lithium hydrogen 7-{(E)-[2-amino-4-(2-naphthyl)-1,3-thiazol-5-yl]diazenyl}naphthalene-1,3,5-trisulfonate (2:1:1)

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:

Test phase 2014-03-06 to 2014-05-23. Report completed 2014-06-19

Reliability:

1 (reliable without restriction)

Rationale for reliability incl. deficiencies:

other: Meets the criteria for classification as reliable without restriction according to Klimisch et al (1997).

Data source

Reference

Reference Type:

study report

Title:

Unnamed

Year:

2014

Report date:

2014

Materials and methods

Test guideline

Qualifier:

according to guideline

Guideline:

OECD Guideline 301 F (Ready Biodegradability: Manometric Respirometry Test)

Deviations:

no

GLP compliance:

yes (incl. QA statement)

Test material

Test material information

Constituent 1

Reference substance name:

lithium hydrogen 7-{(E)-[2-amino-4-(2-naphthyl)-1,3-thiazol-5-yl]diazenyl}naphthalene-1,3,5-trisulfonate (2:1:1)

EC Number:

940-433-0

Molecular formula:

C23H14Li2N4O9S4

IUPAC Name:

lithium hydrogen 7-{(E)-[2-amino-4-(2-naphthyl)-1,3-thiazol-5-yl]diazenyl}naphthalene-1,3,5-trisulfonate (2:1:1)

Test material form:

solid: particulate/powder

Remarks:

migrated information: powder

Study design

Oxygen conditions:

aerobic

Inoculum or test system:

sewage, predominantly domestic, non-adapted

Details on inoculum:

A mixed population of sewage treatment micro-organisms was obtained on 23 April 2014 from the final effluent stage of the Severn Trent Water Plc sewage treatment plant at Loughborough, Leicestershire, UK, which treats predominantly domestic sewage.

The sample of effluent was filtered through coarse filter paper (first approximate 200 mL discarded) and maintained on aeration in a temperature controlled room at 21 ± 1 ºC prior to use.

Duration of test (contact time):

28 d

Initial test substance concentration

Initial conc.:

100 mg/L

Based on:

test mat.

Parameter followed for biodegradation estimation

Parameter followed for biodegradation estimation:

O2 consumption

Details on study design:

Test Item
The test item was dissolved directly in mineral medium. An amount of test item (500 mg) was dissolved in mineral medium with the aid of ultrasonication for approximately 5 minutes and the volume adjusted to 500 mL to give a 1000 mg/L stock solution. An aliquot (50 mL) of this stock solution was diluted with mineral medium (445 mL) and inoculum (5 mL) to give the final test concentration of 100 mg/L. The volumetric flasks containing the stock solution and the test concentration were inverted several times to ensure homogeneity.

Reference Item
A reference item, aniline (C6H5NH2), was used to prepare the procedure control vessels. An initial stock solution of 1000 mg/L was prepared by dissolving the reference item directly in mineral medium with the aid of ultrasonication for approximately 5 minutes. An aliquot (50 mL) of this stock solution was diluted with mineral medium (445 mL) and the inoculum (5 mL), to give the test concentration of 100 mg/L. The volumetric flask containing the stock solution was inverted several times to ensure homogeneity.

Toxicity Control
A toxicity control, containing the test item and aniline, was prepared in order to assess any toxic effect of the test item on the sewage treatment micro-organisms used in the test. An aliquot (50 mL) of the 1000 mg/L test item stock solution and an aliquot (50 mL) of the 1000 mg/L aniline stock solution was diluted with mineral medium (395 mL) and inoculum (5 mL) to give the test concentration of 100 mg test item/L and 100 mg aniline/L.

Preparation of Test System
The following test preparations were prepared and inoculated in 500 mL bottles:
a) Three replicate bottles containing inoculated mineral medium to act as the inoculum control.
b) Two replicate bottles containing inoculated mineral medium and the reference item, aniline, at a concentration of 100 mg/L.
c) Three replicate bottles containing inoculated mineral medium and the test item at a concentration of 100 mg/L.
d) Two replicate bottles containing inoculated mineral medium, the reference item, aniline, at a concentration of 100 mg/L and the test item at a concentration of 100 mg/L to act as toxicity control vessels.
Data from the inoculum control and procedure control vessels was shared with similar concurrent studies.

All vessels were inoculated with the prepared inoculum at a rate of 1% v/v.

On Day 0 the test and reference items were added and the pH of all vessels measured using a Hach HQ40d Flexi handheld meter. The pH values were adjusted to pH 7.4 ± 0.2 using diluted hydrochloric acid solution prior to the addition of the inoculum and the volume in all the vessels being adjusted to 500 mL by the addition of mineral medium.

All inoculum control, test item, procedure control and toxicity control vessels were placed in a CES Multi-Channel Aerobic Respirometer.

The system consists of a sample flask sealed by a sensor head/CO2 trap immersed in a temperature controlled water bath. The samples were stirred for the duration of the test with a magnetically coupled stirrer.

As biodegradation progresses, the micro-organisms convert oxygen to carbon dioxide which is absorbed into the ethanolamine solution (50% v/v) causing a net reduction in gas pressure within the sample flask. The pressure reduction triggers the electrolytic process, generating oxygen and restoring the pressure in the sample flask. The magnitude of the electrolyzing current and the duration of the current is proportional to the amount of oxygen supplied to the micro-organisms. The data generated from the respirometer’s own battery backed memory was collected on the hard disk drive of a non-dedicated computer.

The test was conducted in diffuse light at a temperature of 23 ± 1 ºC.

On Day 28, one inoculum control, one procedure control, two test item and one toxicity control vessel were sampled for pH.

The remaining vessels which were not sampled were discarded and are not reported. Additional replicate vessels were prepared and incubated in order that in the event of a leak in the test system a replicate vessel could be discarded without jeopardizing the integrity of the test.

Reference substance

Reference substance:

aniline

Results and discussion

% Degradation

% Degradation 1

Parameter:

% degradation (O2 consumption)

Value:

2

Sampling time:

5 d

% Degradation 2

Parameter:

% degradation (O2 consumption)

Value:

6

Sampling time:

10 d

% Degradation 3

Parameter:

% degradation (O2 consumption)

Value:

4

Sampling time:

15 d

% Degradation 4

Parameter:

% degradation (O2 consumption)

Value:

4

Sampling time:

20 d

% Degradation 5

Parameter:

% degradation (O2 consumption)

Value:

5

Sampling time:

28 d

Details on results:

The mean BOD of the inoculated mineral medium (control) was 36.32 mg O2/L after 28 days and therefore satisfied the validation criterion given in the OECD Test Guidelines.

The pH of the inoculated test item vessels on Day 28 ranged from 7.5 to 7.7 and hence satisfied the validation criterion given in the OECD Test Guidelines.

The difference between extremes of replicate BOD values at the end of the test was less than 20% and therefore satisfied the validation criterion given in the OECD Test Guidelines.

Biodegradation
The test item attained 5% biodegradation after 28 days, calculated from the oxygen consumption values, and therefore cannot be considered to be readily biodegradable under the strict terms and conditions of OECD Guideline No. 301F.

The toxicity control attained 48% biodegradation after 14 days and 49% biodegradation after 28 days thereby confirming that the test item was not toxic to the sewage treatment micro-organisms used in the test.

Aniline (procedure control) attained 74% biodegradation after 14 days and 78% biodegradation after 28 days thereby confirming the suitability of the inoculum and test conditions.

BOD5 / COD results

Results with reference substance:

Aniline (procedure control) attained 74% biodegradation after 14 days and 78% biodegradation after 28 days thereby confirming the suitability of the inoculum and test conditions.

Applicant's summary and conclusion

Validity criteria fulfilled:

yes

Interpretation of results:

other: Not readily biodegradable

Conclusions:

The test item attained 5% biodegradation after 28 days and therefore cannot be considered as readily biodegradable under the strict terms and conditions of OECD Guideline No. 301F.

Executive summary:

Introduction

The study was performed to assess the ready biodegradability of the test item in an aerobic aqueous media. The method followed was designed to be compatible with the OECD Guidelines for Testing of Chemicals (1992) No. 301F, “Ready Biodegradability; Manometric Respirometry Test” referenced as method C.4-D of Commission Regulation (EC) No. 440/2008 and US EPA Fate, Transport, and Transformation Test Guidelines OCSPP 835.3110 (Paragraph (q)).

 

 Methods…….

The test item at a concentration of 100 mg/L was exposed to sewage treatment micro-organisms with mineral medium in sealed culture vessels in diffuse light at approximately 23 ºC for 28 days. 

 

The biodegradation of the test item was assessed by the measurement of daily oxygen consumption values on Days 0 to 28. Control solutions with inoculum and the reference item, aniline, together with a toxicity control were used for validation purposes.

  

Results…….

The test item attained 5% biodegradation after 28 days and therefore cannot be considered to be readily biodegradable under the strict terms and conditions of OECD Guideline No. 301F.