Extract obtained from defatted powder of Theobroma cacao (Malvaceae) by extraction with ethanol

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:

from 2018

Reliability:

1 (reliable without restriction)

Rationale for reliability incl. deficiencies:

guideline study

Remarks:

The study is performed according to the OECD 301F guideline and under GLP compliance.

Qualifier:

according to guideline

Guideline:

OECD Guideline 301 F (Ready Biodegradability: Manometric Respirometry Test)

Deviations:

no

Principles of method if other than guideline:

Not applicable

GLP compliance:

yes (incl. QA statement)

Specific details on test material used for the study:

None

Oxygen conditions:

aerobic

Inoculum or test system:

activated sludge, domestic, adapted

Details on inoculum:

- Source of inoculum/activated sludge: Pre-conditioning will consist in aerating sludge (in mineral medium) for 5-7 days before the test and at the test temperature.
- Laboratory culture: A filtration through a fine sieve (about 1 mm) will be performed in order to remove coarse particles. After removal of any coarse particles, the sludge will be washed by decantation in a mineral medium until sludge is considered free from excess substrate or inhibitor.
- Inoculum preconditioning: Pre-conditioning will consist in aerating sludge (in mineral medium) for 5-7 days before the test and at the test temperature.

Duration of test (contact time):

28 d

Initial conc.:

100 mg/L

Based on:

test mat.

Parameter followed for biodegradation estimation:

O2 consumption

Details on study design:

TEST ITEM PREPARATION:
- The test item is poorly soluble in water and in order to improve the dissolution the test item was dissolved in silicone oil AR 20 up to final concentration of 1 % v/v. Around 25 mg of test item was stirred with 2.5 mL silicone oil AR 20 by magnetically stirring until completely dissolved.
Then, this solution was directly supplemented or completed into 250 mL with mineral medium and sludge directly in each test vessels
The test medium was the mineral medium, a weakly saline aqueous medium.
The test item was tested at 100 mg/L in mineral medium containing 1 % v/v of silicone oil AR 20.

TEST PRINCIPLE
A measured volume of inoculated medium containing a known concentration of the test item (100 mg/L) as the nominal sole source of Carbon, is stirred in a closed vessel at a constant temperature up to 28 days with possible extension to 60 days.
Microbial respiration consumes O2 and releases CO2 which will be absorbed by soda lime pellets present in the headspace of flasks. This produces a decrease in pressure in the flask proportional to the amount of oxygen taken up by the microbial population during biodegradation, which will be expressed as a percentage of ThOD (Theoretical Oxygen Demand).
Allowance will be made for the endogenous activity of the inoculum by running parallel blanks with inoculum but without the test item. Furthermore, in order to check the procedure, a reference item control (sodium benzoate + inoculum) and a toxicity control (sodium benzoate + inoculum + test item) will be run in parallel.
The pH of the contents will be measured at the start and the end of the experiment.

TEST CONDITIONS
- Composition of medium: Mineral medium according to OECD 301F guideline
- Continuous darkness: Yes
- Test performed in closed vessels: Yes
- The pH of the contents is measured at the start and the end of the experiment.

Reference substance:

benzoic acid, sodium salt

Preliminary study:

No data

Test performance:

No data

Parameter:

% degradation (O2 consumption)

Value:

83.59

Sampling time:

28 d

Details on results:

Based on ThODNH4, the degradation rate of the test item “COCOA OLEORESIN” reached 60% within the 10-Day window
on Day 6 (61.85%) and after 28 days of incubation the biodegradation reached 83.59% (80.12% to 86.24%)
Based on ThODNO3, the degradation rate of the test item “COCOA OLEORESIN” reached 60% within the 10-Day window
on Day 9 (63.13%) and after 28 days of incubation the biodegradation reached 72.07% (69.08 to 74.36%)

Results with reference substance:

>60% (92.28% to 95.28%)

Criteria	Validity	Validity for the test
Oxygen uptake of the inoculum control in 28 days	< 60 mg/L	Yes (12.7 to 14.1 mg/L) (Silicone: 19.7 to 21.1 mg/L)
Biodegradability of the reference item within 14 days	> 60%	Yes (92.28 to 95.28%)
Biodegradability of the toxicity control within 14 days	> 25%	ThODNH4	ThODNO3
		Yes(80.00 and 81.14%)	Yes(72.79 and 73.83%)
Variation between the test item replicates at 28 days	< 20%	Yes (3.76%)
pH of the inoculum control at 28 days	6 to 8.5	Yes(7.2)

Validity criteria fulfilled:

yes

Interpretation of results:

readily biodegradable

Conclusions:

Under the study conditions, the test item is considered as ready biodegradable.

Executive summary:

The aim of this study was to determine the biodegradability of the test item «COCOA OLEORESIN», according to the OECD 301F guideline, within an experimental period of 28 days by respirometric method.

The degradation rate of the test item «COCOA OLEORESIN» reached 83.59%, following the ThODNH4 and 72.07% following ThODNO3 after 28 days of incubation.

Under conditions of OECD Guideline 301F, the test item “COCOA OLERESIN”, is considered to be readily biodegradable.

Description of key information

The registered substance is regarded as readily biodegradable.

Key value for chemical safety assessment

Biodegradation in water:

readily biodegradable

Type of water:

freshwater

Additional information

The aim of this study was to determine the biodegradability of the test item «COCOA OLEORESIN», according to the OECD 301F guideline, within an experimental period of 28 days by respirometric method.

The degradation rate of the test item «COCOA OLEORESIN» reached 83.59%, following the ThODNH4 and 72.07% following ThODNO3 after 28 days of incubation.

Under conditions of OECD Guideline 301F, the test item “COCOA OLERESIN”, is considered to be readily biodegradable.