Molybdenum, bis(dibutylcarbamodithioato)di-μ-oxodioxodi-, sulfurized

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:

07 July 2016 to 25 July 2018

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)

Deviations:

no

Qualifier:

according to guideline

Guideline:

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

Deviations:

no

Qualifier:

according to guideline

Guideline:

EPA OPPTS 835.3110 (Ready Biodegradability)

Deviations:

no

GLP compliance:

yes (incl. QA statement)

Specific details on test material used for the study:

CAS RN: 68412-26-0
Physical state/Appearance: Yellow powder
Purity: This substance has an Unknown or Variable composition, is a Complex reaction product, or a Biological material (UVCB).

Oxygen conditions:

aerobic

Inoculum or test system:

activated sludge, domestic, non-adapted

Details on inoculum:

Determination of the suspended solids level of the activated sewage sludge was carried out by filtering a sample (100 mL) of the washed activated sewage sludge by suction through pre-weighed GF/A filter paper using a Buchner funnel. Filtration was then continued for a further 3 minutes after rinsing the filter three successive times with 10 mL of deionized reverse osmosis water. The filter paper was then dried in an oven at approximately 105 ºC for at least 1 hour and allowed to cool before weighing. This process was repeated and after a further 1-Hour drying period an identical weight was determined thereby indicating that complete dryness had been attained. The suspended solids concentration was equal to 3.2 g/L prior to use.

Duration of test (contact time):

28 d

Initial conc.:

10 other: mg C/L

Based on:

test mat.

Parameter followed for biodegradation estimation:

CO2 evolution

Details on study design:

Based on the results of a preliminary solubility/dispersibility trial and following the recommendations of the International Standards Organisation (ISO 1995), the test item was dissolved in an auxiliary volatile solvent (chloroform) prior to being adsorbed onto a filter paper and subsequent dispersal in test media. Using this method the test item is evenly distributed throughout the test medium and the surface area of test item exposed to the test organisms is increased thereby increasing the potential for biodegradation.

The following test preparations were prepared and inoculated in 5 liter glass test culture vessels each containing 3 liters of solution:

a) Duplicate inoculated control vessels consisting of inoculated mineral medium plus a filter paper.
b) Duplicate procedure control vessels containing the reference item (sodium benzoate) in inoculated mineral medium plus a filter paper to give a final concentration of 10 mg carbon/L.
c) Duplicate test item vessels containing the test item on a filter paper in inoculated mineral medium to give a final concentration of 10 mg carbon/L.
d) A single toxicity control consisting of the test item on a filter paper plus the reference item in inoculated mineral medium to give a final concentration of 20 mg carbon/L.

A filter paper with chloroform evaporated to dryness was added to the inoculum control and procedure control vessels in order to maintain consistency between these vessels and the test item vessels.

One additional vessel containing water only was incubated under the same conditions as the test vessels to allow the temperature to be checked daily.

An abiotic control is designed to concurrently evaluate chemical degradation of the test item. An abiotic control was omitted as it was considered that carbon dioxide (CO2) would not be produced from any such degradation of the test item.

Each vessel was inoculated with the prepared inoculum at a final concentration of 30 mg suspended solids (ss)/L. The test was carried out in a temperature controlled room, in darkness and the temperature of the additional vessel containing water which was incubated under the same conditions as the test ranged from 21 to 24 ºC.

Approximately 24 hours prior to addition of the test and reference items the vessels were filled with 2400 mL of mineral medium and 28.1 mL of inoculum and aerated overnight. On Day 0 the test and reference items were added, as applicable, and the pH of all vessels measured using a Hach HQ40d Flexi handheld meter. The pH was adjusted to pH 7.4 ± 0.2, as necessary, using diluted hydrochloric acid or sodium hydroxide solution prior to the volume in all the vessels being adjusted to 3 liters by the addition of mineral medium, which had been purged overnight with CO2-free air.

The vessels were sealed and CO2-free air bubbled through the solution at a rate of 30 to 100 mL/min per vessel and stirred continuously by magnetic stirrer.

The CO2-free air was produced by passing compressed air through a glass column containing self-indicating soda lime (Carbosorb®) granules.

The CO2 produced by degradation was collected in two 500 mL Dreschel bottles containing 350 mL of 0.05 M NaOH. These CO2 absorbing solutions were prepared using purified water.

Observations
The appearance of the test preparations was recorded on Days 0, 6, 13, 20 and 27.

pH Measurements
The pH of the test preparations was determined on Day 0 and on Day 28 prior to acidification with hydrochloric acid, using a Hach HQ40d Flexi handheld meter.

Temperature Measurements
One additional vessel containing water only was incubated under the same conditions as the test vessels. Temperature was measured daily in this vessel using a Hanna Instruments HI 93510 digital thermometer.

Statistical Analysis
Statistical analysis of the Day 29 IC values for the inoculum control and test item vessels was carried out using a Student’s t-test to determine any statistically significant differences between the test and inoculum control groups. All statistical analyses were performed using the SAS computer software package (SAS, 1999 - 2001).

Reference substance:

benzoic acid, sodium salt

Test performance:

The average total CO2 evolution in the two inoculum control vessels on Day 28 was 26.88 mg/L and therefore satisfied the validation criterion given in the OECD Test Guidelines.

The IC content of the test item suspension in the mineral medium at the start of the test was below 5% of the TC content and hence satisfied the validation criterion given in the OECD Test Guidelines.

The CO2 production at the end of the test for the replicate test item vessels was negligible and the difference between the replicate vessels was <20%, satisfying the validation criterion given in the OECD Test Guidelines.

Biodegradation in the procedure and toxicity controls satisfied the validation criterion given in the OECD Test Guidelines.

Key result

Parameter:

% degradation (CO2 evolution)

Value:

0

Sampling time:

28 d

Details on results:

Acidification of the test vessels on Day 28, followed by analysis on Day 29 was conducted according to the methods specified in the test guidelines. This acidification effectively kills the micro-organisms 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.

The results of the inorganic carbon analysis of samples from the first absorber vessels on Day 29 showed an increase in all replicate vessels with the exception of the toxicity control.

Inorganic carbon analysis of the samples from the second absorber vessels on Day 29 confirmed that no significant carry-over of CO2 into the second absorber vessels occurred.

The test item attained 0% biodegradation after 28 days and therefore cannot be considered to be readily biodegradable according to the criteria set forth in OECD Guideline No. 301B.

Statistical analysis of the Day 29 IC values for the inoculum control and test item vessels showed there were no statistically significant differences (P≥ 0.05) between the inoculum control and the test item.  The test item was therefore considered not to have a toxic effect on the sewage sludge micro-organisms used in the study, which was confirmed by the toxicity control results.

 

The toxicity control attained 37% biodegradation after 14 days and 32% biodegradation after 28 days thereby satisfying the validation criterion given in the OECD Test Guidelines and confirming that the test item did not exhibit an inhibitory effect on the sewage treatment micro-organisms used in the test.  The slight decrease in biodegradation between Days 14 and 28 was attributed to sampling/analytical variation.

 

The reference item, sodium benzoate, attained 74% biodegradation by Day 14 (and within 10 days of biodegradation exceeding 10%) and 66% biodegradation after 28 days thereby satisfying the validation criterion given in the OECD Test Guidelines and confirming the suitability of the inoculum and test conditions.  The slight decrease in biodegradation between Days 14 and 28 was attributed to sampling/analytical variation.

 

Percentage Biodegradation Values

Day	% Biodegradation
	Procedure Control	Test Item	Toxicity Control
0	0	0	0
2	44	0	24
6	59	0	30
8	58	0	32
10	66	0	41
14	74	1	37
21	57^**	0	40
28	66	0	40
29*	66	0	32

* Day 29 values corrected toinclude carry-over of CO2 detected in Absorber 2, if any.

^**The drop in biodegradation between Days 14 and 21 was considered to be duesampling/analytical variation.

 

 

 

 

 

Validity criteria fulfilled:

yes

Interpretation of results:

under test conditions no biodegradation observed

Conclusions:

The test item attained 0% biodegradation after 28 days and therefore cannot be considered to be readily biodegradable under the strict terms and conditions of OECD Guideline No. 301B.

Executive summary:

A study was performed to assess the ready biodegradability of the test item in an aerobic aqueous medium. The method followed was designed to be compatible with the OECD Guidelines for Testing of Chemicals (1992) No. 301B, "Ready Biodegradability: CO2 Evolution Test", Method C.4-C of Commission Regulation (EC) No. 440/2008, and US EPA Fate, Transport, and Transformation Test Guidelines OCSPP 835.3110 (Paragraph (m)).

The test item, at a concentration of 10 mg carbon/L, was exposed to activated sewage sludge micro-organisms with mineral medium in sealed, glass culture vessels in the dark at temperatures between 21 and 24 ºC for 28 days.

Based on the results of a preliminary solubility/dispersibility trial and following the recommendations of the International Standards Organisation (ISO 1995), the test item was dissolved in an auxiliary volatile solvent (chloroform) prior to being adsorbed onto a filter paper and subsequent dispersal in test media. Using this method the test item is evenly distributed throughout the test medium and the surface area of test item exposed to the test organisms is increased thereby increasing the potential for biodegradation.

The biodegradation of the test item was assessed by the determination of carbon dioxide produced. Control solutions with inoculum and the reference item, sodium benzoate, together with a toxicity control were tested concurrently for validation purposes.

All validation criteria were met in this study. The test item attained 0% biodegradation after 28 days and therefore cannot be considered readily biodegradable as defined in OECD Guideline No. 301B.

Description of key information

The test item attained 0% biodegradation after 28 days and therefore cannot be considered to be readily biodegradable under the strict terms and conditions of OECD Guideline No. 301B.

Key value for chemical safety assessment

Biodegradation in water:

under test conditions no biodegradation observed

Type of water:

freshwater