Hexachloroethane

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

Study period:

From 30 MARCH 2021 to 12 MAY 2021

Reliability:

2 (reliable with restrictions)

Rationale for reliability incl. deficiencies:

guideline study with acceptable restrictions

Data source

Reference

Reference Type:

study report

Title:

Unnamed

Year:

2021

Materials and methods

Test guideline

Qualifier:

according to guideline

Guideline:

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

Version / remarks:

No analytical monitoring

GLP compliance:

no

Test material

Test material information

impurity 1

Reference substance name:

Tetrachloroethylene

EC Number:

204-825-9

EC Name:

Tetrachloroethylene

Cas Number:

127-18-4

IUPAC Name:

tetrachloroethene

Constituent 1

Reference substance name:

Hexachloroethane

EC Number:

200-666-4

EC Name:

Hexachloroethane

Cas Number:

67-72-1

Molecular formula:

C2Cl6

IUPAC Name:

hexachloroethane

Sampling and analysis

Analytical monitoring:

no

Test organisms

Test organisms (species):

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

Details on test organisms:

TEST ORGANISM

- Strain: CCAP 278/4
-Source: Culture Collection of Algae and Protozoa at Scottish Marine Institute. Successive generations grown in the Ecotoxicity laboratory of TECNALIA R&I.

Study design

Test type:

static

Water media type:

freshwater

Limit test:

no

Total exposure duration:

72 h

Test conditions

Test temperature:

23 +/- 2°c

pH:

At T=0h between 7.82 and 7.96
At t=72h between 8.23 and 9.28

Nominal and measured concentrations:

Preliminary test concentrations: 0.5 mg/l, 5 mg/l and 50 mg/l.
Definitive test concentrations: 0.25 mg/l, 0.5 mg/l, 1 mg/l, 2 mg/l and 4 mg/l

Details on test conditions:

Location: Algae test chamber
Test vessels: 300 ml Erlenmeyer flasks
Volume: 100 ml per flask
Initial cell concentration: 25,000 cells/mL
Type original medium of OECD TG 201 prepared according to OECD Guideline 201.
Photoperiod: continuous light
Light intensity: 4440-8880 lux (mean value of 5903 lux)

Results and discussion

Effect concentrations

Effect concentrations 1

Key result

Duration:

72 h

Dose descriptor:

NOEC

Effect conc.:

< 0.25 mg/L

Nominal / measured:

nominal

Conc. based on:

test mat.

Basis for effect:

growth rate

Effect concentrations 2

Key result

Duration:

72 h

Dose descriptor:

LOEC

Effect conc.:

0.25 mg/L

Nominal / measured:

nominal

Conc. based on:

test mat.

Basis for effect:

growth rate

Effect concentrations 3

Key result

Duration:

72 h

Dose descriptor:

EC50

Effect conc.:

0.88 mg/L

Nominal / measured:

nominal

Conc. based on:

test mat.

Basis for effect:

growth rate

Details on results:

At the end of the established observation period of the definitive test (72 hours), it has been calculated that the concentration that would cause 50% growth inhibition in algae (EC50) would be 0.88 mg/l with a 95% confidence interval of 0.51 mg/l to 1.52 mg/l.
On the other hand, it has been determined that the highest concentration tested that has not produced effect on algae (NOEC) has not been tested and that the lowest that has produced effect (LOEC) has been 0.25 mg/l.

Any other information on results incl. tables

 

The following table shows the average cell densities of all the replicates (expressed in cells per milliliter and in neperian logarithm) of each concentration and the control at 0, 24, 48 and 72 hours. The cellular densities at 0 hours are the theoretical, due to the error of the cell counter at low densities. The densities at 24, 48 and 72 hours have been measured by cell counter.

Concentration (mg/l)	0 h		24 h		48 h		72 h
	Cell density		Cell density		Cell density		Cell density
	cell/ml	ln	cell/ml	ln	cell/ml	ln	cell/ml	ln
Control	2.50.10^4	10.13	1.39.10^5	11.84	5.49.10^5	13.21	2.70.10^6	14.81
0.25	2.50.10^4	10.13	1.11.10^5	11.62	4.17.10^5	12.94	1.17.10^6	13.97
0.5	2.50.10^4	10.13	8.33.10^4	11.33	2.91.10^5	12.58	8.75.10^5	13.68
1	2.50.10^4	10.13	5.56.10^4	10.93	9.72.10^4	11.48	2.08.10^5	12.25
2	2.50.10^4	10.13	4.17.10^4	10.64	5.56.10^4	10.93	6.94.10^4	11.15
4	2.50.10^4	10.13	4.17.10^4	10.64	2.78.10^4	10.23	2.78.10^4	10.23

The results (obtained at 72 hours) of the percentages of cell growth inhibition (%Ir) with respect to the control and the respective standard deviations are presented in the following table. The percentages have been calculated based on the average specific growth rate (μ) from the mean of the three replicates. The average specific growth rate (μ) is the logarithm of the increase in cell density per unit of time.

The concentrations are presented in neperian logarithm (ln) to represent graphically the percentages of inhibition against the logarithm of the concentrations, as indicated in the OECD 201 guide.

Concentration		Ir
mg/l	ln	%	Standard deviation
0.25	-1.39	17.96	1.56
0.5	-0.69	24.21	3.07
1	0.00	55.00	4.31
2	0.69	79.22	8.53
4	1.39	92.71	6.31

 

 

 

 

 

Applicant's summary and conclusion

Validity criteria fulfilled:

not specified

Conclusions:

At the end of the established observation period of the definitive test (72 hours), it has been calculated that the concentration that would cause 50% growth inhibition in algae (EC50) would be 0.88 mg/l with a 95% confidence interval of 0.51 mg/l to 1.52 mg/l.
On the other hand, it has been determined that the highest concentration tested that has not produced effect on algae (NOEC) has not been tested and that the lowest that has produced effect (LOEC) has been 0.25 mg/l.
Based on these results, it can be concluded that the tested sample is very toxic to Pseudokirchneriella subcapitata.

Executive summary:

A laboratory test according to OECD guideline 201 has been carried out to determine the growth inhibition of freshwater algae (Pseudokirchneriella subcapitata) produced by the sample «HEXACHLOROETHANE».

Exponentially growing algae were exposed to the sample for a period of 72 hours to determine its toxic effect by calculating the concentration that produces a 50% inhibition of algal growth (EC50), the highest concentration that produces no effect on the algae (NOEC) and the lowest concentration that produces an effect (LOEC).

Firstly, a preliminary test has been carried out with a wide range of concentrations and, based on the results obtained, the range of concentrations to be tested in the definitive test has been established.

The results obtained in the definitive test indicate that the concentration of the sample «HEXACHLOROETHANE» that causes a 50% inhibition of growth in algae (EC50) would be 0.88 mg/l with a 95% confidence interval of 0.51 mg/l to 1.52 mg/l. The results further indicate that the highest concentration that would produce no effect on algae (NOEC) has not been tested and that the lowest concentration tested that produced an effect on algae (LOEC) has been 0.25 mg/l. Based on these results, it can be concluded that the tested sample is very toxic to P. subcapitata.