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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:
04 April 2017 - 03 May 2017
Reliability:
1 (reliable without restriction)
Rationale for reliability incl. deficiencies:
guideline study
Qualifier:
according to
Guideline:
OECD Guideline 301 D (Ready Biodegradability: Closed Bottle Test)
Version / remarks:
17 July 1992
Deviations:
yes
Remarks:
ammonium chloride was omitted from the medium to prevent oxygen consumption due to nitrification and river water instead of an effluent/extract/mixture was used as inoculum.
Qualifier:
according to
Guideline:
ISO 10707 Water quality - Evaluation in an aqueous medium of the "ultimate" aerobic biodegradability of organic compounds - Method by analysis of biochemical oxygen demand (closed bottle test)
Version / remarks:
1994
GLP compliance:
yes (incl. certificate)
Specific details on test material used for the study:
Batch no: S 71699
CAS no multi-constituent: 8001-61-4
Composition (molecular formula of constituents):
C10H16 (0.3%)
C15H24 (86.3%)
C15H28 (5.6%)
Unknown (7.8%)
Solubility in water: insoluble in water
Stability: not relevant in biodegradation tests
Storage: at ambient temperature in the dark
Oxygen conditions:
aerobic
Inoculum or test system:
natural water: freshwater
Details on inoculum:
River water was sampled from the Rhine near Heveadorp, The Netherlands (29-03-2017). The nearest plant (Arnhem-Zuid) treating domestic wastewater biologically was 3 km upstream. The river water was aerated for 7 days before use to reduce the endogenous respiration (van Ginkel and Stroo, 1992). River water without particles was used as inoculum. The particles were removed by sedimentation after 1 day while moderately aerating.
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 bottles:
The test was performed in 0.30 L BOD (biological oxygen demand) bottles with glass stoppers.

Deionized water:
Deionized water containing no more than 0.01 mg/L Cu (ISO/IEC 17025; non-GLP analysis) was prepared in a water purification system.

Nutrients, stocks and administration:
The river water used in the Closed Bottle test was spiked per liter of water with 8.5 mg KH2PO4, 21.75 mg K2HPO4, 33.4 mg Na2HPO4·2H2O, 22.5 mg MgSO4·7H2O, 27.5 mg CaCl2, 0.25 mg FeCl3·6H2O. Ammonium chloride was not added to the river water to prevent nitrification. Accurate administering of the test substance was accomplished by preparing a solid stock of 3.0 mg of the test substance per g of silica gel in a 50-mL serum flask. Only part of the top layer of the silica gel was brought into contact with the test substance. The serum flask was closed with a screw top with alumiminium foil and the content was mixed vigorously. Subsequently, 0.20 g of silica gel with the test substance was added to the test bottles. The resulting concentration of test substance in the bottles was 2.0 mg/L. Next the bottles were filled with nutrient medium with inoculum and closed. Sodium acetate was added to the bottles using a stock solution of 1.0 g/L.

Test procedure:
The Closed Bottle test was performed according to the study plan. The study plan was developed from ISO Test Guidelines (1994). Use was made of 10 bottles containing only river water,10 bottles containing river water and silica gel, 10 bottles containing river water and silica gel with test substance, 6 bottles with river water and sodium acetate. The concentrations of the test substance, and sodium acetate in the bottles were 2.0 and 6.7 mg/L, respectively. Each of the prepared solutions was dispensed into the respective group of BOD bottles so that all bottles were completely filled without air bubbles. The zero time bottles were immediately
analyzed for dissolved oxygen using an oxygen electrode. The remaining bottles were closed and incubated in the dark. Two duplicate bottles of all series were withdrawn for analyses of the dissolved oxygen concentration at day 7, 14, 21, and 28.

Test conditions
The pH of the media was 7.9 at the start of the test. The pH of the medium at day 28 was 7.9 (controls) and 7.8 (test). Temperatures were within the prescribed temperature range of 22 to 24°C.
Reference substance:
acetic acid, sodium salt
Remarks:
Purity > 99%
Test performance:
The validity of the test is demonstrated by an endogenous respiration of 1.3 mg/L at day 28 . Furthermore, the differences of the replicate values at day 28 were less than 20%. The biodegradation percentage of the reference compound, sodium acetate, at day 14 was 87% of its theoretical oxygen demand. Finally, the validity of the test is shown by oxygen concentrations >0.5 mg/L in all bottles during the test period.
Key result
Parameter:
% degradation (O2 consumption)
Value:
77
Sampling time:
28 d
Parameter:
% degradation (O2 consumption)
Value:
71
Sampling time:
21 d
Parameter:
% degradation (O2 consumption)
Value:
55
Sampling time:
14 d
Parameter:
% degradation (O2 consumption)
Value:
32
Sampling time:
7 d
Details on results:
Copaiba Balsam oil was biodegraded by 77% at day 28 in the Closed Bottle test. The time-day window concept assumes that biodegradation of a single organic compound in a ready biodegradability test is a growth-linked process which follows an S-shaped growth curve. Copaiba Balsam oil is a mixture of chemicals (UVCB). The biodegradation kinetics (lag period, growth rate, and yield) of the individual chemicals in a mixture are not necessarily same. The biodegradation of a (fragrance) substance consisting of many constituents is therefore an addition of different biodegradation curves. It is thus possible that individual compounds meet the time window criterion whereas the biodegradability curve of the multiconstituent suggests that the test substance is not readily biodegradable. The time window that can be applied to Closed Bottle tests is therefore not considered applicable to this multiconstituent substance (OECD, 2006), even though it is met. Over 60% biodegradation was achieved in a period of approximately 14 days immediately following the attainment of 10% biodegradation at day 2 (Figure), therefore fulfilled the 14-day time window (10-day time window for other OECD 301 tests) criterion for ready biodegradable compounds. Copaiba Balsam oil should as a consequence be classified as readily biodegradable.
Results with reference substance:
The biodegradation percentage of the reference compound, sodium acetate, at day 14 was 87%.

Inhibition of the degradation of a well-degradable compound, e.g. sodium acetate by the test substance in the Closed Bottle test was not determined because possible toxicity of Copaiba Balsam oil to microorganisms degrading acetate is not relevant. Inhibition of the endogenous respiration of the inoculum by the test substance at day 7 was not detected. Therefore, no inhibition of the biodegradation due to the "high" initial test substance

concentration is expected.

Validity criteria fulfilled:
yes
Remarks:
See test performance
Interpretation of results:
readily biodegradable
Remarks:
Copaiba Balsam oil is a mixture of chemicals (UVCB), the time window applicable for the closed bottle test should therefore not be applied to this multi-constituent substance (OECD, 2006).
Conclusions:
The substance is biodegraded by 77% at day 28 in the Closed Bottle test and should therefore be classified as readily biodegradable.
Executive summary:

The ready biodegradability of Copaiba Balsam oil was investigated in a study conducted in accordance with OECDTG 301D (closed bottle test) and GLP. In this study river water was exposed to 2 mg/L of the substance for 28 days. The substance did not cause a reduction in the endogenous respiration. Furthermore, the validity criteria of the test were met. Copaiba Balsam oil was biodegraded by 77% at day 28 in the standard Closed Bottle screening test and should therefore be classified as readily biodegradable.

Description of key information

The ready biodegradability of Copaiba Balsam oil was investigated in a study conducted in accordance with OECDTG 301D (closed bottle test) and GLP. In this study river water was exposed to 2 mg/L of the substance for 28 days. The substance did not cause a reduction in the endogenous respiration. Furthermore, the validity criteria of the test were met. Copaiba Balsam oil was biodegraded by 77% at day 28 in the standard Closed Bottle screening test and should therefore be classified as readily biodegradable.

Key value for chemical safety assessment

Biodegradation in water:
readily biodegradable

Additional information