disodium 4,4'-bis[(prop-2-enoyl)amino][1,1'-biphenyl]-2,2'-disulfonate

Toxicity to aquatic algae and cyanobacteria

Administrative data

Endpoint:

toxicity to aquatic algae and cyanobacteria

Type of information:

experimental study

Adequacy of study:

key study

Study period:

27 Feb - 02 Mar 2018

Reliability:

2 (reliable with restrictions)

Rationale for reliability incl. deficiencies:

guideline study with acceptable restrictions

Data source

Reference

Reference Type:

study report

Title:

Unnamed

Year:

2018

Report date:

2018

Materials and methods

Test guideline

Qualifier:

equivalent or similar to guideline

Guideline:

OECD Guideline 201 (Alga, Growth Inhibition Test)

Version / remarks:

2006

Deviations:

yes

Remarks:

Temperature during test was 20.1–23.6 °C (OECD 201: 21-24 °C ± 2°C), pre-culture preparation: 5 days before test start (OECD 201: 2-4 days before stest start). As exponential growth was observed in the blank control, this was stated as uncritical.

GLP compliance:

yes (incl. QA statement)

Remarks:

Landesamt für Umwelt, Rheinland-Pfalz, Mainz, Germany

Test material

Test material information

Constituent 1

Reference substance name:

disodium 4-[(1-hydroxyprop-2-en-1-yl)amino]-4'-(prop-2-enamido)-[1,1'-biphenyl]-2,2'-disulfonate

EC Number:

812-549-0

Cas Number:

881539-89-5

Molecular formula:

C18H14N2Na2O8S2

IUPAC Name:

disodium 4-[(1-hydroxyprop-2-en-1-yl)amino]-4'-(prop-2-enamido)-[1,1'-biphenyl]-2,2'-disulfonate

Sampling and analysis

Analytical monitoring:

yes

Details on sampling:

- Concentrations: 0 (control), 10, 22, 46, 100 and 200 mg/L (0, 72 h)

Test solutions

Vehicle:

no

Details on test solutions:

PREPARATION AND APPLICATION OF TEST SOLUTION (especially for difficult test substances)
- Method: stock solution containing 220.0 mg/L in algal medium, lower treatments were prepared by dilution of this stock solution with algal medium
- Controls: nutrient medium only mixed with algal pre-culture

Test organisms

Test organisms (species):

Desmodesmus subspicatus (previous name: Scenedesmus subspicatus)

Details on test organisms:

TEST ORGANISM
- Common name: green alga
- SAG Strain Number: 86.81
- Source (laboratory, culture collection): obtained in January 2016 by MBM Sciencebridge GmbH (Institut für Pflanzenphysiologie of Universität Göttingen, Germany)
- Age of inoculum (at test initiation): Five days before the start of each test, an aliquot of the permanent culture was brought into nutrient medium and incubated under continuous lighting for 96 hours (5000 lux, 20.4 – 23.1°C)
- Method of cultivation: Kept as stock culture on solid agar at 2 - 8°C. From the stock culture, a permanent culture was prepared. From an aliquot of the permanent culture, the pre-culture was prepared.

Study design

Test type:

static

Water media type:

freshwater

Limit test:

no

Total exposure duration:

72 h

Test conditions

Test temperature:

20.1 - 23.6°C

pH:

7.4 - 7.8

Nominal and measured concentrations:

Nominal concentrations: 0 (control), 10, 22, 46, 100 and 220 mg/L
Measured concentrations initial (t-=0) /test end (t-72h):

Details on test conditions:

TEST SYSTEM
- Test vessel: glass flasks, total volume 65 mL
- Type: open: covered with perforated plastic foil acting as a stopper
- Fill volume: 45 ± 1 mL
- Aeration: no
- Initial cells density: 2.3*10^3 cells/mL
- Control end cells density: 157.623*10^3
- No. of vessels per concentration (replicates): 3
- No. of vessels per control (replicates): 6

GROWTH MEDIUM
- Standard medium used: yes: OECD TG 201 medium

TEST MEDIUM / WATER PARAMETERS
- Intervals of water quality measurement: pH (0, 72 h)

OTHER TEST CONDITIONS
- Sterile test conditions: yes
- Adjustment of pH: no
- Photoperiod: continuous light
- Light intensity and quality: 5000 lux

EFFECT PARAMETERS MEASURED (with observation intervals if applicable) : cell number (0, 24, 48, 72 h), appearance of the algae (72 h)
- Determination of cell concentrations: electronic particle counter (Cell Counter Beckman Coulter Z2)

TEST CONCENTRATIONS
- Spacing factor for test concentrations: 2.09 - 2.2

Reference substance (positive control):

yes

Remarks:

Potassium dichromate (K2Cr2O7)

Results and discussion

Effect concentrations

Effect concentrations 1

Duration:

72 h

Dose descriptor:

NOEC

Effect conc.:

100 mg/L

Nominal / measured:

nominal

Conc. based on:

test mat.

Basis for effect:

growth rate

Remarks:

yield

Effect concentrations 2

Duration:

72 h

Dose descriptor:

EC50

Effect conc.:

> 220 mg/L

Nominal / measured:

nominal

Conc. based on:

test mat.

Basis for effect:

growth rate

Remarks:

yield

Details on results:

- Exponential growth in the control (for algal test): yes
- Observation of abnormalities (for algal test): normal and healthy shape in all tested concentration

Results with reference substance (positive control):

- 72h ErC50: 0.90 mg/L (95% confidence limits: 0.77 – 1.1 mg/L)
- Reference substance results are within the recommended range of 1.19 mg/L ± 0.27 mg/L (ISO guideline 8692)

Reported statistics and error estimates:

The estimation of the biological data was accomplished using the software ToxRat® Professional, version 3.2.1.

Any other information on results incl. tables

Table 1: Cell number/mL

 

Nominal Concentration in mg/L	Parameter	Cell Number/mL
		0 h	24 h	48 h	72 h
Blank control	Mean	2340	8747	36457	157623
Blank control	SD	0	884	2310	11232
10	Mean	2340	8587	35660	155993
10	SD	0	147	1652	15427
22	Mean	2340	8647	35913	156287
22	SD	0	162	1508	18212
46	Mean	2340	8780	34520	146667
46	SD	0	473	2589	23275
100	Mean	2340	9000	35253	145593
100	SD	0	714	1798	6972
220	Mean	2340	9320	34700	134393
220	SD	0	278	322	4302

Table 2: Daily growth rates of the blank controls

 

Growth rates	days 0 – 1	days 1 – 2	days 2 – 3	CV of sectional daily growth rates	days 0 – 3
Replicate 1	1.280	1.338	1.518	9%	1.379
Replicate 2	1.340	1.427	1.532	7%	1.433
Replicate 3	1.178	1.634	1.465	16%	1.426
Replicate 4	1.354	1.396	1.482	5%	1.411
Replicate 5	1.472	1.297	1.387	6%	1.385
Replicate 6	1.261	1.488	1.399	8%	1.383
Mean	1.314	1.430	1.464	9%	1.403
Standard deviation	0.100	0.120	0.060		0.024
CV	8%	8%	4%		2%

Table 3: Measured concentration and % of nominal

 

Nominal Concentration Test Item	Measured concentration t = 0 h	Measured concentration t = 72 h	% of Nominal concentration t = 0 h	% of Nominal concentration t = 72 h
mg/L	mg/L	mg/L	mg/L	mg/L
Blank control	< LOQ	<LOQ	--	--
10	8.27	9.64	82.74	96.39
22	20.85	22.35	94.77	101.58
46	43.01	45.16	93.50	98.17
100	96.00	98.24	96.00	98.24
220	204.48	216.84	92.95	98.56

LOQ (Limit of quantification) = 4 mg/L

The recovery of the QC sample of 93.7% (0h) and 96.5% (72h) was taken into account

Table 4: Validity criteria for OECD 201.

Criterion from the guideline	Outcome	Validity criterion fulfilled
The biomass in the control cultures should have increased exponentially by a factor of at least 16 within the 72-hour test period.	67	yes
The mean coefficient of variation for section-by-section specific growth rates (days 0-1, 1-2 and 2-3, for 72-hour tests) in the control cultures must not exceed 35%	9%	yes
The coefficient of variation of average specific growth rates during the whole test period in replicate control cultures must not exceed 7% in tests with Pseudokirchneriella subcapitata and Desmodesmus subspicatus. For other less frequently tested species, the value should not exceed 10%.	2%	yes

Applicant's summary and conclusion

Validity criteria fulfilled:

yes

Remarks:

For further details please refer to “Any other information on results incl. tables”.