1-(2-bromo-4-chloro-phenyl)-4-(trifluoromethyl)triazole

Short-term toxicity to aquatic invertebrates

Administrative data

Endpoint:

short-term toxicity to aquatic invertebrates

Type of information:

experimental study

Adequacy of study:

key study

Study period:

25 Jan - 28 Jan 2022

Reliability:

1 (reliable without restriction)

Rationale for reliability incl. deficiencies:

guideline study

Data source

Reference

Reference Type:

study report

Title:

Unnamed

Year:

2022

Report date:

2022

Materials and methods

Test guideline

Qualifier:

according to guideline

Guideline:

OECD Guideline 202 (Daphnia sp. Acute Immobilisation Test)

Version / remarks:

adopted April 13, 2004

Deviations:

no

GLP compliance:

yes (incl. QA statement)

Test material

Test material information

Constituent 1

Reference substance name:

1-(2-bromo-4-chlorophenyl)-4-(trifluoromethyl)-1H-1,2,3-triazole

EC Number:

863-757-3

Cas Number:

1445866-55-6

Molecular formula:

C9H4BrClF3N3

IUPAC Name:

1-(2-bromo-4-chlorophenyl)-4-(trifluoromethyl)-1H-1,2,3-triazole

Sampling and analysis

Analytical monitoring:

yes

Details on sampling:

- Sampling method: Duplicate samples from the freshly prepared test media of all test concentrations and the control were taken at the start of the test.
For the determination of the stability of the test item under the test conditions and of the maintenance of the test item concentrations during the test period, duplicate samples from the test media of all test concentrations and the control were collected at the end of the test (after 48 hours) by pouring together the contents of the test beakers of each treatment. All samples were diluted by a factor of 2 with acetonitrile. Additional samples of the untreated controls and the sample dilution solvent were taken at each sampling test start and test end without any sample treatment.
For the determination of the stability of the test item under the test conditions and of the maintenance of the test item concentrations during the test period.
- Sample storage conditions before analysis: deep frozen

Test solutions

Vehicle:

no

Details on test solutions:

PREPARATION AND APPLICATION OF TEST SOLUTION
The test item is not well soluble in test medium. To avoid physical effects of undissolved test item on the daphnids, no concentrations above the solubility limit of the test item in test water was tested. Therefore, a supersaturated stock solution with a loading rate of nominal 100 mg test item/L was prepared by suspending 103.2 mg test item in 1032 mL test water. The stock suspension was stirred for 24 hours at room temperature in the dark to dissolve as much test item as possible. Then, non-dissolved fractions of the test item were separated from the test medium by membrane filtration (0.45 µm cellulose acetate filter). The solution with dissolved test item was used as the test medium of the highest test concentration and to prepare the desired 1:3.3, 1:10.9, 1:35.9 and 1:118.6 dilutions.
The test media were prepared just before introduction of the daphnids (= start of the test).
In the control, test water was used without addition of the test item.

Test organisms

Test organisms (species):

Daphnia magna

Details on test organisms:

TEST ORGANISM
Daphnia magna (Straus), clone 5
Age at Test Start: From 0.75 to 18.25 hours old
Source: The daphnids introduced in the test were taken from ibacon's in-house laboratory culture.
The daphnids were bred in the laboratories of ibacon under similar temperature and light conditions as used in the test. The cultivation of the parental daphnids was performed in Elendt M4 medium. The test organisms were not first brood progeny. The daphnids in the stock culture were fed at least on all working days with green algae (Desmodesmus subspicatus) freshly grown in the laboratories of ibacon.
Feeding during the test: none
Acclimatisation: was not necessary, since the test was performed in the same medium as the culturing.

Study design

Test type:

static

Water media type:

freshwater

Limit test:

no

Total exposure duration:

48 h

Test conditions

Hardness:

Water Hardness: 2.5 mmol/L (= 250 mg/L) as CaCO3

Test temperature:

18.1 to 19.1 °C at test start;
18.1 to 18.8 °C at test end

pH:

7.9 to 8.0 at test start;
7.8 to 8.0 at test end; and thus the pH-value did not vary by more than 1.5 unit

Dissolved oxygen:

9.2 to 9.7 mg/L at test start;
11.2 to 12.4 mg/L at test end

Nominal and measured concentrations:

A filtrate with a loading rate of 100 mg test item/L and dilutions of 1:3.3, 1:10.9, 1:35.9 and 1:118.6, and a control, corresponding to following arithmetic mean measured concentrations of the test item:
4.89, 1.47, 0.456, 0.100 and 0.025 mg test item/L, and a control.

The initial concentrations and the maintenance of the exposure concentrations during the test were verified in the analytical part. All reported results refer to nominal values since the concentrations of the test item were within ± 20% of the nominal concentrations during the test.

Details on test conditions:

Test Vessel: Glass beakers of 100 mL volume containing approximately 60 mL of test medium. The test vessels were covered with a lid to reduce the loss of water due to evaporation and to avoid the entry of dust into the solutions.
Light Regime: 16 h light : 8 h dark
Light Intensity: The light intensity was 1060 to 1080 lux (measured once during the test). Target: ≤ 1500 lux
Introduction of Daphnids: 20 daphnids per control and test concentration, divided into 4 groups of 5 animals, each group in 60 mL test medium
Replicates: The test was performed with four replicates per treatment group. Exposure Time: 48 hours
Test Procedure: A static test was performed.
Feeding: None
Sampling: Samples were taken at test start (0 hours) and test end (48 hours)

Test Parameter:
Immobility: The mobility of the daphnids was determined by visual observation after 24 and 48 hours. Those animals that are not able to swim within 15 seconds after gentle agitation of the test beaker were considered to be immobile (even if they could still move their antennae).

No pre-experiments were performed.

Reference substance (positive control):

yes

Remarks:

For the evaluation of the quality of the Daphnia clone and the consistency of the experimental conditions, the reference item potassium dichromate is tested at least twice a year to demonstrate satisfactory test conditions.

Results and discussion

Effect concentrations

Effect concentrations 1

Key result

Duration:

48 h

Dose descriptor:

EC50

Effect conc.:

2.53 mg/L

95% CI:

> 1.15 - 4.89

Nominal / measured:

nominal

Conc. based on:

test mat.

Basis for effect:

mobility

Effect concentrations 2

Duration:

48 h

Dose descriptor:

EC10

Effect conc.:

1.32 mg/L

95% CI:

> 0.38 - < 4.61

Nominal / measured:

nominal

Conc. based on:

test mat.

Basis for effect:

mobility

Effect concentrations 3

Duration:

48 h

Dose descriptor:

NOEC

Effect conc.:

1.47 mg/L

Nominal / measured:

nominal

Conc. based on:

test mat.

Basis for effect:

mobility

Effect concentrations 4

Duration:

48 h

Dose descriptor:

LOEC

Effect conc.:

4.89 mg/L

Nominal / measured:

nominal

Conc. based on:

test mat.

Basis for effect:

mobility

Details on results:

Signs of Intoxication after 48 Hours:
After 48 hours of exposure no immobilisation of the test animals was observed in the control and up to and including the test item concentration of 0.100 mg test item/L. At the concentrations of 0.456 and 1.47 mg test item/L, one animal was immobile. All animals were immobile at the concentration of 4.89 mg test item/L.

Results with reference substance (positive control):

Results of the most recent Test with the Reference Item Potassium Dichromate in July 2021
Dose response test: 0.40, 0.60, 0.89, 1.33, 2.0 mg/L
EC50 24h: 1.21 mg/L
NOEC 24 h: 0.60 mg/L
LOEC 24 h: 0.89 mg/L
EC50 48 h: 0.844 mg/L
NOEC 48 h: 0.40 mg/L
LOEC 48 h: 0.60 mg/L

The reference test was valid; The results indicate that the sensitivity of the daphnids was consistent with the level proposed by the OECD 202 guideline (EC50-24 h between 0.6 and 2.1 mg potassium dichromate/L).

Reported statistics and error estimates:

The 24-hour and 48-hour EC50, EC20 and EC10 and the 95 % confidence limits were calculated by weibull analysis.
The NOEC and LOEC after 24 hours were calculated by Fisher´s exact binominal test with Bonferroni corrections and the NOEC and LOEC after 48 hours were calculated by Step-down Cochran Armitage test procedure.
The software used to perform the statistical analysis was ToxRat Professional, Version 3.3.0, ToxRat(R) Solutions GmbH.

Any other information on results incl. tables

Validity criteria

Criterion	value required	value obtained	fullfilled
Control immobilisation rate	< 10%	0%	yes
Dissolved oxygen concentration	at the end of the test >= 3 mg O2/L	≥ 11.2 mg O2/L in in all treatment groups at the end of the test	yes

Applicant's summary and conclusion

Validity criteria fulfilled:

yes

Remarks:

see "Any other information on results incl. tables"

Conclusions:

The toxic effect of the test item to Daphnia magna was assessed in a static concentration-response test. The 48-hour NOEC was determined to be 1.47 mg/L. The 48-hour LOEC was determined to be 4.89 mg/L and the 48-hour EC50 value was calculated to be 2.53 mg/L.
The initial concentrations and the maintenance of the exposure concentrations during the test were verified in the analytical part. All reported results refer to arithmetic mean concentrations, since the test item concentrations were not within ± 20 % of the nominal concentrations during the test.

Executive summary:

Test Species: Daphnia magna, clone 5; 0.75 to 18.25 hours  old

Test Design: This study encompassed 6 treatment groups (5 dose rates of the test item and a control) each containing 20 individuals. The mobility of the daphnids was determined in a static 48-hour test by visual observation after 24 and 48 hours. 

Endpoints: Number of immobile organisms after 24 and 48 hours

Test Concentrations: A filtrate with a loading rate of 100 mg test item/L and dilutions of 1:3.3, 1:10.9, 1:35.9 and 1:118.6, and a control, corresponding to following arithmetic mean measured concentrations of the test item:

4.89, 1.47, 0.456, 0.100 and 0.025 mg test item/L, and a control.

Test Conditions: Water temperature: 18.1 to 19.1 °C (target: 18 °C – 22 °C); pH value: 7.8 to 8.0 (target: 6 - 9); dissolved oxygen concentration: 9.2 to 12.4 mg/L (target: ≥ 3 mg/L); photoperiod: 16 h light - 8 h dark; light intensity: 1060 to 1080 lux (target: < 1500 lux); and thus were within the ranges requested by guideline OECD 202

Results: After 48 hours of exposure no immobilisation of the test animals was observed in the control and up to and including the test item concentration of 0.100 mg/L. At the concentrations of 0.456 and 1.47 mg/L, one animal was immobile. All animals were immobile at the concentration of 4.89 mg/L. No sublethal effects on the daphnids were observed in any treatment.

Parameter
	after 24 h	after 48 h
EC50 mg test item/L	> 4.89	2.53
EC20 mg test item/L	> 4.89	1.71
EC10 mg test item/L	> 4.89	1.32
NOEC mg test item/L	>= 4.89 mg/L	1.47
LOEC mg test item/L	> 4.89	4.89

Analytical Results: The quantification of the test item in the test samples was performed using liquid chromatography with UV detection.

The concentrations of the test item were determined in the duplicate test media samples from all test concentrations (test item stock solution and its dilutions) and the duplicate control samples from all sampling times. The mean measured values are

4.89 mg test item/L in the filtrate,

1.47 mg test item/L in the 1:3.3 dilution of filtrate,

0.456 mg test item/L in the 1:10.9 dilution of filtrate,

0.100 mg test item/L in the 1:35.9 dilution of filtrate,

0.025 mg test item/L in the 1:118.6 dilution of filtrate.

Conclusion: The toxic effect of the test item to Daphnia magna was assessed in a static concentration-response test. The 48-hour NOEC was determined to be 1.47 mg/L. The 48-hour LOEC was determined to be 4.89 mg/L and the 48-hour EC₅₀ value was calculated to be 2.53 mg/L.The initial concentrations and the maintenance of the exposure concentrations during the test were verified in the analytical part. All reported results refer to arithmetic mean concentrations, since the test item concentrations were not within ± 20 % of the nominal concentrations during the test.