Benzenamine, 3,3'-[[1,1'-biphenyl]-4,4'-diylbis(oxy)]bis-

Short-term toxicity to aquatic invertebrates

Administrative data

Endpoint:

short-term toxicity to aquatic invertebrates

Type of information:

experimental study

Adequacy of study:

key study

Reliability:

1 (reliable without restriction)

Rationale for reliability incl. deficiencies:

other: see 'Remark'

Remarks:

Study was in compliance with EU Methods for Determination of Ecotoxicity, Annex to Directive 92/69/EEC (O.J. No. L383A, 29.12.92) Part C, Method 2 "Acute toxicity to Daphnia" and the OECD Guideline for Testing of Chemicals No. 202, "Daphnia Acute Immobilisation Test" (2004). No deviations from the protocol were reported.

Data source

Reference

Reference Type:

study report

Title:

Unnamed

Year:

2008

Report date:

2008

Materials and methods

Test guideline

Qualifier:

according to guideline

Guideline:

OECD Guideline 202 (Daphnia sp. Acute Immobilisation Test)

Deviations:

no

GLP compliance:

yes (incl. QA statement)

Remarks:

The Department of Health of the Government of the United Kingdom

Test material

Test material information

Constituent 1

Reference substance name:

3-{[4'-(3-aminophenoxy)-[1,1'-biphenyl]-4-yl]oxy}aniline

EC Number:

700-102-1

Cas Number:

105112-76-3

Molecular formula:

C24H20N2O2

IUPAC Name:

3-{[4'-(3-aminophenoxy)-[1,1'-biphenyl]-4-yl]oxy}aniline

Details on test material:

- Name of test material (as cited in study report): 3,3-(4,4-biphenylenedioxy)dianiline
- Physical state: Off-white powder
- Analytical purity: > 99%
- Lot/batch No.: 20070501
- Expiration date of the lot/batch: 30 April 2008
- Storage condition of test material: Room temperature
- Other: Sample received on 28 August 2007

Sampling and analysis

Analytical monitoring:

yes

Details on sampling:

- Concentrations:
- Sampling method: At the start of the definitive test, four samples (100 mL) were taken from the freshly prepared control and test media. After 48 hours, the contents of the test vessels from each group were pooled and further samples were taken for analysis.
- Sample storage conditions before analysis: Acetonitrile (50 mL) was added to each sample to stabilise the test substance prior to analysis. On each occasion, two of the samples were analysed and the others were stored frozen in case furter analysis was required.

Test solutions

Vehicle:

yes

Details on test solutions:

PREPARATION AND APPLICATION OF TEST SOLUTION (especially for difficult test substances)
- Method: On each occasion, the test substance (20 mg) was dissolved in dimethylformamide (DMF; 10 ml) to provide a primary stock solution at 2 mg/ml. This primary solution was serially diluted, with DMF, to produce two intermediate solutions at 20 and 200 μg/ml. An aliquot (100 μl) of the solvent stock at 20 μg/ml was added to the dilution medium (1 l) in a volumetric flask. The contents of the flask were shaken vigorously before being poured into the test vessels.

- Controls: As an intermediate vehicle was used to facilitate the preparation of the test medium, an additional control group containing dimethylformamide and dilution water (100 μl/l) was included in the study. The solvent control vessel was filled with dilution medium containing the same concentration of auxiliary substance as present in the test concentration. The solvent selected for use in this study was based on information provided by the Sponsor.
- Chemical name of vehicle (organic solvent, emulsifier or dispersant): dimethylformamide
- Evidence of undissolved material (e.g. precipitate, surface film, etc): The test medium was clear and colourless.

Test organisms

Test organisms (species):

Daphnia magna

Details on test organisms:

TEST ORGANISM
- Common name: Daphnia magna
- Strain: Straus
- Source: National Institute for Applied Chemical Research (IRCHA), France
- Age at study initiation (mean and range, SD): less than 24 hours

Study design

Test type:

static

Water media type:

freshwater

Limit test:

yes

Total exposure duration:

48 h

Test conditions

Hardness:

250

Test temperature:

18.8 to 20.1 °C

pH:

Control: 7.42 at start and 7.53 after 48 hours;
Solvent control: 7.55 at start and 7.7 after 48 hours;
with test substance: 7.53 at start and 7.72 after 48 hours

Dissolved oxygen:

% of air saturation value:
Control: 99 at start and 98 after 48 hours
Solvent control: 99 at start and 99 after 48 hours
with test substance: 98 at start and 98 after 48 hours

Nominal and measured concentrations:

The test was conducted at a nominal 2ug/L; this concentration intentionally slightly exceeded the limit of the solubility of the test substance (stated value, 1.7 ug/L). The measured concentrations during the definitive test ranged between 2.71 and 2.83 ug/L. After 48 hours, in samples of expired medium, the measured concentrations had decreased, ranging from below the limit of detection of the analytical method (0.25 ug/L) to less than the limit of quantification (0.5 ug/L). A geometric mean measured concentration of 0.83 ug/L was calculated.

Details on test conditions:

TEST SYSTEM
- Test vessel:
- Material, size, headspace, fill volume: 150 mL capacity crystallising dishes, filled with 100 mL of medium, loosely covered with watch glasses
- Aeration: no supplementary aeration
- Renewal rate of test solution (frequency/flow rate): Daphnia were exposed to the test or control conditions for a period of 48 hours without renewal of test media
- No. of organisms per vessel: five
- No. of vessels per concentration (replicates): 4
- No. of vessels per control (replicates): 4
- No. of vessels per solvent control (replicates): 4


TEST MEDIUM / WATER PARAMETERS
- Source/preparation of dilution water: Deionised, reverse osmosis water
- Total organic carbon:
- Particulate matter:
- Metals:
- Pesticides:
- Chlorine:
- Alkalinity: 50 mg/L as CaCO3
- Ca/mg ratio:
- Conductivity:
- Culture medium different from test medium: Elendt M4 Medium
- Intervals of water quality measurement:


OTHER TEST CONDITIONS
- Adjustment of pH:
- Photoperiod: 16 hours light and 8 hours dark was maintained, subdued lightning at the beginning and end of each light phase
- Light intensity:


EFFECT PARAMETERS MEASURED (with observation intervals if applicable) :
Daphnia were considered to be immobile if they were unable to swin within approximately 15 seconds following gentle agitation of the test vessel. The numbers of mobile, immobile and floating Daphnia were counted approximately 24 and 48 hours after the start of the study.

TEST CONCENTRATIONS
- Spacing factor for test concentrations:
- Justification for using less concentrations than requested by guideline:
- Range finding study
- Test concentrations: The range finding test employed nominal test concentrations of 0.1, 1 and 10 ug/L.
- Results used to determine the conditions for the definitive study: After 48 hours, no immobility or adverse effects on Daphnia were noted at nominal 10 ug/L, where a mean level of 3.86 ug/L was maintained. Based on this result, the definitive (limit) test was conducted at a nominal 2 ug/L

Results and discussion

Effect concentrations

Duration:

48 h

Dose descriptor:

EC50

Effect conc.:

> 0.83 µg/L

Nominal / measured:

meas. (geom. mean)

Conc. based on:

dissolved

Basis for effect:

mobility

Details on results:

- Any observations (e.g. precipitation) that might cause a difference between measured and nominal values: During the definitive test, the measured concentrations of 3,3(4,4-biphenylenedioxy)dianiline ranged between 2.71 and 2.83 μg/l in samples of the freshly prepared medium; 136 and 142% of the nominal value. After 48 hours, in samples of expired medium, the measured concentration had decreased, ranging from below the limit of detection of the analytical method (0.25 μg/l) to less than the limit of quantification (0.5 μg/l). These results indicate that 3,3-(4,4-biphenylenedioxy)dianiline was unstable under the conditions of the test. Based on a geometric mean, an overall mean measured concentration of 0.83 μg/l was calculated (see Table 1) and this value has been used in test calculations.

Any other information on results incl. tables

Table 1: Measured concentrations (microgram/L)

Nominal exposure concentration (microgram/L)	Measured 3,3'-[biphenyl-4,4'-diylbis(oxy)]dianiline concentration (microgram/L)					Overall geometric mean
	0 hours	%N	48 hours	%N	%ti
Control	nd, nd	-	nd, nd	-	-	nd
Solvent control	nd, nd	-	nd, nd	-	-	nd
2*	2.71, 2.83	136, 142	0.25**, 0.25**	13	9	0.83

nd : none detected (0.25 microgram/L)

%N: mean measured concentration expressed as a percentage of the nominal concentration

%ti: mean measured concentration after 48 hours expressed as percentage of the mean starting concentration

*: as 3,3-(4,4-biphenylenedioxy)dianiline

**: for calculation of the test results, the limit of detection (0.25 microgram/L) has been used as the mean measured concentration present at 72 hours

Table 2: Environmental parameters

Nominal exposure concentration (microgram/L)		Temperature (°C)		pH		Dissolved oxygen (%ASV)
Nominal	Measured	0 h	48 h	0 h	48 h	0 h	48 h
Control	nd	20.4	19.2	7.42	7.53	99	98
Solvent control	nd	20.2	19.1	7.55	7.70	99	99
2*	0.83**	20.1	18.8	7.53	7.72	98	98

nd : none detected ( 0.25 microgram/L)

ASV: air saturation value

*: 3,3-(4,4-biphenylenedioxy)dianiline

**: geometric mean

The total hardness and alkalinity of the batch of Elendt M4 medium used were 250 and 50 mg/L as CaCO, respectively.

Continuous monitoring of an additional vessel containing dilution medium = 19.4 to 20.4 °C.

Applicant's summary and conclusion

Validity criteria fulfilled:

yes

Conclusions:

Under the conditions of the study, 3,3-(4,4-biphenylenedioxy)dianiline was not found to be acutely toxic to Daphnia magna when tested at a starting concentration (2.77 ug/L) that exceeded the limit of aqueous solubility of the test substance (1.7 ug/L), or in a range finding test where Daphnia were exposed to a mean measured level of 3.86 ug/L.

Consequently, the 48-hour EC50 value for 3,3-(4,4-biphenylenedioxy)dianiline with Daphnia magna could not be calculated but must be >0.83 ug/L, and the "no observed effect concentration" was 0.83 ug/L.

Executive summary:

The acute toxicity of 3,3'-[biphenyl-4,4'-diylbis(oxy)]dianiline to Daphnia magna was assessed under static exposure conditions (Wilby et al. 2008b). The study was conducted in accordance with EU Methods for Determination of Ecotoxicity, Annex to Directive 92/69/EEC, Part C, Method 2 “Acute toxicity to Daphnia” and the OECD Guideline for Testing of Chemicals No. 202, “Daphnia Acute Immobilisation Test” (2004).

A group of twenty Daphnia, less than 24 hours old, was exposed for 48 hours to 3,3'-[biphenyl-4,4'-diylbis(oxy)]dianiline at a nominal concentration of 2 μg/L, which was slightly in excess of the limit of solubility of the test substance (stated value, 1.7 μg/L) to ensure that the Daphnia were exposed to the maximum attainable concentration. To aid dissolution, the test substance was dissolved in an auxiliary solvent before its addition to the dilution medium (Elendt M4). To minimise losses by adsorption, test vessels were pre-exposed to the test substance before use in the definitive test. Measured concentrations of 3,3'-[biphenyl-4,4'-diylbis(oxy)]dianiline ranged between 2.71 and 2.83 μg/L in samples of freshly prepared media and decreased to less than 0.5 μg/L in samples of expired (48-hour-old) media, giving an overall geometric mean measured level of 0.83 μg/L. These results indicate that the test substance was not stable under the conditions of the test. Observations of the Daphnia in each control and test vessel were made after 24 and 48 hours. No immobilisation or adverse effects on the Daphnia were noted at a mean measured concentration of 0.83 μg/L. Based on these findings, the following values have been estimated:

48-hour EC50 value: >0.83 μg/L

NOEC: ≥0.83 μg/L

The NOEC must be greater than 0.83 μg/L. Under the conditions of the study, 3,3'-[biphenyl-4,4'-diylbis(oxy)]dianiline was not found to be acutely toxic to Daphnia magna when tested at a starting concentration (2.77 μg/L) that exceeded the limit of aqueous solubility of the test substance (1.7 μg/L), or in a range finding test where Daphnia were exposed to a mean measured level of 3.86 μg/L.