Trisodium bis[3-[(4,5-dihydro-3-methyl-5-oxo-1-phenyl-1H-pyrazol-4-yl)azo]-4-hydroxy-5-nitrobenzenesulphonato(3-)]chromate(3-)

Short-term toxicity to aquatic invertebrates

Link to relevant study record(s)

Reference

Endpoint:

short-term toxicity to aquatic invertebrates

Type of information:

other: read-across from supporting substance (structural analogue or surrogate)

Adequacy of study:

key study

Study period:

From June 03 to July 31, 2014

Reliability:

2 (reliable with restrictions)

Rationale for reliability incl. deficiencies:

guideline study

Remarks:

The Read Across approach justification is reported in the endpoint summary

Qualifier:

according to guideline

Guideline:

OECD Guideline 202 (Daphnia sp. Acute Immobilisation Test)

Version / remarks:

adopted 13 April 2004

GLP compliance:

yes (incl. QA statement)

Analytical monitoring:

yes

Details on sampling:

After sampling, the test medium samples (0.5 ml) were mixed with 0.5 ml methanol and stored deep-frozen (below -18 °C) until analysis.

Vehicle:

no

Details on test solutions:

The concentrations in the main test were spaced by a factor of 2.2. The necessary amount of the test item for preparing the stock solution was weighed on weighing scoops and transferred to a volumetric flask. Test medium was added up to the bench mark and the solution was homogenised by shaking. The lower test solutions were prepared by dilution of the stock solution. Defined volumes of the prepared solution were transferred to each test vessels. The test solution volume was 50 ml per test vessel.

Test organisms (species):

Daphnia magna

Details on test organisms:

TEST ORGANISM
- Common name: Daphnia magna STRAUS, Clone V
- Source: the animals are originally purchased in a healthy condition from the Umweltbundesamt in Berlin/Germany.
- Age at study initiation: freshly hatched daphnids max. 24 hours old were used for the test.
- Breeding: the animals are continuously bred in the laboratory. Daphnia magna was bred as single culture (one daphnid per 100 ml) in Elendt M4 medium.
- Feeding during test: none
- Food type: animals were fed with single cell green algae (Desmodesmus subspicatus, former Scenedesmus subspicatus) at least three times a week.

CONDITIONS
- Medium: medium was changed three times per week
- pH: range of 6.0 – 9.0
- Dissolved oxygen: above 60 % saturation.
- Total hardness: total hardness 140 - 250 mg/l (as CaCO3), corresponding to 7.8 - 14 °dH.
- Aerations: yes
- Temperature: 20 ± 2 °C
- Light cycle day: 16 h of illumination and 8 h of darkness.

Test type:

static

Water media type:

freshwater

Limit test:

no

Total exposure duration:

48 h

Hardness:

13 °dH (232 mg/l as CaCO3)

Test temperature:

20.1 - 21.3 °C

pH:

7.95 - 8.08

Dissolved oxygen:

≥ 8.3 mg/l

Nominal and measured concentrations:

1.94, 4.27, 9.39, 20.7, 45.5 and 100 mg/l

Details on test conditions:

TEST SYSTEM
- Test vessel: 100 ml glass beakers.
- Loading: at least 10 ml of test solution for each animal.
- Material, size, headspace, fill volume: filled with 50 ml.
- Aeration: none.
- No. of organisms per vessel: 20 per concentration.
- No. of vessels per concentration: four replicates.

TEST MEDIUM
- Elendt M4 medium was used as test medium.

OTHER TEST CONDITIONS
- Photoperiod: 16 hours photoperiod / 8 hours darkness daily.

observations
After 24 h and 48 h the immobilised daphnids were counted. All daphnids not able to swim within 15 seconds after gentle agitation of the test vessel were
considered to be immobilised. If present, behavioural changes of daphnids were recorded at 24 and 48 hours after starting the test.

MEASUREMENTS
The test temperature and the pH-value as well as the oxygen concentration rate of the test media were measured at all concentrations at t = 0 h, 24 h and 48 h in one replicate per test item concentration.

Reference substance (positive control):

yes

Remarks:

potassium dichromate

Duration:

48 h

Dose descriptor:

EC50

Effect conc.:

24.1 mg/L

Nominal / measured:

nominal

Conc. based on:

test mat.

Basis for effect:

mobility

Remarks on result:

other: 15.9 - 36.6

Duration:

48 h

Dose descriptor:

NOEC

Effect conc.:

9.39 mg/L

Nominal / measured:

nominal

Conc. based on:

test mat.

Basis for effect:

mobility

Details on results:

After 24 hours of exposure no immobilisation above the allowed control immobilisation was observed in the control and up to 45.5 mg/l. 70 % of the daphnids were found immobile at 100 mg/l.
After 48 hours of exposure no immobilisation above the allowed control immobilisation was observed in the control and up to 9.39 mg/l. 20 % of the daphnids were immobilised at 20.7 mg/l. At the two highest test item concentrations of 45.5 and 100 mg/l all daphnids were found immobile.

Results with reference substance (positive control):

The results indicate an EC50 (24 h) of the reference item potassium dichromate between 1.0 and 2.0 mg/l in the range-finding test. Since the results fall within the range recommended by the guideline (OECD 202), the daphnids were suitable for the determination of the toxicological effects of the test item.

Reported statistics and error estimates:

The 24 h and 48 h EC50 are the estimated concentrations where 50 % of the daphnids were immobilised after 24 and 48 hours, respectively.
For the evaluation of the 24 h and 48 h EC50, logit analysis using linear max. likelihood regression was used. The evaluation of data was performed by using ToxRat Professional 2.10.
The NOEC was established based on the highest concentration at which the immobilisation is not higher than the allowed control immobilisation (≤ 10 % immobilisation).

Nominal and measured concentrations of the test item.

Nominal conc. [mg/l]	Sampling [h]	Measured conc. [mg/l]	% of nominal
Control	0 fresh	n.d.	-
	48 aged	n.d.	-
1.94	0 fresh	1.85	95
	48 aged	1.82	94
4.27	0 fresh	4.03	94
	48 aged	3.98	93
9.39	0 fresh	8.9	95
	48 aged	8.67	92
20.7	0 fresh	20.1	97
	48 aged	19.3	93
45.5	0 fresh	44	97
	48 aged	43.3	95
100	0 fresh	99.2	99
	48 aged	96.5	97

- = not calculated; n.d. = not detectable; LOQ = 1.00 mg/l test item

The initial mean measured content of the test item was 96 % of nominal, the arithmetic mean of fresh and aged samples was 95 % of nominal.

Since the mean measured concentration of the test item was between 80 – 120 % of nominal, the toxicological endpoints were evaluated using nominal concentrations of the test item.

Validity criteria fulfilled:

yes

Remarks:

in the control not more than 10 % of animals were immobilised; the dissolved oxygen concentration was ≥ 3 mg/l at the end of the test in control and test vessels

Conclusions:

EC50 (48 h) for immobilisation: 24.1 mg test item/l (nominal).

Executive summary:

Method

Acute toxicity of substance to Daphnia Magna was investigated in a 48 hours test. Twenty organisms per test concentration (4 replicates of 5 animals each) were used. A static main test with concentrations of 0, 1.94, 4.27, 9.39, 20.7, 45.5 and 100 mg/l was performed.

Result

According to the results of the test, the EC50 (48 h) for immobilisation was determined to be 24.1 mg test item/l (nominal). The corresponding NOEC (48 h) was 9.39 mg test item/l (nominal).

Description of key information

EC50 (48 h) for immobilisation: 24.1 mg test item/l (nominal).

Key value for chemical safety assessment

Fresh water invertebrates

Fresh water invertebrates

Effect concentration:

24.1 mg/L

Additional information

No studies on the short-term toxicity to aquatic invertebrates are available for the Acid Red 357 (AR357).

Nevertheless, a study conducted on the structural analogue Acid Red 315 (AR315 - Similar Substance 01) is available. Acute toxicity of substance to Daphnia Magna was investigated in a 48 hours test. Twenty organisms per test concentration (4 replicates of 5 animals each) were used and the static main test was conducted at the concentrations of 0, 1.94, 4.27, 9.39, 20.7, 45.5 and 100 mg/l.

After 48 hours of exposure no immobilisation above the allowed control immobilisation was observed in the control and up to 9.39 mg/l. 20 % of the daphnids were immobilised at 20.7 mg/l. At the two highest test item concentrations of 45.5 and 100 mg/l all daphnids were found immobile.

Since the mean measured concentration of the test item was between 80 – 120 % of nominal, the test results can be considered as reliable and representative.

AR357 and AR315 are both textile pre-metalized complex dyes 1:2 and they are manufactured for the same use, thus the characteristics of the final product are strictly similar and share the same functionality.

Both AR357 and AR315 are chromate pyrazol sodium salts: AR357 is a trisodium, while AR315 is a disodium salt. They share the same structure, except for the fact that AR357 has two sulphonated groups, instead one as AR315. It is expected that this difference has not impact on the potential of short-term toxicity to aquatic invertebrates.

The typical commercial batches present some differences. The main component AR357 is commonly greater than 55 percent, while in the case of AR315 the main component was stated at 80 %. From this point of view, the major active ingredient concentration of AR315 leads to consider the read-across approach as conservative.

The major difference in the composition is due to the elevated concentration of inorganic salts (i.e. sodium chloride and disodium sulphate) in the AR357, which is a consequence of the salification procedure. In general, the impurities do not impact the experimental results and the endpoint assessment.

Considering that the substances are both salts derived from strong acids (sulphonic acid derivatives), it is expected that they would be completely dissociated and stable in water for the whole test period in that form.