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Environmental fate & pathways

Biodegradation in water: screening tests

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Reference
Endpoint:
biodegradation in water: ready biodegradability
Type of information:
experimental study
Adequacy of study:
key study
Study period:
11 February 2016 to 10 March 2016
Reliability:
1 (reliable without restriction)
Rationale for reliability incl. deficiencies:
guideline study
Remarks:
GLP study performed according to OECD Guideline No. 301D. All validity criteria were fulfilled.
Qualifier:
according to
Guideline:
OECD Guideline 301 D (Ready Biodegradability: Closed Bottle Test)
Deviations:
no
Qualifier:
according to
Guideline:
EU Method C.4-E (Determination of the "Ready" Biodegradability - Closed Bottle Test)
Principles of method if other than guideline:
Not applicable
GLP compliance:
yes (incl. certificate)
Remarks:
05 March 2015
Specific details on test material used for the study:
PHYSICO-CHEMICAL PROPERTIES
- Specific Gravity at 20 °C: 0.8 -0.9
- Solubility in water: Not determined
Oxygen conditions:
aerobic
Inoculum or test system:
other: river water
Details on inoculum:
- Origin: River water was sampled from “L’Adour”, situated downstream of a plant treating domestic wastewater (Aureilhan, France).
- Pre-conditioning: The river water was aerated for about 7 days before use as inoculum at the test temperature to reduce endogenous respiration.
Duration of test (contact time):
28 d
Initial conc.:
2 mg/L
Based on:
test mat.
Parameter followed for biodegradation estimation:
O2 consumption
Details on study design:
TEST CONDITIONS
- Test water: Mineral medium, Reconstituted water, as prescribed by the OECD Guideline No. 301D
- Composition of medium: Mineral medium (8.5 mg KH2PO4, 21.75 mg K2HPO4, 33.40 mg Na2HPO4•2H2O, 0.50 mg NH4Cl; 36.40 mg CaCl2•2H2O, 22.50 mg MgSO4•7H2O and 0.25 mg FeCl3•6H2O) per litre
- River water was aerated for about 7 days before use as inoculum at the test temperature to reduce endogenous respiration.
- Administration of test item on silica gel (as recommended in ISO 10634 (1995) “Water quality - Guidance for the preparation and treatment of poorly water-soluble organic compounds for the subsequent evaluation of their biodegradability in an aqueous medium”)
- Test temperature: 21.1-22.1 °C throughout the test (average value: 21.8 °C)
- Continuous darkness: Yes

TEST SYSTEM
- Culturing apparatus: All-glass sealed with ground glass stoppers BOD (Biological Oxygen Demand) bottles of approximately 300 mL capacity.
- Inoculum: A concentration of inoculum of approximately 10^5 cells/L was used.
- Number of culture flasks/concentration: 10 bottles containing test item, inoculum, mineral medium and silica gel (test suspension)

SAMPLING
- Sampling frequency: Determinations of the concentration of dissolved oxygen using an oxygen electrode in duplicate vessels taken from each group (except the procedure control) were made at day 0, 7, 14, 21 and 28, and only at the start of the test and after 7 and 14 days of incubation for the procedure control.
- Temperature: Measured continuously in the growth chamber.

CONTROL AND BLANK SYSTEM
- Inoculum blank: 10 bottles containing inoculum, mineral medium and silica gel (inoculum blank + with silica gel); 10 bottles containing inoculum and mineral medium (inoculum blank without silica gel)
- Procedure control: 6 bottles containing reference substance, inoculum and mineral medium (procedure control)
- Toxicity control: No
Reference substance:
benzoic acid, sodium salt
Remarks:
2 mg/L
Preliminary study:
None
Key result
Parameter:
% degradation (O2 consumption)
Value:
81
Sampling time:
28 d
Details on results:
The test item was biodegraded by 81 % at day 28 in the Closed Bottle test. Substances are considered to be readily biodegradable in this test if oxygen consumption is equal to or greater than 60 % of the ThOD value within ten days from the day by which the level exceeding 10 %, and this pass level is attained within 28 days. In this study, the pass level of 60 % was reached after less than 7 days (approximately 6 days) upon achieving 10 % biodegradation. The test item therefore fulfilled the 10-day time window criterion for ready biodegradable compounds.
Calculated ThOD of the test item: 3.29 mg O2/mg
Results with reference substance:
Calculated ThOD of the reference substance: 1.67 mg O2/mg. The biodegradation percentage of the reference substance, sodium benzoate, was 74 % at day 14.

Table 5.2.1/1: Percentage biodegradation of the test item and of the reference substance throughout the test

Test item and reference substance

Replicates

Time (days)

0

7

14

21

28

Test Suspension

(2 mg/L)

Replicate 1

0.00

58.05

75.15

69.60

81.91

Replicate 2

0.00

63.68

74.39

75.68

81.00

Mean

0

61

75

73

81

Procedure Control (2 mg reference substance/L)

Replicate 1

0.00

76.95

75.00

N.A.

N.A.

Replicate 2

0.00

77.54

73.80

N.A.

N.A.

Mean

0

77

74

N.A.

N.A.

Validity criteria of the test:      

Inoculum blank: Oxygen depletion in the inoculum blank did not exceed 1.5 mg dissolved oxygen/L after 28 days.

Residual [O2]: Oxygen concentrations were > 0.5 mg/L in all bottles during the test period.

Replicate values: The differences of the replicate (test suspension) values at day 28 were less than 20 %.

Reference substance: The biodegradation percentage of the reference substance, sodium benzoate, was 74 % at day 14.

Thus all validity criteria of the test have been fulfilled in the present study.

Validity criteria fulfilled:
yes
Interpretation of results:
readily biodegradable
Conclusions:
The test item, REACTION MASS OF BETA-PHELLANDRENE AND D-LIMONENE AND L-LIMONENE, is considered as readily biodegradable based on the biodegradation percentage of 81% at day 28 and fulfilling the 10 days time window requirement (ca. less than 7 days).
Executive summary:

A ready biodegradability study was performed to assess the biotic degradation of the test item, REACTION MASS OF BETA-PHELLANDRENE AND D-LIMONENE AND L-LIMONENE, according to Guideline OECD 301 D and in compliance with GLP.

Ten BOD bottles were filled with mineral medium inoculated with river water (5 mL/L) and test item added at a nominal concentration of 2 mg/L with the help of silica gel. In the meantime, two series of ten blanks (with and without silica gel) were filled with inoculated mineral medium. Furthermore, six bottles containing the reference substance sodium benzoate (2 mg/L) were tested in order to check the procedure. The concentrations of dissolved oxygen in duplicate bottles from each group incubated at 22 ± 2°C in darkness were measured at the start of the test and thereafter once a week until the end of the test (28 days for each group except the procedure control until 14 days).

The validity of the test was demonstrated by an endogenous respiration < 1.5 mg O2/L and by a biodegradation of the reference substance of 74% of its ThOD after 14 days of incubation. Moreover, the differences of the replicate values at day 28 were less than 20% and oxygen concentrations were > 0.5 mg/L in all bottles during the test period. The test item was biodegraded by 81% at day 28 in the present study. Substances are considered to be readily biodegradable in the Closed Bottle test if oxygen consumption is equal to or greater than 60% of the ThOD value within ten days from the day by which the level exceeding 10%. In this study, the pass level of 60% was reached after less than 7 days (approximately 6 days) upon achieving 10% biodegradation.

Therefore, the test item is considered as readily biodegradable based on the biodegradation percentage of 81% at day 28 and fulfilling the 10 days time window requirement (ca. less than 7 days).

Description of key information

Test item Reaction mass of beta-phellandrene and d-limonene and l-limonene is considered as readily biodegradable according to the criteria of OECD Guideline 301 D.

Key value for chemical safety assessment

Biodegradation in water:
readily biodegradable

Additional information

A ready biodegradability study was performed to assess the biotic degradation of the test item, REACTION MASS OF BETA-PHELLANDRENE AND D-LIMONENE AND L-LIMONENE, according to Guideline OECD 301 D and in compliance with GLP.

The validity of the test was demonstrated by an endogenous respiration < 1.5 mg O2/L and by a biodegradation of the reference substance of 74% of its ThOD after 14 days of incubation. Moreover, the differences of the replicate values at day 28 were less than 20% and oxygen concentrations were > 0.5 mg/L in all bottles during the test period. The test item was biodegraded by 81% at day 28 in the present study. Substances are considered to be readily biodegradable in the Closed Bottle test if oxygen consumption is equal to or greater than 60% of the ThOD value within ten days from the day by which the level exceeding 10%. In this study, the pass level of 60% was reached after less than 7 days (approximately 6 days) upon achieving 10% biodegradation.

Therefore, the test item is considered as readily biodegradable based on the biodegradation percentage of 81% at day 28 and fulfilling the 10 days time window requirement (ca. less than 7 days).