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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:
The study will be available 30/06/2018
Reliability:
1 (reliable without restriction)
Rationale for reliability incl. deficiencies:
guideline study

Data source

Reference
Reference Type:
study report
Title:
Unnamed
Year:
2018
Report Date:
2018

Materials and methods

Test guidelineopen allclose all
Qualifier:
according to
Guideline:
EU Method C.4-E (Determination of the "Ready" Biodegradability - Closed Bottle Test)
Deviations:
no
Qualifier:
according to
Guideline:
OECD Guideline 301 D (Ready Biodegradability: Closed Bottle Test)
Deviations:
no
Principles of method if other than guideline:
The solution of the test item in mineral medium is inoculated with a relatively small number of microorganisms from i mixed population and kept in completely full closed bottles in the dark at constant temperature.
Degradation is determined by analysis of dissolved oxygen over a 28-day period.
The following determinations were done simultaneously under the same conditions:
- check of inoculum activity with the reference item
- toxicity test of the test item for used inoculum in mixtrue of the test and reference item
- determination of ixygen consumption for nitrification
- determination of blank (inoculated mineral medium).
The amount of oxygen taken up by the test chemical, corrected for uptake by the blank inoculum run i parallel, is expressed as a percentage of ThOD.
ThOD (Theoretical oxygen demand (mg) is the total amount of oxygen required to oxidise a chemical completely; it is calculated from the molecular formula and it is also expressed as mg oxygen required per mg test compound.
GLP compliance:
yes

Test material

Reference
Name:
Unnamed
Type:
Constituent
Test material form:
liquid

Study design

Oxygen conditions:
aerobic
Inoculum or test system:
sewage, predominantly domestic (adaptation not specified)
Details on inoculum:
THE PREPARATION OF INOCULUM
The inoculum contained a mixed population of microorganisms obtained from the secondary effluent of the waste water treatment plant of Pardubice processing predominantly minucipal sewage.
The fresh collected waste water was filtered through paper filter and aerated till using.
The inoculum preparation is in conformity with the recommendations of the test guideline.
The volume of waste water for the inoculation of mineral medium was chosen 1.8 mL per 1 L of medium. 30 L aerated mineral medium was prepared for the test and before the beginning of the test it was inoculated with 54 mL of modified waste water. The pH value of medium: 7.4
Duration of test (contact time):
ca. 28 d
Initial test substance concentration
Initial conc.:
ca. 2.91 - 3.05 mg/L
Based on:
ThOD
Remarks:
More information - Table 1 (Concentration of the test item in bottles)
Parameter followed for biodegradation estimationopen allclose all
Parameter followed for biodegradation estimation:
O2 consumption
Remarks:
for nitrification
Parameter followed for biodegradation estimation:
other: toxicity test of the test item for used inoculum in mixture of the test and reference item
Parameter followed for biodegradation estimation:
other: check of inoculum activity with the reference item
Parameter followed for biodegradation estimation:
other: determination of blank (inoculated mineral medium)
Details on study design:
TEST ITEM: The test item was weighed directly into bottle and the bottles were completed to the whole volume with inoculated medium. Mixtures prepared in this way contained 2.91 - 3.05 mg.L-1 of the test item.
REFERENCE ITEM: The stock solution of sodium benzoate was prepared in concentration 1.0000 g.L-1 of deionized water. From this solution the 12.6 mL (2.1 mL.L-1) were transfered to a large glass bottle and completed with the inoculated medium to the volume of 6 L. The solution preparad in this wqy contained 2.10 mg.L-1 of sodium benzoate. The pH value of solution: 7.5
TOXICITY TEST: The test item was weighed directl into bottle and the bottles were completed to the whole volume with inoculataed solution of sodium benzene. The mixture prepared in this way contained 2.91 - 3.06 mg.L-1 of the test item (Table 2) and 2.10 mg.L-1 of sodium benzoate.

TEST CONDITIONS Exposition time: 28 days
Temperature: 20 ± 1 °C
Illumination: no illumination, in the dark

TEST DESIGN
The number of bottles for individual series:
Test item series: 2 x 9 bottles with test item and inoculum
Reference item series: 2 x 9 bottles with reference item and inoculum
Blank series: 2 x 9 bottles with inoculated medium only
Series for nitrification determination: 1 x 9 bottles with test item and inoculum
Toxicity test series: 2 x 5 bottles with test item, reference item and inoculum
The prepared dosing solutions for each series were filled in two parallel bottle replicates (except the series for nitrification determination) . The bottles were placed into thermostat.
MEASUREMENTS: The first two bottles from each series were immediately submitted to analysis for dissolved oxygen (zero-time) and other bottles are placed in the thermostat.
On the 3rd, 7th, 10th, 14th, 17th, 21st, 24th and 28th day of the test, two bottles (parallel replicate) from the test item series, from the reference item series, from the blank series and one bottle from series for nitrification determination were taken off and the determinations were performed.
On the 3rd, 7th, 10th, 14th day of the test, two bottles from the toxicity test series were taken off and the determinations was performed.
The analysis of dissolved oxygen was performed electrochemically using oximeter dievice (oximeter WTW OXI 730).
On the 3rd, 7th, 10th, 14th, 17th, 21st, 24th and 28th day of the test, one bottle from the series for nitrification determination was taken off and stored in freezer. The determinations of nitrite-N concetration and nitrate-N concentration were carried out by ion chromatography at one time at the end of the test. These analyses were performed as GLP work.

Reference substance
Reference substance:
benzoic acid, sodium salt

Results and discussion

Test performance:
MEASUREMENTS: The first two bottles from each series were immediately submitted to analysis for dissolved oxygen (zero-time) and other bottles are placed in the thermostat.
On the 3rd, 7th, 10th, 14th, 17th, 21st, 24th and 28th day of the test, two bottles (parallel replicate) from the test item series, from the reference item series, from the blank series and one bottle from series for nitrification determination were taken off and the determinations were performed.
On the 3rd, 7th, 10th, 14th day of the test, two bottles from the toxicity test series were taken off and the determinations was performed.
The analysis of dissolved oxygen was performed electrochemically using oximeter dievice (oximeter WTW OXI 730).
On the 3rd, 7th, 10th, 14th, 17th, 21st, 24th and 28th day of the test, one bottle from the series for nitrification determination was taken off and stored in freezer. The determinations of nitrite-N concetration and nitrate-N concentration were carried out by ion chromatography at one time at the end of the test. These analyses were performed as GLP work.
% Degradation
Key result
Parameter:
% degradation (DOC removal)
Value:
ca. 33.8
Sampling time:
28 d
Remarks on result:
readily biodegradable based on QSAR/QSPR prediction
Details on results:
According to paragraph 10 of OECD TG 301 the pass level for ready biodegradability is 60% of ThOD values for respirometric methods.

BOD5 / COD results

BOD5 / COD
Parameter:
ThOD
Value:
ca. 2.185 other: mg.mg-1

Applicant's summary and conclusion

Validity criteria fulfilled:
yes
Interpretation of results:
readily biodegradable
Conclusions:
In this 28-day study of ready biological degradability the degradation of 33.8 % of the test
item, Pentyl nitrite, was attained in the end of study.
Executive summary:

The test item , Pentyl nitrite, was tested for the ready biological degradability in Closed Bottle

Test.

Test performance

The test was performed according to:

Method C.4E - Closed Bottle Test, Council Regulation (EC) No. 440/2008, published

in O.J .L 142, 2008

OECD Test Guideline No. 301 D - Closed Bottle Test. Adopted July 17, 1992

The test item is non-soluble in the mineral medium. For these reasons the direct dosing of the

test item into each bottle was implemented.

Sodium benzoate was used as the reference item. The dosage was carried out from the stock

solution.

In parallel to the main test the toxicity test was performed.

Based on the test item contains nitrogen in the chemical composition , the oxidized nitrogen

forms were dete1mined and the correction for nitrification was carried out.

The test was performed at temperature of 20 ± I °C with the pH values of solutions 7 - 8

at the beginning of the test.

The calculations of biological degradation are based on ThOD (theoretical oxygen demand)

values of the test and reference item.

Validity of the test

The prescribed validity criteria in the test were fulfilled.

The test item was not inhibiting for the used inoculum.

Since all criteria of acceptability were met, this study is considered to be valid.

Test results

In this 28-day study of ready biological degradability the degradation of 33.8 % of the test

item, Pentyl nitrite, was attained in the end of study.