2-nitrophenyl acetate

Biodegradation in water: screening tests

Administrative data

Endpoint:

biodegradation in water: ready biodegradability

Type of information:

experimental study

Adequacy of study:

key study

Reliability:

1 (reliable without restriction)

Data source

Reference

Reference Type:

study report

Title:

Unnamed

Year:

2015

Report date:

2015

Materials and methods

Test guideline

Qualifier:

according to guideline

Guideline:

EU Method C.4-E (Determination of the "Ready" Biodegradability - Closed Bottle Test)

Deviations:

no

GLP compliance:

yes (incl. QA statement)

Test material

Test material information

Constituent 1

Reference substance name:

Acetic acid, 2-nitrophenyl ester

IUPAC Name:

Acetic acid, 2-nitrophenyl ester

Constituent 2

Reference substance name:

2-nitrophenyl acetate

EC Number:

210-233-1

EC Name:

2-nitrophenyl acetate

Cas Number:

610-69-5

Molecular formula:

C8H7NO4

IUPAC Name:

2-nitrophenyl acetate

Test material form:

solid: crystalline

Details on test material:

- Name of test material (as cited in study report): 2-Nitrophenyl Acetate

- Physical state: solid light yellow
- Analytical purity: 95 % w/w

- Impurities (identity and concentrations):
Phenol <5.0 % (w/w)
2,4-dinitrophenol <5.0 % (w/w)
2,6-dinitrophenol <5.0 % (w/w)
Acetic acid <1.0 % (w/w)
Water <5.0 % (w/w)
2-nitrophenol <5.0 % (w/w)

- Lot/batch No.: 201408
- Expiration date of the lot/batch: unlisted

- Storage condition of test material: The test substance will be stored in dry room in dark in
closed container at the room temperature

Study design

Oxygen conditions:

aerobic

Inoculum or test system:

sewage, predominantly domestic (adaptation not specified)

Details on inoculum:

- Preparation of inoculum for exposure: The fresh collected waste water was filtered through paper filter. COD was determined and waste water was aerated till using.

Initial test substance concentration

Initial conc.:

3.4 mg/L

Based on:

COD

Parameter followed for biodegradation estimation

Parameter followed for biodegradation estimation:

O2 consumption

Details on study design:

TEST CONDITIONS
- Composition of medium:
Inoculated medium
The mineral medium was strongly aerated for 20 minute and let standing for 20 hours at the test temperature. The volume of waste water for the inoculation of mineral medium was chosen 2.1 mL per 1 L of medium (COD of waste water: 61 mg·L-1).
30 L aerated mineral medium was prepared for the test and before the beginning of the test it was inoculated with 63 mL of modified waste water. The pH value of medium: 7.4.
Test substance
The stock solution of the test substance was prepared in concentration 0.1007 g·L-1 of deionized water.
From this solution 120 mL (20 mL·L-1) were transferred to a large glass bottle and completed with the inoculated medium to the volume of 6 L.
The solution prepared in this way contained 2.01 mg·L-1 of the test substance, and had COD 3.40 mg·L-1. The pH value of solution: 7.4.
Reference substance
The stock solution of sodium benzoate was prepared in concentration 1.0000 g·L-1of water. From this solution the 12.6 mL (2.1 mL·L-1) were transferred to a large glass bottle and completed with the inoculated medium to the volume of 6 L.
The solution prepared in this way contained 2.10 mg·L-1 of sodium benzoate, and had COD 3.45 mg·L-1. The pH value of solution: 7.4.
Toxicity test
The solution was prepared by dosing 80 mL of stock solution of test substance and 8.4 mL of stock solution of reference substance into inoculated medium and completed with this medium to the volume of 4 L.
The mixture prepared in this way contained 2.01 mg·L-1 of test substance and 2.10 mg·L-1 of sodium benzoate and had COD 6.85 mg·L-1.
Control (blank) determination
The inoculated mineral medium without test substance was used for the blank determination.


- Test temperature: 20 +- 1.0°C

- Aeration of dilution water: yes

- Continuous darkness: yes

TEST SYSTEM
- Culturing apparatus:
- Number of culture flasks/concentration:
- 2 x 9 bottles with test substance and inoculated medium
- 2 x 9 bottles with reference substance and inoculated medium for the check of inoculum activity
- 9 bottles with test substance and inoculated medium for the determination of oxidized nitrogen forms
- 2 x 5 bottles with test and reference substance mixture and inoculated medium for the toxicity test
- 2 x 9 bottles with inoculated medium only for the blank determination


- Measuring equipment:
pH meter WTW Inolab pH 730
- analytical balance XS 105 DU (Mettler Toledo)
- UV/VIS spectrophotometer Helios Alpha
- equipment for the determination of nitrite
- equipment for the determination of nitrate
- volumetric flasks, flasks, beakers, pipettes and further common laboratory equipment

SAMPLING
- Sampling frequency: At the zero-time and at the 3rd, 7th, 10th, 14th, 17th, 21st, 24th and 28th day of test the appropriate bottles from each series were taken off

Reference substance

Reference substance:

benzoic acid, sodium salt

Results and discussion

% Degradation

Parameter:

% degradation (O2 consumption)

Value:

70.4

Sampling time:

28 d

BOD5 / COD results

BOD5 / COD

BOD5 / COD 1

Parameter:

BOD5

Value:

0.248 other: mg/mg

BOD5 / COD 2

Parameter:

COD

Value:

1.688 other: mg/mg

Results with reference substance:

COD of the reference substance: 1.641 mg.mg-1

Any other information on results incl. tables

The oxygen consumption and percentage of the biodegradation of the test substance for each test interval and each bottle were calculated.

In time intervals when the increase of oxidized nitrogen forms was detected, the correction of oxygen consumption for nitrification was carried out.

The evaluation of the reference substance and of the test and reference substance mixture was done by the same way.

All the calculations were related to the initial values of COD of the test and reference substance.

Formulae for the calculations are given in tables 4 and 5.

The results of the test were plotted into a graph from which the degree of degradability in percentage in each time interval can be deduced.

Applicant's summary and conclusion

Validity criteria fulfilled:

yes

Conclusions:

In this 28-day study of ready biodegradability a degradation of 32.6 % of the test substance, 2-Nitrophenyl Acetate , was attained in the end of study.

Executive summary:

The test substance, 2-Nitrophenyl Acetate, 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.

The results of biological degradation are related to experimentally determined COD values of the test and reference substance at the beginning of the test.

The test substance was sufficiently soluble in used mineral medium so the dosage from the stock solution was carried out.

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

COD of the test substance in medium at the beginning of the main test: 3.40 mg·L-1

COD of reference substance in medium at the beginning of the main test: 3.45 mg·L-1

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

Based on the test substance contains nitrogen in the chemical composition, the oxidized nitrogen forms were determined and the correction for nitrification was carried out.

The test was performed at temperature of 20 ± 1 °C with the pH values of solutions 7-8 at the beginning of the test.

Validity of the test

The prescribed validity criteria in the test were fulfilled.

The test substance 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 32.6 % of the test substance, 2-Nitrophenyl Acetate, was attained in the end of study.