Reaction products of 2,4,6-trichloro-1,3,5-triazine copuled with 3-aminopropyldiethylamine, coupled with 3-acetamidoanilinium chloride, diazotised, coupled with 2,5-Xylidine, coupled with 3-amino-4-hydroxybenzenesulphonamide, further coupled with resorcinol and metallized with iron chloride.

Short-term toxicity to aquatic invertebrates

Link to relevant study record(s)

Reference

Endpoint:

short-term toxicity to aquatic invertebrates

Type of information:

experimental study

Adequacy of study:

key study

Study period:

From May 15 to 17, 2019

Reliability:

1 (reliable without restriction)

Rationale for reliability incl. deficiencies:

guideline study

Qualifier:

according to guideline

Guideline:

OECD Guideline 202 (Daphnia sp. Acute Immobilisation Test)

Version / remarks:

2004

GLP compliance:

yes (incl. QA statement)

Analytical monitoring:

yes

Details on sampling:

- Concentrations: 100 mg/l

Vehicle:

no

Test organisms (species):

Daphnia magna

Details on test organisms:

TEST ORGANISM
- Common name: Daphnia
- Strain/clone: Daphnia magna (Straus)
- Age at study initiation (mean and range, SD): Less than 24 h old at the beginning of the test.
- Sex: female
- Breeding: the Daphnia are bred in the laboratory of TOXI-COOP ZRT
- Source: Laboratory of Hydrobiology (Central Agricultural Office, Directorate of Plant-, and Soil Protection) 2100 Gödöllő, Kotlán S. u. 3. Hungary
- Feeding during test : were not fed during the exposure
- Food type: the Daphnia were fed with concentrated algal suspension of Raphidocelis subcapitata during holding.

ACCLIMATION
- Acclimation period: test animals were bred under similar (or the same) conditions as that used during the exposure period (holding water, temperature, background colour etc.), therefore additional acclimatisation before the test was not necessary. Brood daphnids were maintained in dilution water at the test temperature for at least 48 hours prior to the start of the test.

Test type:

static

Water media type:

freshwater

Limit test:

yes

Total exposure duration:

48 h

Remarks on exposure duration:

The immobility or mortality of the Daphnia was determined by visual observation after 24 and 48 hours of exposure.

Hardness:

The hardness of reconstituted water (ISO medium) used in the study was measured as 249.2 mg/l (as CaCO3).

Test temperature:

The test temperature was in the range of 20.7 – 21.0 °C measured in the test vessels during the test. The additionally measured ambient temperature within the climate chamber was between 20.0 – 21.5°C.

pH:

The pH of the test solutions was not adjusted and not varied by more than 1.5 units in any one test. The pH was in the range of 7.58 – 7.97 during the test.

Dissolved oxygen:

The dissolved oxygen concentration was in the range of 8.20 – 8.84 mg/l during the test.

Nominal and measured concentrations:

Nominal: 100 mg/l
Measured: start: 106 ± 2.6 mg/l, end: 108 ± 3.2 mg/l.

Details on test conditions:

TEST SYSTEM
- Test vessel: beaker
- Material, size, headspace, fill volume: glass beaker; volume app. 50 ml
- Volume of solution: each test vessel contained approximately 40 ml of test solution and was uniquely identified with study number, test group and replicate.
- Aeration: the test solutions were not aerated during the experiment
- No. of organisms per vessel: five animals
- No. of vessels per concentration: four replicates
- No. of vessels per control: four replicates

OTHER TEST CONDITIONS
- Adjustment of pH: the pH of the test solutions was not adjusted
- Light/dark cycle: the experiment was carried out in complete darkness

Reference substance (positive control):

yes

Remarks:

potassium dichromate

Duration:

48 h

Dose descriptor:

NOEC

Effect conc.:

100 mg/L

Nominal / measured:

nominal

Conc. based on:

test mat.

Basis for effect:

mobility

Duration:

48 h

Dose descriptor:

LOEC

Effect conc.:

> 100 mg/L

Nominal / measured:

nominal

Conc. based on:

test mat.

Basis for effect:

mobility

Details on results:

Main test
- Behavioural abnormalities: no
- Other biological observations: no
- Mortality of control: no
- Immobilisation of control: no
- Abnormal responses: no
- Effect concentrations exceeding solubility of substance in test medium: no

Preliminary range-finding test
A non-GLP preliminary range-finding test was conducted to determine the approximate toxicity of the test item. In the static preliminary range-finding test, the nominal concentration of 100 mg/l was prepared by suspending 0.0202 g of test item in 202 ml ISO medium. The further test solutions were prepared by the appropriate diluition of this test item stock solution. Untreated control ran parallel in the test.
Daphnids in the test concentrations and control were exposed for 48 hours. Based on the results of the non-GLP preliminary range-finding test, the main test was a limit test including a loading rate of 100 mg test item/l and a concurrent control group. Measured test item concentration remained within ± 20 % of the nominal over the test period of 48 hours, therefore the biological results are based on the nominal concentrations.

Immobilisation of the test animals in the main test

Nominal concentration	Replicate	Number of treated animals	Number of immobilised animals
[mg/l]			24 h	48 h
Control	1	5	0	0
	2	5	0	0
	3	5	0	0
	4	5	0	0
100	1	5	0	0
	2	5	0	0
	3	5	0	0
	4	5	0	0

Results of the preliminary range-finding test

Nominal concentrations	Untreated	0.1	1	10	100
(mg/l)	control
Number of treated animals	10	10	10	10	10
Number of immobilised animals(at 48h)	0	0	0	0	0

Validity criteria fulfilled:

yes

Conclusions:

In this 48-hour acute toxicity test on Daphnia magna, the obtained results show that the test item Basic Brown 22 has not any toxic effect on the mobility of Daphnia at 100 mg/l test concentration.
The 48-h NOEC was determined to be 100 mg/l.
The 48-h LOEC was determined to be > 100 mg/l.
All validity criteria were met. The results are based on nominal test item concentration.

Executive summary:

Basic Brown 022 was tested on Daphnia magna (Straus) in ISO medium. Nominal concentration of 100 mg/l was used in the main study (limit test). Concurrent control ran.

Measured test item concentration remained within ± 20 % of the nominal over the test period of 48 hours, thus the biological results are based on the nominal concentration.

Twenty animals, divided into four replicates (glass beaker; volume app. 50 ml) of five animals each were exposed to the test concentration or run as control for 48 hours in a static test. The immobility or mortality of the Daphnia was determined by visual observation after 24 and 48 hours of exposure.

For determination of the test item concentration, samples were taken from the test item treated group and control at the start and at the end of the test. Concentration was determined using UV/VIS spectrophotometric method. The measured concentration was 106 % of the nominal at the start and 108 % of the nominal at the end of the test.

NOEC and LOEC were determined directly from the raw data.

All validity criteria were met and therefore the study can be considered as valid. Overall, after a 48 -hour exposure:

NOEC = 100 mg/l

LOEC > 100 mg/l.

Description of key information

48 -h NOEC was determined to be 100 mg/L.

48-h LOEC was determined to be > 100 mg/L.

Additional information

Basic Brown 22 was tested according to OECD guideline 202 at nominal concentration of 100 mg/l in a limit test.

Analytical monitoring of solutions was carried out; measured concentrations remained within ± 20 % of the nominal over the test period of 48 hours, therefore the biological results are based on the nominal concentration.

Twenty animals, divided into four replicates (glass beaker; volume ca. 50 ml) of five animals each were exposed for 48 hours in a static test. Toxic effects were determined in terms of immobility or mortality of daphnids.

All validity criteria were met and therefore the study can be considered as valid.