Reaction mass of hydrogen [1-[(2-hydroxy-4-nitrophenyl)azo]-2-naphtholato(2-)][1-[(2-hydroxy-5-nitrophenyl)azo]-2-naphtholato(2-)]chromate(1-) , compound with 3-[(2-ethylhexyl)oxy]propylamine (1:1) and hydrogen bis[1-[(2-hydroxy-4-nitrophenyl)azo]-2-naphtholato(2-)]chromate(1-) , compound with 3-[(2-ethylhexyl)oxy]propylamine (1:1) and hydrogen bis[1-[(2-hydroxy-5-nitrophenyl)azo]-2-naphtholato(2-)]chromate(1-) , compound with 3-[(2-ethylhexyl)oxy]propylamine (1:1)

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:

26 September 2017 to 28 September 2017

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

Deviations:

no

Qualifier:

according to guideline

Guideline:

EPA OPPTS 850.1010 (Aquatic Invertebrate Acute Toxicity Test, Freshwater Daphnids)

Deviations:

no

Qualifier:

according to guideline

Guideline:

other: Guidelines for studies on the new chemical substance as required by the Law Concerning the Evaluation of Chemical Substances and Regulation of their Manufacture, etc (Chemical Substance Control Law)

Version / remarks:

1973, amended 2009 under the reference of YAKUSHOKHATSU No. 1121002, SEIKYOKU No.2 and KANPOKIHATSU No. 021121002 and partially amended 2006 as the joint ordinance of The Japanese Ministry of Economy Trade and Industry (METI), Ministry of Health, Labour and Welfare (MHLW) and Ministry of the Environment (MOE)

Deviations:

no

Qualifier:

according to guideline

Guideline:

other: JMAFF 2-7-2-1 The guidelines related to the study reports for the registration application of pesticide

Version / remarks:

Ref. No. 12-Nousan-8147 on 24 November 2000) & Ref. No.13-Seisan-3986 on 10 October 2001.

Deviations:

no

Qualifier:

according to guideline

Guideline:

EU Method C.2 (Acute Toxicity for Daphnia)

GLP compliance:

yes (incl. QA statement)

Analytical monitoring:

yes

Details on sampling:

- The test material was measured in the test concentration (100 mg/L nominal loading rates WAF) at the start and end of the experiment.

Vehicle:

no

Details on test solutions:

- Because the Test Material is an UVCB, the test solutions were prepared using the saturated solution method according to the Guidance Document on Aquatic Toxicity Testing of Difficult Substances and Mixtures, OECD No. 23. The saturated Test Material solution (mg/L nominal loading rate WAF) was prepared by dispersing/dissolving the needed amount of Test Material into the test medium (ISO Medium) two days (Day-2) before the start of the treatment.
- This solution was shaken for approximately 24 hours at approximately 30 °C (Day -2) and then was equilibrated for approximately 24 hours at approximately 20 °C (Day -1). The non-dissolved test materials were removed by filtration through a fine (0.22 μm) filter to give the 100 % saturated solution.
- As only limit test was carried out, further dilution of this saturated solution was not performed. Test solution was distributed into test vessels prior to introduction of daphnia.

Test organisms (species):

Daphnia magna

Details on test organisms:

TEST ORGANISM
- Breeding: The Daphnia are bred in Ecotoxicological Laboratory of CiToxLAB Hungary Ltd. The health of the stock animals is continuously monitored by visual daily checking. Abnormal behaviour or significant decrease of population is recorded.
- Age: They were less than 24 h old at the beginning of the test.
- Feeding during test: No

ACCLIMATION
- Acclimation period: There was no acclimatisation because the water used was similar to the culture water.

Test type:

static

Water media type:

freshwater

Limit test:

yes

Total exposure duration:

48 h

Hardness:

248 mg/L (as CaCO3)

Test temperature:

20.4 - 20.7 °C

pH:

7.07 - 7.19

Dissolved oxygen:

7.4 - 8.7 mg/L

Nominal and measured concentrations:

- Nominal: 100 mg/L (WAF)
- Measured mean concentration: 69.45 mg/L
- As the measured concentration at the end of the exposure deviated more than 20 % from the measured concentration at the beginning, biological results are based on the measured mean concentration 69.45 mg/L

Details on test conditions:

TEST SYSTEM
- Test vessels: Glass beakers
- Material, size, headspace, fill volume: At least 5 mL test solution/animal.
- No. of organisms per vessel: 5
- No. of vessels per concentration: 4
- No. of vessels per control: 4

TEST MEDIUM / WATER PARAMETERS
- Reconstituted water (ISO medium, according to OECD 202) was used as dilution water for both the range finding and definitive tests. The same composition of reconstituted water was used for the tests and for breeding the test animals.
- Source/preparation of dilution water: Separate stock solutions of individual trace elements were first prepared in deionised water. The medium was prepared of these different stock solutions, so it contained all trace elements (combined solution). Components of medium: CaCl2.2H2O 11.76 g/L 25 mL/L, MgSO4.7H2O 4.93 g/L 25 mL/L, KCl 0.23 g/L 25 mL/L and NaHCO3 2.59 g/L 25 mL/L.
- Culture medium different from test medium: No
- Intervals of water quality measurement: The water temperature was measured daily, the oxygen concentrations and pH of the control and the test solution were measured at the beginning and at the end of the experiment.

OTHER TEST CONDITIONS
- Adjustment of pH: No
- Photoperiod: 16-hour light and 8-hour dark cycle

EFFECT PARAMETERS MEASURED:
- The immobility or mortality of the Daphnia was determined by visual observation 24 and 48 hours after the start of the test. Those animals not able to swim within 15 seconds after gentle agitation of the test beaker are considered to be immobile.
- The number of immobilised animals and the percentage of immobility were determined at 24 and 48 hours.

RANGE-FINDING STUDY
- Test concentrations (nominal): 0.1, 1, 10, and 100 mg/L (WAF)
A concentration range-finding test was conducted to determine the approximate toxicity of the test item so that appropriate test concentrations could be selected for use in the definitive test. Ten daphnia (divided into two replicates) in each test concentration and control were exposed for 48 hours under semi-static conditions.
- Results used to determine the conditions for the definitive study: Yes, because toxic response was not observed during the preliminary concentration rangefinding test, a limit test was carried out using only one test concentration (100 mg/L nominal loading rate WAF) and one control group in a static system.

Reference substance (positive control):

yes

Remarks:

Potassium dichromate

Key result

Duration:

48 h

Dose descriptor:

EC50

Effect conc.:

> 69.45 mg/L

Nominal / measured:

meas. (geom. mean)

Conc. based on:

test mat.

Basis for effect:

mobility

Key result

Duration:

48 h

Dose descriptor:

EC100

Effect conc.:

> 69.45 mg/L

Nominal / measured:

meas. (geom. mean)

Conc. based on:

test mat.

Basis for effect:

mobility

Key result

Duration:

48 h

Dose descriptor:

NOEC

Effect conc.:

69.45 mg/L

Nominal / measured:

meas. (geom. mean)

Conc. based on:

test mat.

Basis for effect:

mobility

Key result

Duration:

48 h

Dose descriptor:

LOEC

Effect conc.:

> 69.45 mg/L

Nominal / measured:

meas. (geom. mean)

Conc. based on:

test mat.

Basis for effect:

mobility

Key result

Duration:

24 h

Dose descriptor:

EC50

Effect conc.:

> 69.45 mg/L

Nominal / measured:

meas. (geom. mean)

Conc. based on:

test mat.

Basis for effect:

mobility

Details on results:

IMMOBILISATION
- No immobilisation was observed at 24 or 48 hours.
- In addition to immobility, no abnormal behaviour or appearance of test animals was detected.

CONCLUSIONS
- The test material had no toxic effect at the limit of solubility at a loading rate of 100 mg/L, which was found to be 69.45 mg/L WAF; the EC50 results and the LOEC are higher than the solubility level of the test material in the test medium.

Results with reference substance (positive control):

The 24h EC50: 0.64 mg/L, (95 % confidence limits: 0.59 – 0.68 mg/L).

Reported statistics and error estimates:

No statistical analysis was performed because of the lack of toxic effects.
The EC50, NOEC, LOEC and EC100 values were determined directly from the raw data.

Validity

- There were no immobilised animal in the control group and the dissolved oxygen concentration at the end of the test in control and test vessels was more than 3 mg/L.

- All validity criteria were within acceptable limits and therefore the study can be considered as valid.

Validity criteria fulfilled:

yes

Conclusions:

Under the conditions of this study the 24 and 48 hour EC50 of the test material was >69.45 mg/L WAF. The 48h EC100 value and LOEC were also >69.45 mg/L WAF. The NOEC was 69.45 mg/L WAF. The test material had no toxic effect at the limit of solubility.

Executive summary:

The toxicity of the test material to aquatic invertebrates was investigated in accordance with the standardised guidelines OECD 202, OCSPP 850.1010 and JMAFF guidelines under GLP conditions.

Acute toxicity of the test material on Daphnia magna was assessed in an Acute immobilisation test, over an exposure period of 48 hours in a static test system.

Because no toxic response was observed during the preliminary range-finding test, only a limit test was carried out using only one test concentration (100 mg/L nominal loading rate WAF) and one control group in the definitive study in a static system.

The concentration of the test material was analytically determined at the start and at the end of the experiment. Measured concentration was 79.2 mg/L at the start and 59.7 mg/L at the end of the experiment. As the measured concentration at the end of the exposure deviated more than 20 % from the measured concentration at the beginning, biological results are based on the measured mean concentration 69.45 mg/L WAF.

Twenty animals, divided into four replicates (glass beaker) of five animals each were used at the test concentration and for the control. All validity criteria were met during this study.

Under the conditions of this study the 24 and 48 hour EC50 of the test material was >69.45 mg/L WAF. The 48h EC100 value and LOEC were also >69.45 mg/L WAF. The NOEC was 69.45 mg/L WAF.

The test material had no toxic effect at the limit of solubility at a loading rate of 100 mg/L, which was found to be 69.45 mg/L WAF.

Description of key information

Under the conditions of this study the 24 and 48 hour EC50 of the test material was >69.45 mg/L WAF. The 48h EC100 value and LOEC were also >69.45 mg/L WAF. The NOEC was 69.45 mg/L WAF. The test material had no toxic effect at the limit of solubility.

Additional information

The toxicity of the test material to aquatic invertebrates was investigated in accordance with the standardised guidelines OECD 202, OCSPP 850.1010 and JMAFF guidelines under GLP conditions. The study was awarded a reliability score of 1 in accordance with the criteria set forth by Klimisch et al. (1997).

Acute toxicity of the test material on Daphnia magna was assessed in an Acute immobilisation test, over an exposure period of 48 hours in a static test system.

Because no toxic response was observed during the preliminary range-finding test, only a limit test was carried out using only one test concentration (100 mg/L nominal loading rate WAF) and one control group in the definitive study in a static system.

The concentration of the test material was analytically determined at the start and at the end of the experiment. Measured concentration was 79.2 mg/L at the start and 59.7 mg/L at the end of the experiment. As the measured concentration at the end of the exposure deviated more than 20 % from the measured concentration at the beginning, biological results are based on the measured mean concentration 69.45 mg/L WAF.

Twenty animals, divided into four replicates (glass beaker) of five animals each were used at the test concentration and for the control. All validity criteria were met during this study.

Under the conditions of this study the 24 and 48 hour EC50 of the test material was >69.45 mg/L WAF. The 48h EC100 value and LOEC were also >69.45 mg/L WAF. The NOEC was 69.45 mg/L WAF.

The test material had no toxic effect at the limit of solubility at a loading rate of 100 mg/L, which was found to be 69.45 mg/L WAF.