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Environmental fate & pathways

Biodegradation in water: screening tests

Administrative data

Endpoint:
biodegradation in water: ready biodegradability
Type of information:
experimental study
Adequacy of study:
key study
Study period:
11 March - 8 April 1991
Reliability:
2 (reliable with restrictions)
Rationale for reliability incl. deficiencies:
guideline study without detailed documentation
Remarks:
The study was carried out in accordance with a modified OECD 301B (headspace test) and in accordance with GLP principles. However, the report is very summary (for example the amount of replicate bottles of each treatment is unclear and only the calculated percentages biodegradation were reported, thus no results of the inorganic carbon analyses) and a toxicity control was not included in the study. Furthermore, the report and QA statement have been issued 5 years after the experimental phase. For this reason a reliability 2 was assigned.

Data source

Reference
Reference Type:
study report
Title:
Unnamed
Year:
1991
Report Date:
1993

Materials and methods

Test guideline
Qualifier:
equivalent or similar to
Guideline:
OECD Guideline 301 B (Ready Biodegradability: CO2 Evolution Test)
Version / remarks:
(1981)
Deviations:
yes
Remarks:
see "principles of method"
Principles of method if other than guideline:
Sealed vessel test based on OECD Guideline 301B and studies conducted by the OECD aimed at harmonising the various tests of ready biodegradability. Development and validation of the method is described in Birch et al 1991. The study is equivalent or similar to OECD guideline 310, which was published in 2006 as OECD guideline 310. This corresponds to a sealed vessel test in which CO2 production is determined by measuring the increase in the concentration of inorganic carbon (IC). The test system is closed and is therefore suitable for the examination of volatile substances.

Birch, R.R. and R.J. Fletcher (1991). the application of dissolved inorganic carbon measurements to the study of aerobic biodegradability. Chemosphere 23: 507-524.
GLP compliance:
yes

Test material

Reference
Name:
Unnamed
Type:
Constituent
Test material form:
other: Oily Liquid
Details on test material:
- Name of test material (as cited in study report): ELINTAAL FORTE (Q)
- Physical state: Oily liquid (colourless)
- Storage condition of test material: At room temperature in the dark

Study design

Oxygen conditions:
aerobic
Inoculum or test system:
other: Secondary effluent from activated sludge plant (non-adapted)
Details on inoculum:
- Source of inoculum/activated sludge (e.g. location, sampling depth, contamination history, procedure): Secondary effluent from an unacclimatised activated sludge plant at Unilever Research Laboratory North
- The inoculum used was 10% by volume of activated sludge plant secondary effluent, filtered through a Whatman filter paper to remove coarse particulate matter. The level of dissolved inorganic carbon (DIC) was reduced by sparging the filtered effluent with nitrogen after prior adjustment of the pH to 6.5.
Duration of test (contact time):
28 d
Initial test substance concentration
Initial conc.:
10 mg/L
Based on:
other: Organic carbon
Parameter followed for biodegradation estimation
Parameter followed for biodegradation estimation:
inorg. C analysis
Details on study design:
TEST CONDITIONS
- Composition of medium: The mineral salts medium was as recommended in the OECD Ring Test (1988) on the harmonisation of ready biodegradability tests with the following deviation: the Ferric chloride stock solution contained 0.25 g not 0.2 g. The slight difference in composition of the mineral salts medium should have no effect on the results.
- Test temperature: 18 - 22 °C
- pH: The pH of the secondary effluent that was used as inoculum was adjusted to 6.5

TEST SYSTEM
The test is conducted in 160 mL vessels (hypovials) containing 100 mL mineral salts medium inoculated with secondary effluent and the test substance. Multiple vessels are prepared per substance sealed with a butyl rubber septum and an aluminium crimp seal. The headspace in each vessel has a volume of 60 mL and, when filled with air, contains approximately 6 times the mass of oxygen required for the complete oxidation of the substance. The sealed vessels are incubated at 20 °C on a rotary shaker.

SAMPLING
- Sampling frequency: Day 4, 8, 14, 22, 24 and 28
- Sampling method: a vessel was removed and concentration of carbon dioxide in the headspace gas determined. The seal was then broken and the concentration of inorganic carbon in the test medium was also determined.

CONTROL AND BLANK SYSTEM
- Inoculum blank: yes
- Abiotic sterile control: No
- Toxicity control: No
Reference substance
Reference substance:
other:
Remarks:
A standard reference substance of known biodegradability was not tested in parallel to check the test procedure & viability of the inoculum. However, a number of other test items were tested in parallel that met the ready biodegradability crieria.

Results and discussion

% Degradationopen allclose all
Parameter:
% degradation (inorg. C analysis)
Value:
7.1
Sampling time:
4 d
Parameter:
% degradation (inorg. C analysis)
Value:
22.1
Sampling time:
8 d
Parameter:
% degradation (inorg. C analysis)
Value:
44
Sampling time:
14 d
Parameter:
% degradation (inorg. C analysis)
Value:
57.2
Sampling time:
22 d
Key result
Parameter:
% degradation (inorg. C analysis)
Value:
62.3
Sampling time:
28 d
Remarks on result:
other: 95% CI: 53.8 - 70.8
Details on results:
The 10-d window begins when the degree of biodegradation has reached 10% (i.e. between day 4 and day 8 for Elintaal Forte). Therefore the end of the 10d window is between days 14-18. Given the fact that the % biodegradation was only 44% at day 14 and 57.2% at day 22, the 10-day window criterion was not met.
However, Elintaal is a mixture of diastereoisomers and thus the degradation curve will be the sum of different individual growth curves associated with each isomer. The OECD Guidelines for Testing of Chemicals, Section 3. Part 1: Principles and strategies related to the testing of degradation of organic chemicals (adopted 23 March 2006), states that if a substance consists of “constituents with different chain-lengths, degree and/or site of branching or stereo-isomers, even in their most purified commercial forms” and “it is anticipated that a sequential biodegradation of the individual structures is taking place, then the 10-day window should not be applied to interpret the results of the test.” The 10-day time-window should therefore be ignored as a pass / fail criterion for Elintaal. Thus, Elintaal should be regarded as readily biodegradable according to this test based only on the biodegradation percentage of 62.3% at day 28.

BOD5 / COD results

Results with reference substance:
- No reference substance was tested. However, several of the other test substances tested in parallel to Elintaal Forte met the validity criteria for reference compounds of > 60% biodegradation within 14 days (delta-decalactone, menthone, methyl nonyl ketone, prenyl acetate, tridecene-2-nitrile and Cervolide). Thus, it can be concluded that the inoculum was sufficiently active.

Applicant's summary and conclusion

Validity criteria fulfilled:
yes
Remarks:
Substances tested in parallel met the "reference compound" validity criterion of > 60% biodegradation by day 14. Confidence limits of 53.8 to 70.8% for Elintaal Forte, indicate that the extreme replicate values differed by < 20% at test end.
Interpretation of results:
readily biodegradable
Conclusions:
Under the conditions of a modified OECD 301B sealed vessel test, 62.3% biodegradation (95% CI: 53.8-70.8%) was observed for Elintaal forte (Q) after 28 days. The 10-day window criterion is not applicable to Elintaal Forte because it is a mixture of diastereoisomers. Thus, Elintaal forte is regarded as readily biodegradable according to this test based on > 60% degradation at day 28.
Executive summary:

The ready biodegradability of Elintaal forte (Q) was tested according to a modified OECD 301B (headspace test) and principles of Good Laboratory Practice. The test substance was tested at a nominal concentration of 10 mg/L organic carbon. After 28 days, 62.3% biodegradation of Elintaal forte (Q) was observed (95% CI: 53.8 -70.8%). The substance was concluded to be readily biodegradable, but failing to pass the 10-day window under the conditions of this test. However, Elintaal forte is a mixture of diastereoisomers and thus the degradation curve will be the sum of different individual growth curves associated with each isomer. The OECD Guidelines for Testing of Chemicals, Section 3. Part 1: Principles and strategies related to the testing of degradation of organic chemicals (adopted 23 March 2006), states that if a substance consists of “constituents with different chain-lengths, degree and/or site of branching or stereo-isomers, even in their most purified commercial forms” and “it is anticipated that a sequential biodegradation of the individual structures is taking place, then the 10-day window should not be applied to interpret the results of the test.” The 10-day time-window should therefore be ignored as a pass / fail criterion for Elintaal Forte. Thus, Elintaal forte should be regarded as readily biodegradable according to this test based only on the biodegradation percentage of 62.3% at day 28.