1-(3,5-Dichloro-4-fluoro-phenyl)-2,2,2-trifluoro-ethanone

Toxicity to aquatic algae and cyanobacteria

Administrative data

Endpoint:

effects on growth of green algae

Type of information:

experimental study

Adequacy of study:

key study

Reliability:

1 (reliable without restriction)

Rationale for reliability incl. deficiencies:

guideline study

Data source

Reference

Reference Type:

study report

Title:

Unnamed

Year:

2020

Report date:

2020

Materials and methods

Test guideline

Qualifier:

according to guideline

Guideline:

OECD Guideline 201 (Freshwater Alga and Cyanobacteria, Growth Inhibition Test)

Version / remarks:

2011

Deviations:

no

GLP compliance:

yes (incl. QA statement)

Test material

Test material information

Constituent 1

Reference substance name:

1-(3,5-dichloro-4-fluorophenyl)-2,2,2-trifluoroethan-1-one

EC Number:

829-719-5

Cas Number:

1190865-44-1

Molecular formula:

C8H2Cl2F4O

IUPAC Name:

1-(3,5-dichloro-4-fluorophenyl)-2,2,2-trifluoroethan-1-one

Test material form:

liquid

Sampling and analysis

Analytical monitoring:

yes

Details on sampling:

For the determination of the test item concentrations, duplicate samples (10 mL) were taken from the test media of all test concentration groups and the control at the start and at the end of the exposure (96 hours). For sampling at the end of the exposure, the contents of the replicate test vessels were combined prior to sampling.

Test solutions

Vehicle:

no

Details on test solutions:

Test was conducted in reconstituted test water (AAP Medium) prepared according to the test guideline. Analytical grade salts were dissolved in sterile purified water to obtain the following concentrations: 15 mg/L NaHCO3, 1.044 mg/L K2HPO4, 14.6 mg/L MgSO4 x 7 H2O, 12.16 mg/L MgCl2 x 6 H2O, 4.41 mg/L CaCl2 x 2 H2O, 25.5 mg/L NaNO3, 0.186 mg/L H3BO3, 0.415 mg/L MnCl2 x 4 H2O, 0.00327 mg/L ZnCl2, 0.00143 mg/L CoCl2 x 6 H2O, 0.000012 mg/L CuCl2 x 2 H2O, 0.00726 mg/L Na2MoO4 x 2 H2O, 0.16 mg/L FeCl3 x 6 H2O and 0.3 mg/L Na2EDTA x 2 H20.
A stock solution of the nominal concentration of 100 mg/L was prepared by dissolving 55.32 mg of test item completely in 553 mL of test water using ultrasonic treatment for 15 minutes and intense stirring for 15 minutes at room temperature. The stock solution was diluted with test water to prepare the test medium of the highest test concentration.

Test organisms

Test organisms (species):

Raphidocelis subcapitata (previous names: Pseudokirchneriella subcapitata, Selenastrum capricornutum)

Details on test organisms:

Raphidocelis subcapitata (formerly Pseudokirchneriella subcapitata, occasionally also listed as Selenastrum capricornutum), Strain No. 61.81 SAG, supplied by the Collection of Algal Cultures (SAG, Institute for Plant
Physiology, University of Göttingen, 37073 Göttingen / Germany). The algae were cultivated at the test facility under standardized conditions according to the test guidelines.
An inoculum culture of algae was set up three days before the start of the test. The algae were cultivated under the test conditions and kept in the exponential growth phase until inoculation of the test solutions. The inoculum culture was incubated under the same conditions and dilution media as the test cultures.

Study design

Test type:

static

Water media type:

freshwater

Limit test:

no

Total exposure duration:

96 h

Test conditions

Hardness:

15 mg/L as CaCO3

Test temperature:

21 °C

pH:

7.4 at test start to 8.5 at end of experiment (control), 7.4 at test start to 8.9 at end of experiment (test media)

Nominal and measured concentrations:

Nominal: 0, 0.1, 0.32, 1.0, 3.2, 10 and 32 mg/L
Mean measured:

Details on test conditions:

The test design included three replicates per test concentration and six replicates of the control.
The test was started using a nominal algal cell density of 5000 cells/mL. The initial cell density was selected according to the recommendations of the OECD test guideline. The algal cell density in the pre-culture was determined using an electronic particle counter (Cell Counter CASY TT, OLS, Bremen/Germany).
Erlenmeyer flasks of volume 75 mL were used as incubation vessels. The volume of test solution in each test flask was 30 mL per replicate. Each test flask was loosely covered with a glass lid, thereby adequate CO2 be transferred from surrounding air to the test solution.
The test flasks were incubated in a temperature controlled orbital shaker (approximately 125 rpm, Multitron-Pro, Infors HT, Bottmingen/Switzerland) at a temperature maintained at 21 °C. The test flasks were positioned randomly and repositioned daily. They were continuously illuminated by LED lights installed above the test flasks. The light intensity at the level of the test solutions was approximately 73 μE/sm2 (range: 70 to 75 μE/sm2, measured at nine places in the experimental area). The light intensity over the incubation area was within a ±15%-deviation from the average light intensity as recommended by the guideline.

Reference substance (positive control):

yes

Remarks:

For evaluation of the algal quality and experimental conditions, potassium dichromate is tested as a positive control twice a year in the test facility to demonstrate satisfactory test conditions.

Results and discussion

Effect concentrations

Effect concentrations 1

Key result

Duration:

72 h

Dose descriptor:

EC50

Effect conc.:

3.6 mg/L

Nominal / measured:

meas. (geom. mean)

Conc. based on:

test mat.

Basis for effect:

growth rate

Remarks on result:

other: 95% confidence limits of 3.3 and 3.9 mg/L

Effect concentrations 2

Key result

Duration:

72 h

Dose descriptor:

EC10

Effect conc.:

0.85 mg/L

Nominal / measured:

meas. (geom. mean)

Conc. based on:

test mat.

Basis for effect:

growth rate

Remarks on result:

other: 95% confidence limits of 0.71 and 0.99 mg/L

Effect concentrations 3

Duration:

72 h

Dose descriptor:

NOEC

Effect conc.:

0.2 mg/L

Nominal / measured:

meas. (geom. mean)

Conc. based on:

test mat.

Basis for effect:

growth rate

Details on results:

The microscopic examination of the algal cells at the end of the test showed no difference between the algae growing at the mean measured concentration of 2.1 mg/L (3.2 mg/L nominal) and the algal cells in the control. The shape and size of the algal cells were not affected by the test item up to at least this concentration.

Results with reference substance (positive control):

Results of the reference test for potassium dichromate confirmed sensitivity of the test system, with a 72-hour EC50 for growth rate of 0.92 mg/L with 95% confidence limits of 0.85 and 0.99 mg/L determined.

Reported statistics and error estimates:

The values for the validity criteria of the test were calculated by a statistical program. The test was valid since the following performance criteria (according to the OECD 201) were met.
- In the control, the biomass increased by a factor of 116 over 72 hours. (Criterion: increase by at least a factor of 16 within three days).
- The mean coefficient of variation of the daily growth rates in the control (section-bysection growth rates) during 72 and 96 hours was 11 and 17%, respectively. (Criterion: must not be higher than 35%).
- The coefficient of variation of the average specific growth rates in the replicates of the control after 72 and 96 hours was 1.2 and 0.8%, respectively. (Criterion: must not be higher than 7%).

Any other information on results incl. tables

Table 1: Average growth rates

Mean measured concentration (mg/L)	Replicate number	Growth rate μ (1/day), 24 hrs	Growth rate μ (1/day), 48 hrs	Growth rate μ (1/day), 72 hrs	Growth rate μ (1/day), 96 hrs
Control	Mean	1.42	1.55	1.59	1.49
	Stdev	0.139	0.0212	0.0187	0.0122
0.061	Mean	1.50	1.59	1.59	1.49
	Stdev	0.179	0.0358	0.011	0.0043
0.20	Mean	1.33	1.52	1.57	1.47
	Stdev	0.0729	0.0129	0.0176	0.0126
0.64	Mean	1.30	1.42	1.46	1.38
	Stdev	0.136	0.037	0.0103	0.0104
2.1	Mean	1.25	1.23	1.08	0.963
	Stdev	0.175	0.009	0.0229	0.0117
7.0	Mean	0.636	0.612	0.483	0.300
	Stdev	0.144	0.0331	0.0591	0.0525
23	Mean	-0.138	-0.265	-0.357	-0.256
	Stdev	0.114	0.0619	0.112	0.111

Table 2: Inhibition of Area Under the Growth Curves

Mean measured concentration	0-24 h		0-48 h		0-72 h		0-96 h
(mg/L)	AUC	IAUC (%)	AUC	IAUC (%)	AUC	IAUC (%)	AUC	IAUC (%)
Control	28.7	0.0	250.3	0.0	1490.9	0.0	5992.1	0.0
0.061	31.1	-11.8	275.1	-9.9	1562.7	-4.8	6171.6	-3.0
0.20	25.3	12.0	233.0*	6.9	1415.6	5.1	5659.7	5.5
0.64	24.4	15.2	196.3*	21.6	1067.1#	28.4	4013.7#	33.0
2.1	23.1	19.5	143.9*	42.5	465.3#	68.8	1107.8#	81.5
7.0	8.2*	71.6	38.2*	84.7	90.2#	93.9	141.9#	97.6
23	-1.1*	104.0	-6.0*	102.4	-15.6#	101.0	-27.0#	100.5

*: Mean value statistically significantly lower than in the control (Williams t-test one-sided smaller, alpha = 0.05)

#: Mean value statistically significantly lower than in teh control (Welch t-test one-sided smaller, alpha = 0.05)

Applicant's summary and conclusion

Validity criteria fulfilled:

yes

Conclusions:

The 72-hour ErC50 and ErC10 values determined in this static test with freshwater green algae (Raphidocelis subcapitata) were 3.6 mg/L and 0.85 mg/L, respectively.

Executive summary:

The effect of the test item on the growth of the freshwater green algal species Raphidocelis subcapitata was investigated under GLP in a 96-hour static test according to the OECD TG 201 (2006, corrected 2011). The nominal test item concentrations tested were 0.10, 0.32, 1.0, 3.2, 10 and 32 mg/L. A control (test water without test item) was tested in parallel. The measured concentrations of the substance in the test media at the start of the test ranged from 72 to 75% of the nominal values and were between 50 and 72% of nominal at the end of the test. Therefore, the biological results were calculated based on the geometric mean measured concentrations of the test item of 0.061, 0.20, 0.64, 2.1, 7.0 and 23 mg/L. The test design included three replicates per test concentration and six replicates of the control. The test was started using a nominal algal cell density of 5000 cells/mL. The test flasks were incubated in a temperature controlled orbital shaker (approximately 125 rpm, Multitron-Pro, Infors HT, Bottmingen/Switzerland) at a temperature maintained at 21 °C. The test flasks were positioned randomly and repositioned daily. They were continuously illuminated by LED lights installed above the test flasks. The test was valid since the performance criteria specified in the test guideline were all met. Furthermore, a parallel study with the positive test substance potassium dichromate demonstrated the suitability and sensitivity of the test system.