Qualifier:

equivalent or similar to guideline

Guideline:

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

Qualifier:

equivalent or similar to guideline

Guideline:

EPA OPPTS 835.3110 (Ready Biodegradability)

GLP compliance:

yes (incl. QA statement)

Remarks:

The Department of Health of the Government of the United Kingdom

Oxygen conditions:

aerobic

Inoculum or test system:

activated sludge, domestic (adaptation not specified)

Details on inoculum:

- Source of inoculum/activated sludge: A mixed population of activated sludge microorganisms was obtained from the Severn Trent Water Plc sewage treatment plant at Loughborough, Leicestershire, UK on 30 July 2012.
- Preparation of inoculum for exposure: the activated sludge was washed twice by settlement and resuspension in mineral medium to remove any excessive amounts of dissolved organic carbon (DOC) that may have been present. The washed sample was then maintained on continuous aeration at approximately 21 °C and used on the day of collection.
- Suspended solids concentration: 2.9 g/L prior to use

Duration of test (contact time):

28 d

Initial conc.:

14.2 mg/L

Based on:

test mat.

Initial conc.:

10 mg/L

Based on:

other: Carbon

Parameter followed for biodegradation estimation:

CO2 evolution

Details on study design:

TEST CONDITIONS
- Composition of medium: According to OECD Guideline:
Solution a KH2PO4 8.50 g/L
K2HPO4 21.75 g/L
Na2HPO4.2H2O 33.40 g/L
NH4Cl 0.5 g/L

Solution b CaCl2 27.5 g/L
Solution c MgSO4.7H2O 22.5 g/L
Solution d FeCl3.6H20 0.25 g/L
10 mL of solution a and 1 mL of solutions b, c and d were added to 1 L of purified water
- Solubilising agent (type and concentration if used): Due to the low solubility of the test substance in water (< 0.05 mg/L) a pre-study on the solubility and stability of the substance was performed to establish the most suitable mode of preparation, which turned out to be the preparation of the solution in a volatile solvent. Therefore, for the final test, chloroform was used to prepare the stock solution. An amount of test item (1065 mg) was dissolved in 10 mL of chloroform to give a 1065 mg/10 mL solvent stock solution. An aliquot (400 μL) of this solvent stock solution was dispensed onto a filter paper and the solvent allowed to evaporate to dryness for approximately 15 minutes. The filter paper was dispersed in approximately 400 mL of mineral medium with the aid of high shear mixing (approximately 7500 rpm, 5 minutes) prior to addition to inoculated mineral medium. The volume was then adjusted to 3 liters.
- Test temperature: ca. 22 °C
- pH: 7.4 ± 0.2
- pH adjusted: yes
- Suspended solids concentration: 30 mg/L
- Continuous darkness: yes

TEST SYSTEM
- Culturing apparatus: 5 L test culture vessels, each containing 3 L of solution.
- Number of culture flasks/concentration: 2 per blank control, reference control and test item vessels. Only one replicate in the toxicity control
- Method used to create aerobic conditions: the test vessels were sealed and CO2-free air bubbled through the solution at a rate of 30-100 mL/min per vessel and stirred continuously by magnetic stirrer.
- Measuring equipment: TOC analyzer. Samples (50, 135, 180, 300 or 540 μL) were injected into the IC (Inorganic Carbon) channel of the TOC analyzer. Inorganic carbon analysis occurs by means of the conversion of an aqueous sample to CO2 by orthophosphoric acid using zero grade air as the carrier gas. Calibration was by reference solutions of sodium carbonate (Na2CO3). Each analysis was carried out in triplicate.
- Details of trap for CO2 and volatile organics if used: the CO2 produced by degradation was collected in two 500 mL Dreschel bottles containing 350 mL of 0.05 M NaOH. The CO2 absorbing solutions were prepared using purified de-gassed water

SAMPLING
- Sampling frequency: Samples (2 mL) were taken from the first CO2 absorber vessels on Days 0, 2, 6, 8, 10, 14, 21, 28 and 29. The second absorber vessels were all sampled on Days 0 and 29.
- Sampling method: The samples taken on Days 0, 2, 6, 8, 10, 14, 21, 28 and 29 were analyzed for CO2 immediately. On Day 28, 1 mL of concentrated hydrochloric acid was added to each vessel to drive off any inorganic carbonates formed. The vessels were resealed, aerated overnight and the final samples taken from both absorber vessels on Day 29.
- Inorganic Carbon/Total Carbon Analysis: Samples (30 mL) were removed from the test item vessels on Day 0 prior to the addition of the test item in order to calculate the Inorganic Carbon content in the test media. The samples were filtered through 0.45 μm Gelman AcroCap filters (first approximate 5 mL discarded in order to pre-condition the filter) prior to DOC analysis. Samples (30 mL) were also removed from the inoculum control vessels on Day 0 and filtered through 0.45 μm Gelman AcroCap filters (first approximate 5 mL discarded in order to pre-condition the filter) prior to DOC analysis. IC/TC analysis of the test item dispersions after dosing was not possible due to the insoluble nature of the test item in water.

CONTROL AND BLANK SYSTEM
- Inoculum blank: yes, 2 replicates
- Reference control: yes, 2 replicates
- Toxicity control: yes, 1 replicate

Reference substance:

benzoic acid, sodium salt

Remarks:

10 mg C/L

Parameter:

% degradation (CO2 evolution)

Value:

60

Sampling time:

29 d

Details on results:

- Acidification of the test vessels on Day 28 followed by the final analyses on Day 29 was conducted according to the methods specified in the Test Guidelines. This acidification effectively kills the microorganisms present and drives off any dissolved CO2 present in the test vessels. Therefore any additional CO2 detected in the Day 29 samples originated from dissolved CO2 that was present in the test vessels on Day 28 and hence the biodegradation value calculated from the Day 29 analyses is taken as being the final biodegradation value for the test item.
- Biodegradation in the toxicity control reached 62% after 14 days, fulfilling the validity criterion (=> 25% biodegradation within 14 days). Therefore, the test substance is considered to be non-inhibitory of activated sludge microorganisms activity.

Results with reference substance:

Sodium benzoate reached 96% biodegradation after 14 days of exposure, and therefore fulfills the validity criterion for the reference substance (=> 60% within 14 days)

Table 1. Percentage biodegradation values

Day	% Degradation Reference control (Sodium benzoate)	% Degradation Test Item	% Degradation Toxicity control (Test Item plus Sodium benzoate)
0	0	0	0
2	61	6	38
6	83	11	47
8	80	14	45
10	108	35	60
14	96	47	62
21	101	58	72
28	89	75	84
29	91	60	82

Validity criteria fulfilled:

yes

Interpretation of results:

readily biodegradable, but failing 10-day window

Conclusions:

The pass level of 60% was reached in a ready test (OECD 301) within 28 days, but the criteria for "readily biodegradable" were not met (10-day window)

Description of key information

Readily biodegradable, but failing 10-day window: 60% in 28 days (OECD 301B)

Additional information

One study evaluating the biodegradation potential of propane-1,2,3-triyl 3,5,5-trimethylhexanoate (CAS No. 56554-53-1) is available (Bayliss, 2012). The test was conducted according to OECD Guideline 301B, under GLP conditions. Activated sludge microorganisms were exposed to the test substance for a period of 28 days, at 22°C and biodegradation followed by measuring the CO2 production in the test vessels. After 28 days, propane-1,2,3-triyl 3,5,5-trimethylhexanoate reached 60% biodegradation, not fulfilling the 10-day window criterion and therefore, the test substance cannot be considered as readily biodegradable. Nevertheless, the results obtained in the toxicity control (62% biodegradation within 14 days) show that propane-1,2,3-triyl 3,5,5-trimethylhexanoate is not toxic to and therefore non-inhibitory of activated sludge microorganisms activity.