Disodium; 4-Amino-3-{4-[4-(2,4-diamino-phenylazo)-benzenesulfonylamino]-phenylazo}-5-hydroxy-6-phenylazo-naphthalene-2,7-disulfonate

Toxicity to aquatic plants other than algae

Link to relevant study record(s)

Reference

Endpoint:

toxicity to aquatic plants other than algae

Type of information:

experimental study

Adequacy of study:

key study

Study period:

2020

Reliability:

1 (reliable without restriction)

Rationale for reliability incl. deficiencies:

test procedure in accordance with generally accepted scientific standards and described in sufficient detail

Qualifier:

according to guideline

Guideline:

OECD Guideline 221 (Lemna sp. Growth Inhibition Test)

GLP compliance:

yes (incl. QA statement)

Analytical monitoring:

yes

Details on sampling:

Stock solution, media preparation: 119 mg test item/L were freshly prepared with dilution water. The dispersion was agitated until the solution was visually clear.
Test concentrations: The stock solution was diluted to receive the required test item concentrations. Seven concentrations were tested with a spacing factor of √10: 0.119 – 0.376 – 1.19 – 3.76 – 11.9 – 37.6 – 119 mg/L (0.100 – 0.316 – 1.00 – 3.16 – 10.0 – 31.6 – 100 mg a.s./L), corresponding to the geometric mean measured test item concentrations: 0.0827 – 0.142 – 0.260– 0.492 – 1.32 – 4.64 – 20.5 mg test item/L. The concentration levels are based on the results of a preliminary range finding test (non-GLP).

Vehicle:

yes

Test organisms (species):

Lemna gibba

Details on test organisms:

Test organism: Duckweed, Lemna gibba, Lemnaceae, Arales, Arecidae, Monocotyledonae Young, rapidly growing plants without visible lesions or discolouration (chlorosis) were used for the test.
Reason for the selection of the test organism: According to the guideline, Lemna gibba is a suitable species because it is a representative of temperate areas commonly used for toxicity tests
Origin: EUROFINS-GAB GMBH, Eutinger Str. 24, 75223 NiefernÖschelbronn, Germany
Cultivation at test facility: The species is cultured in the test facility. Density is kept low to prevent conglomerates of plants on the surface. At least once per week, plants are transferred to freshly prepared growth medium. Growth media and culturing vessels are autoclaved before use to enable the breeding of axenic cultures. The cultures are visibly free from contamination by other organisms such as algae.
Breeding vessels: Crystallisation dishes of glass, Vol. 900 mL, filled with ca. 500 mL growth medium, covered with glass tops
Medium 20X-AAP-medium (Algal Assay Procedure medium),
pH-value 7.5 ± 0.1
Temperature 24 ± 2 °C
Light regime Continuous fluorescent light, 1100 – 4440 lux

Test type:

static

Water media type:

freshwater

Limit test:

yes

Total exposure duration:

7 d

Hardness:

274 [mg CaCO3/L]

Test temperature:

24 ± 2 °C

pH:

7.5 ± 0.1

Conductivity:

1388 [μS/cm]

Nominal and measured concentrations:

Based on a preliminary test, 5 test item concentrations were tested in a geometrical series with a dilution factor of √10: 0.119 – 0.376 – 1.19 – 3.76 – 11.9 – 37.6 – 119 mg test item/L (0.100 – 0.316 –1.00 – 3.16 – 10.0 – 31.6 – 100 mg a.s./L), corresponding to the geometric mean measured test item concentrations: 0.0827 – 0.142 – 0.260– 0.492 – 1.32 – 4.64 – 20.5 mg test item/L.

Reference substance (positive control):

yes

Remarks:

3,5-dichlorophenol

Duration:

7 d

Dose descriptor:

EC50

Effect conc.:

ca. 22.1 mg/L

Nominal / measured:

nominal

Conc. based on:

test mat.

Details on results:

The concentrations of the test item Acid Black 234 and the control were analysed via HPLC-DAD at the beginning and end of the exposure. The measured concentrations of the test item were in the range of 98 to 117% of the nominal test item concentrations at 0 days and at 7 days in the range of ˂ LOQ to 3% of the nominal test item concentrations.
All effect values are given based on the geometric mean measured test item concentrations.
The EC50-values for inhibition of the specific growth rate of fronds (ErC50) and dry weight (ErdwC50) were 22.1 (18.8 - > 20.5) mg/L and 18.3 (15.1 - > 20.5) mg/L, respectively. The EC50-values for inhibition of yield (fronds, EyC50) and dry weight inhibition of yield (EydwC50) were 8.55 (4.25 – 12.6) mg/L and 1.97 (0.893 – 5.00) mg/L, respectively.

Validity criteria fulfilled:

yes

Conclusions:

Acid Black 234 was found to inhibit the growth of the monocotyledonous aquatic plant Lemna gibba after 7-day exposure under static conditions, with the following effect values
(geometric mean measured test item concentrations): The EC50-values for inhibition of the specific growth rate of fronds (ErC50) and dry weight (ErdwC50) were 22.1 (18.8 - > 20.5) mg/L and 18.3 (15.1 - > 20.5) mg/L, respectively. The EC50-values for inhibition of yield (fronds, EyC50) and dry weight inhibition of yield (EydwC50) were 8.55 (4.25 – 12.6) mg/L and 1.97 (0.893 – 5.00) mg/L, respectively.

Executive summary:

The effects of the test item Acid Black 234 on the growth of the monocotyledonous aquatic plant species Lemna gibba were determined according to the principles of OECD 221 and Council Regulation (EC) No. 266/2016 Method C. 26.

Lemna gibba was exposed to the test item for 7 days under static conditions. Based on a preliminary test, 5 test item concentrations were tested in a geometrical series with a dilution factor of √10: 0.119 – 0.376 – 1.19 – 3.76 – 11.9 – 37.6 – 119 mg test item/L (0.100 – 0.316 – 1.00 – 3.16 – 10.0 – 31.6 – 100 mg a.s./L), corresponding to the geometric mean measured test item concentrations: 0.0827 – 0.142 – 0.260– 0.492 – 1.32 – 4.64 – 20.5 mg test item/L. Three replicates were investigated for each test concentration and six for the control. Frond numbers were assessed on days 0, 2, 5 and 7. Environmental parameters (light, pH and temperature) were within the acceptable limits. The validity criteria of the test guideline were fulfilled. The test item concentrations were concentration-dependent blue-green colored from pale blue to deep bluegreen (almost black).

The concentrations of the test item Acid Black 234 and the control were analysed via HPLC-DAD at the beginning and end of the exposure. All effect values are given based on the geometric mean measured test item concentrations.

The EC50-values for inhibition of the specific growth rate of fronds (ErC50) and dry weight (ErdwC50) were 22.1 (18.8 - > 20.5) mg/L and 18.3 (15.1 - > 20.5) mg/L, respectively. The EC50-values for inhibition of yield (fronds, EyC50) and dry weight inhibition of yield (EydwC50) were 8.55 (4.25 – 12.6) mg/L and 1.97 (0.893 – 5.00) mg/L, respectively.

Description of key information

EC 50: 22.1 mg/l for dry weight

Key value for chemical safety assessment

EC50 for freshwater plants:

22.1 mg/L

Additional information

The effects of the test item Acid Black 234 on the growth of the monocotyledonous aquatic plant species Lemna gibba were determined according to the principles of OECD 221 and Council Regulation (EC) No. 266/2016 Method C. 26.

Lemna gibba was exposed to the test item for 7 days under static conditions.

The concentrations of the test item Acid Black 234 and the control were analysed via HPLC-DAD at the beginning and end of the exposure. All effect values are given based on the geometric mean measured test item concentrations.

The EC50-values for inhibition of the specific growth rate of fronds (ErC50) and dry weight (ErdwC50) were 22.1 (18.8 - > 20.5) mg/L and 18.3 (15.1 - > 20.5) mg/L, respectively. The EC50-values for inhibition of yield (fronds, EyC50) and dry weight inhibition of yield (EydwC50) were 8.55 (4.25 – 12.6) mg/L and 1.97 (0.893 – 5.00) mg/L, respectively.

Moreover, a supporting test on similar substance 1 is available (BMG, 2012) following OECD 221. The tested aquatic plant is Lemna minor and the EC50 for grow rate and biomass is > 2000 mg/l.