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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:
2009-02-19 to 2009-03-20
Reliability:
1 (reliable without restriction)

Data source

Reference
Reference Type:
study report
Title:
Unnamed
Year:
2009
Report Date:
2009

Materials and methods

Test guideline
Qualifier:
according to
Guideline:
OECD Guideline 301 B (Ready Biodegradability: CO2 Evolution Test)
Deviations:
no
GLP compliance:
yes

Test material

Reference
Name:
Unnamed
Type:
Constituent
Details on test material:
- Name of test material (as cited in study report): Sorbitanmonocaprylat
- Physical state: Liquid, brown
- Analytical purity: >99 % (w/w)
- Purity test date: 2012-06-15
- Lot/batch No.: ESD0006420
- Expiration date of the lot/batch: 2013-09-01
- Stability under test conditions: Not applicable
- Storage condition of test material: Room temperature, protected from light, in original container.

Study design

Oxygen conditions:
aerobic
Inoculum or test system:
activated sludge, non-adapted
Details on inoculum:
- Source of inoculum/activated sludge (e.g. location, sampling depth, contamination history, procedure): Municipal sewage treatment plant, D-31137 Hildesheim, Germany
- Pretreatment/Concentration of sludge: The activated sludge was washed twice with autoclaved tap water. After the second washing the settled sludge was resuspended in mineral salts medium and was maintained in an aerobic condition by aeration for 2 hours. Thereafter the sludge was homogenized with a blender. The supernatant was decanted and maintained in an aerobic condition by aeration until test start.
10 mL/L were used to initiate inoculation.
- Initial cell/biomass concentration:5.8*E8 CFU/L
Duration of test (contact time):
28 d
Initial test substance concentration
Initial conc.:
20 mg/L
Based on:
test mat.
Parameter followed for biodegradation estimation
Parameter followed for biodegradation estimation:
CO2 evolution
Details on study design:
TEST CONDITIONS
- Composition of medium: Mineral salts medium acc. to OECD 301 B / CO2 Evolution Test
- Test temperature: 22 +/- 2°C (20.0 - 23.0 °C, for 4 h down to 19.5 °C
- pH adjusted: no
- Continuous darkness: no, low light conditions (brown glass bottles)



TEST SYSTEM
- Culturing apparatus: 5000 mL brown glass flasks
- Number of culture flasks/concentration: 1 for the reference item, 1 for toxicity control (test and reference item), 2 for the control, 2 for the test item
- Method used to create aerobic conditions: Aeration with 30 - 100 mL/min
- Measuring equipment: Visual check of aeration twice per day
- Details of trap for CO2 and volatile organics if used: CO2 adsorption vessels were connected to the air outlets of the incubation vessels via a series of 3 gas wash bottles, each containing 100 mL of a 0.0125 mol/L Ba(OH)2 solution.
- Other: Application: The necessary amounts of bidistilled water, mineral salts medium and inoculum were placed in each of the incubation vessels. The vessels were aerated for 24 h with CO2 free air. After 24 h the CO2 adsorption vessels were connected to the air outlets of the incubation vessels. Test and reference item were weighed out and were transferred into the incubation vessels with bidistilled water. The vessels were made up to 3 L with bidistilled water and connected to the system for the production of CO2 free air.


SAMPLING
- Sampling frequency: Backtitration of the residual Ba(OH)2 with 0.05 N HCL was carried out three times a week during the first ten days and thereafter twice weekly.
- Sampling method: For each titration the first gas wash bottle was removed and a new bottle was connected to the last one.



CONTROL AND BLANK SYSTEM
- Inoculum blank: Test medium without test and reference item
- Abiotic sterile control: No
- Toxicity control: Test item and reference item in test concentration



STATISTICAL METHODS:
The theoretical production of carbon dioxide (ThCO2) of the test item and functional control is calculated by the carbon content and the molecular formula, respectively.
The produced CO2 was calculated by: 1 mL HCl (c = 0.05 mol/L) = 1.1 mg CO2
The net amount of CO2 produced is calculated by correcting the results of the test item and functional control for endogenous CO2 production of the inoculum controls.
The biodegradation is calculated from the ratio theoretical CO2 production to net CO2 production.
Reference substance
Reference substance:
benzoic acid, sodium salt
Remarks:
20 mg/L

Results and discussion

Preliminary study:
Not performed
% Degradation
Parameter:
% degradation (CO2 evolution)
Value:
88
Sampling time:
28 d
Details on results:
The adaptation phase of the functional control changed after 2 days into the degradation phase (degradation  10 %). The course of the degradation was rapid and the functional control reached the pass level of 60 % within 7 days and a biodegradation of 86 % after 28 days. The validity criterion degradation  60 % after 14 days is fulfilled.
The biodegradation of the test item is shown in comparison to the readily degradable functional control and the toxicity control. The mean 10 % level (beginning of biodegradation) was reached after 4 days and the mean 60 % pass level was reached in the 10-day window. After 28 days a mean biodegradation of 88 % was reached.
In the inoculum control the total CO2 production was 15.1 mg CO2/L after 28 days.

BOD5 / COD results

Results with reference substance:
In the toxicity control containing both test and reference item a biodegradation rate of 68 % was determined within 14 days and it came to 94 % after 28 days. The biodegradation of the reference item was not inhibited by the test item in the toxicity control.

Any other information on results incl. tables

Result tables of the original test report are available on request.

Applicant's summary and conclusion

Validity criteria fulfilled:
yes
Interpretation of results:
readily biodegradable
Conclusions:
The test item fulfills the criteria of the OECD 301B Guideline on readily biodegradable after 28 days and within the 10-d-window.
Executive summary:

The ready biodegradability of the test item Sorbitanmonocaprylat was determined with a non adapted activated sludge over a test period of 28 days in the Modified Sturm Test. The study was conducted from 2009 -02 -19 to 2009 -03 -20 according to OECD 301 B at Dr.U.Noack-Laboratorien. The test item was tested at a concentration of 20 mg/L in duplicates, corresponding to a carbon content (TOC) of 0.578 mgC/mg in the test vessels. The biodegradation of the test item was followed by titrimetric analysis of the quantity of CO2 produced by the respiration of bacteria. The degradation was stopped on day 28 by acidification of the test solutions. The last titration was made on day 29, after residual CO2 had been purged from the test solutions over a period of 24 hours. The percentage CO2production was calculated in relation to the theoretical CO2production (ThCO2) of the test item. The biodegradation was calculated for each titration time.

To check the activity of the test system, sodium benzoate was used as functional control. The percentage degradation of the functional control reached the pass level of 60 % after 7 days and a biodegradation of 78 % after 28 days.

In the toxicity control containing both test and reference item a biodegradation rate of 68 % was determined within 14 days and it came to 94 % after 28 days. The biodegradation of the reference item was not inhibited by the test item in the toxicity control.

The biodegradation of the test item is shown graphically in comparison to the readily degradable functional control and the toxicity control. The mean 10 % level (beginning of biodegradation) was reached after 4 days and the mean 60 % pass level was reached in the 10-day window. After 28 days a mean biodegradation of 63 % was reached.

Biodegradation of the Test Item Sorbitanmonocaprylat in Comparison to the Functional Control and Toxicity Control

Biodegradation [%]

Study Day [d]

6

14

22

28

Test Item, 1st Replicate

35

72

79

86

Test Item, 2nd Replicate

26

65

78

90

Functional Control

58

76

76

78

Toxicity Control
Test Item + Reference Item

29

68

82

94