Paraquat-dichloride

Toxicity to aquatic algae and cyanobacteria

Link to relevant study record(s)

Reference

Reference 1

Endpoint:

toxicity to aquatic algae and cyanobacteria

Type of information:

experimental study

Adequacy of study:

key study

Study period:

30 Mar 1992 to 3 Apr 1992

Reliability:

1 (reliable without restriction)

Rationale for reliability incl. deficiencies:

guideline study

Qualifier:

according to guideline

Guideline:

OECD Guideline 201 (Alga, Growth Inhibition Test)

Version / remarks:

7 June 1984

Qualifier:

according to guideline

Guideline:

other: EPA 540/09-82-020, Pesticide Assessment Guideline Subdivision J Hazard Evaluation: Non target Plants

Version / remarks:

October 1982.

GLP compliance:

yes

Analytical monitoring:

yes

Details on sampling:

No analyses were made for the concentrations of test substance in the exposure solutions, since the highest nominal test concentration was below the detection limit determined. Therefore, all the results were calculated using nominal test concentration values expressed as μg of technical solution per litre. Analyses were performed on samples taken from the excess of primary and intermediate stock solutions (nominally 51200 μg/L and 102.4 μg/L respectively) prepared, in sterile test culture medium, at the start of the study. 100 mL volumes of these stock solutions were incubated under test conditions and were re-sampled at the end of the test.

Vehicle:

no

Details on test solutions:

PREPARATION OF TEST SOLUTION
- Method: A 1000 mL volume primary stock solution (nominal concentration 51200 μg/L) was prepared by the direct addition of test substance to sterile culture medium. A 2.0 mL aliquot of this was used in the preparation of a 1000 mL intermediate stock solution (nominal concentration 102.4 μg/L), also in sterile culture medium. Aliquots of this intermediate stock solution were used in the preparation of the three highest test substance containing solutions (nominally 1.28, 2.56 and 5.12 μg/L). All remaining test substance containing solutions were similarly prepared using aliquots of the 5.12 μg/L solution.
- Evidence of undissolved material: After preparation, a visual assessment of the test solutions showed them all to be clear and colourless.

Test organisms (species):

Navicula pelliculosa

Details on test organisms:

TEST ORGANISM
- Common name: Green Algae
- Strain: UTEX 667
- Source: laboratory cultures maintained under axenic conditions
- Age of inoculum: A culture of the diatom in the exponential growth phase was used as inoculum for the test.
- Method of cultivation: According to OECD 201

Test type:

static

Water media type:

freshwater

Limit test:

no

Total exposure duration:

96 h

Test temperature:

24.0 - 25.1 °C; the daily temperature measurements recorded (by thermometer reading) in the incubator ranged from 24.0 to 24.2°C. The hourly temperature measurements, recorded automatically, ranged from 24.7 to 25.1°C.

pH:

- At the start of the test: 8.2 to 8.6
- At the end of the test: 7.4 to 7.8

Nominal and measured concentrations:

- Nominal concentrations: 0 (control), 0.040, 0.080, 0.16, 0.32, 0.64, 1.28, 2.56 and 5.12 μg technical solution/L
- Analytical verification of stock solutions: Analysis of the primary and intermediate stock solutions for the test substance gave values of 46000 μg/L (90% of nominal) and 70 μg/L (68% of nominal) respectively. Re-analysis at the end of the test of the 100 mL samples of the primary and intermediate stock solutions incubated under test conditions gave virtually identical values of 46000 μg/L (90% of nominal) and 72 μg/L (70% of nominal) respectively.

Details on test conditions:

TEST SYSTEM
- Test vessel: Borosilicate glass conical flasks of 250 mL nominal capacity in an orbital incubator set at 140 rpm
- Type: closed with polyurethane foam bungs
- Fill volume: 100 mL
- Initial cells density: nominal cell density of 0.300E+04 cells/mL. Three 100 mL volumes of Coulter electrolyte, inoculated in the same manner, had a mean measured cell density of 0.311E+04 cells/mL. This value was used for growth calculations.

GROWTH MEDIUM
- Standard medium used: yes (composition according to guideline OECD 201

TEST MEDIUM / WATER PARAMETERS
Composition according to guideline OECD 201

OTHER TEST CONDITIONS
- Photoperiod: Continuous illumination
- Light intensity and quality: "cool white" light. The light intensity, measured once during the study, was 4,010 Lux (by cosine receptor). This was also measured in terms of quantum response and was 53.5 microeinsteins/square metre/second.

WATER QUALITY MEASUREMENTS
- pH: The pH of each test solution was measured at the start of the test, using the excess remaining after f illing the test vessels. At the end of the test the pH of two of the replicate test solutions (containing th e freshwater diatom) from each control and test concentration was determined. The measurements were made using a Corning Model 240 pH meter.
- Temperature: The temperature of the incubator was measured daily by a thermometer and was continuously monitored, with hourly recording of values, using an automatic recording system linked to a thermistor.

EFFECT PARAMETERS MEASURED; cell density
After 24, 48, 72 and 96 hours, samples were removed from each test and blank vessel. The appropriate blank particle count was subtracted from that of the test culture to obtain the cell density. The algal cell densities of the inoculum and test cultures were determined by electronic particle counting, using a Coulter counter model ZB, counting at a lower threshold equivalent spherical diameter of approximately 2.3 μm.

Reference substance (positive control):

no

Key result

Duration:

96 h

Dose descriptor:

NOEC

Effect conc.:

0.29 µg/L

Nominal / measured:

nominal

Conc. based on:

other: pure test substance

Basis for effect:

growth rate

Remarks on result:

other: recalculated value, expressed as pure substance, see ‘any other information on results incl. tables’ for respective calculation

Duration:

96 h

Dose descriptor:

NOEC

Effect conc.:

0.64 µg/L

Nominal / measured:

nominal

Conc. based on:

test mat.

Basis for effect:

growth rate

Remarks on result:

other: Original value presented in study

Key result

Duration:

96 h

Dose descriptor:

EC50

Effect conc.:

0.47 µg/L

Nominal / measured:

nominal

Conc. based on:

other: pure test substance

Basis for effect:

growth rate

Remarks on result:

other: recalculated value, expressed as pure substance, see ‘any other information on results incl. tables’ for respective calculation

Duration:

96 h

Dose descriptor:

EC50

Effect conc.:

1.03 µg/L

Nominal / measured:

nominal

Conc. based on:

test mat.

Basis for effect:

growth rate

Remarks on result:

other: 95% confidence limits: 0.72 to 1.47 μg/L

Remarks:

Original value presented in study

Duration:

96 h

Dose descriptor:

LOEC

Effect conc.:

1.28 µg/L

Nominal / measured:

nominal

Conc. based on:

test mat.

Basis for effect:

growth rate

Remarks on result:

other: Original value presented in study

Duration:

96 h

Dose descriptor:

NOEC

Effect conc.:

0.64 µg/L

Nominal / measured:

nominal

Conc. based on:

test mat.

Basis for effect:

biomass

Remarks:

(as Area Under the Growth curve (AUG))

Remarks on result:

other: Original value presented in study

Duration:

96 h

Dose descriptor:

EC50

Effect conc.:

0.69 µg/L

Nominal / measured:

nominal

Conc. based on:

test mat.

Basis for effect:

biomass

Remarks:

(as Area Under the Growth curve (AUG))

Remarks on result:

other: 95% confidence limits: 0.51 to 0.96 μg/L

Remarks:

Original value presented in study

Duration:

96 h

Dose descriptor:

LOEC

Effect conc.:

1.28 µg/L

Nominal / measured:

nominal

Conc. based on:

test mat.

Basis for effect:

biomass

Remarks:

(as Area Under the Growth curve (AUG))

Remarks on result:

other: Original value presented in study

Details on results:

Inhibition of growth rate and biomass was observed from concentrations of 1.28 μg/L and above. For more details see 'Any other information on results incl. tables'.

Reported statistics and error estimates:

Areas under the growth curve and growth rates were examined by one-way analysis of variance, and Dunnett's procedure was used to identify significant differences (P=0.05) from the control.

Table 1: Mean algal density (1E+04 cells/ mL)

Concentration of technical solution (μg/L)	Basis	Day 1	Day 2	Day 3	Day 4
Control	Mean (n=6)	0.295	0.555	2.55	10.1
0.040	Mean (n=3)	0.285	0.528	2.06	12.2
0.080	Mean (n=3)	0.296	0.785	3.43	14.7
0.16	Mean (n=3)	0.270	0.536	2.02	13.2
0.32	Mean (n=3)	0.252	0.575	2.65	18.1
0.64	Mean (n=3)	0.217	0.410	0.905	4.63
1.28	Mean (n=3)	0.341	0.501	0.845	2.13
2.56	Mean (n=3)	0.174	0.149	0.199	0.265
5.12	Mean (n=3)	0.226	0.183	0.477	0.371

Table 2: Mean area under the growth curve

Concentration of technical solution (μg/L)	Mean area under the growth curve (0 -96 hr)	% of control
Control	7.4	-
0.040	7.9	106
0.080	10.8	146
0.16	8.4	113
0.32	11.4	155
0.64	2.8	37
1.28	1.7*	22
2.56	-0.4*	-6
5.12	0.0*	0

*Significant difference from the control, (P=0.05)

Table 3: Mean growth rate

Concentration of technical solution (μg/L)	Mean growth rate (0-96 hr)	% of control
Control	0.871	-
0.040	0.917	105
0.080	0.963	111
0.16	0.938	108
0.32	1.016	117
0.64	0.675	78
1.28	0.180*	55
2.56	-0.040*	-5
5.12	0.044*	5

*Significant difference from the control, (P=0.05)

Calculation of key result

The doses of the test substance were expressed in technical test material, which relates to an aqueous solution of the registered substance (recalculated purity = 45.2%, purity not reported in the study report). The effect values are calculated by correction for the amount of water:

- ErC50: 0.452 x 1.03 µg technical test material/L = 0.47 µg pure test substance/L.

- NOErC: 0.452 x 0.64 µg technical test material = 0.29 µg pure test substance/L.

Validity criteria fulfilled:

yes

Conclusions:

The 96-h ErC50 and 96-h NOErC values are 1.03 and 0.64 µg technical solution/L, respectively in freshwater green algae (Navicula pelliculosa). These values correspond to recalculated values of 0.47 and 0.29 µg pure substance/L.

Executive summary:

The toxicity to aquatic alga was determined in a study according to OECD TG 201 and incompliance with GLP criteria. In this study, cultures of freshwater green algae (Navicula pelliculosa) at an initial concentration of ca. 0.3E+04 cells/mL were exposed to the test substance at nominal concentrations 0 (medium control), 0.040, 0.080, 0.16, 0.32, 0.64, 1.28, 2.56 and 5.12 µg a.i./L for 96 hours. No analyses were made for the concentrations of test substance in the exposure solutions, since the highest nominal test concentration was below the detection limit determined. Therefore, all the results were calculated using nominal test concentration values expressed as μg technical solution/L. Instead, analyses were performed on samples taken from the excess of primary and intermediate stock solutions (nominally 51200 μg/L and 102.4 μg/ respectively) prepared, in sterile test culture medium, at the start and re-sampled at the end of the test. These analysis gave values corresponding to 90% and 68% of nominal concentrations at the start of the experiment, respectively. Re-analysis at the end of the test gave values of 90% and 70% of the nominal concentration for these samples. Cell densities were measured throughout the test and growth rate and biomass (as Areas Under the Curve) determined. Significant inhibition of biomass was observed at concentrations of 1.28, 2.56 and 5.12 µg/L (22, -6 and 0% of control biomass, respectively). For growth rate significant inhibition was observed at these concentrations as well (55, -5 and 5% of growth control, respectively). Based on these findings, the 96-h EC50 values for growth rate and biomass were determined 1.03 and 0.69 µg/L, respectively. The 96-h NOEC was determined to be 0.64 µg/L for both growth rate and biomass. The recalculated 96H ErC50 and 96-h NOErC values are 0.47 and 0.29 µg pure substance/L.

Reference 2

Endpoint:

toxicity to aquatic algae and cyanobacteria

Type of information:

experimental study

Adequacy of study:

key study

Reliability:

1 (reliable without restriction)

Rationale for reliability incl. deficiencies:

guideline study

Qualifier:

equivalent or similar to guideline

Guideline:

EPA OPP 123-3 (Algal Toxicity, Tiers I and II)

Version / remarks:

described as FIFRA 123-2 Guideline, 1982

Deviations:

no

Qualifier:

according to guideline

Guideline:

other: US EPA (1982). Pesticide Assessment Guideline Subdivision J Hazard Evaluation: Non target Plants. EPA 540/09-82-020, October 1982.

Version / remarks:

1982

Deviations:

no

Qualifier:

according to guideline

Guideline:

OECD Guideline 201 (Alga, Growth Inhibition Test)

Version / remarks:

1984

Deviations:

no

GLP compliance:

yes

Analytical monitoring:

yes

Vehicle:

no

Details on test solutions:

A 1600 mL volume of the highest nominal concentration test solution (32 mg/L) was prepared by the direct addition of test substance to sterile culture medium. All lower test concentrations were prepared using sterile medium, to a final volume of 1000 mL, by the addition of aliquots of the nominal 32 mg/L solution..

Test organisms (species):

Skeletonema costatum

Details on test organisms:

- Test species/strain: Marine alga, Skeletonema costatum

Test type:

static

Water media type:

saltwater

Limit test:

no

Total exposure duration:

96 h

Hardness:

Not reported

Test temperature:

Daily measurements recorded by thermometer reading in the incubator: 19.8 to 20.7 °C
Automatically recorded hourly measurements: 18.9 to 20.1 °C

pH:

Start: 8.3 to 8.4
End: 8.2 to 8.9

Dissolved oxygen:

Not reported

Salinity:

The salinity of the test culture medium, measured post filter sterilisation was 30.9‰

Conductivity:

Not reported

Nominal and measured concentrations:

Nominal: 0 (control) and nominal concentrations 0.25, 0.50, 1.0, 2.0, 4.0, 8.0, 16 and 32 mg of test substance/L.
Measured, mean of concentrations at exposure time 0 and 96 hours: 0, 0.22, 0.47, 0.85, 2.1, 4.3, 7.7, 16, 32 mg/L

Details on test conditions:

- Experimental design: Six replicate cultures of the control, and triplicate cultures at each concentration of the test substance were employed. One blank vessel (without algal inoculum) for each control and treatment was incubated concurrently. The positions of the vessels in the incubator were re-randomised, daily, by rows.
- Inoculum: The cell density of the culture used as the inoculum for the test was determined by microscope counting using a haemocytometer chamber and was 0.835 x 106 cells/mL.
- Particle density determinations: the algal particle densities of the inoculum and test cultures were determined by electronic particle counting, using a Coulter counter model ZB, counting at a lower threshold equivalent spherical diameter of approximately 2.9 µm.
After 24, 48, 72 and 96 hours, samples were removed from each test and blank vessel. The appropriate blank particle count was subtracted from that of the test culture to obtain the particle density.

Reference substance (positive control):

no

Key result

Duration:

96 h

Dose descriptor:

NOEC

Effect conc.:

0.21 mg/L

Nominal / measured:

nominal

Conc. based on:

other: pure test substance

Basis for effect:

growth rate

Remarks on result:

other: recalculated value, expressed as pure substance, see 'Any other information on results incl. tables' for respective calculation.

Duration:

96 h

Dose descriptor:

NOEC

Effect conc.:

0.47 mg/L

Nominal / measured:

nominal

Conc. based on:

test mat.

Basis for effect:

growth rate

Remarks on result:

other: Original value as presented in the study report

Duration:

96 h

Dose descriptor:

EC50

Effect conc.:

8.13 mg/L

Nominal / measured:

nominal

Conc. based on:

other: pure test substance

Basis for effect:

growth rate

Remarks on result:

other: recalculated value, expressed as pure substance, see 'Any other information on results incl. tables' for respective calculation.

Duration:

96 h

Dose descriptor:

EC50

Effect conc.:

18 mg/L

Nominal / measured:

nominal

Conc. based on:

test mat.

Basis for effect:

growth rate

Remarks on result:

other: Original value as presented in the study report

Results based onconcentrations as mg of test material per litre:

Based on areas under the growth curve the results obtained mean of determined were:

- No observed effect (P=0.05) concentration (NOEC) = 0.22 mg/L

- Lowest significant effect (P=0.05) concentration (LOEC) = 0.47 mg/L

- Median effective concentration, biomass (EbC50) = 2.2 mg/L; 95% confidence limits = 0.41 to 8.6 mg/L.

 

Based on logarithmic growth rate over the test period, the results obtained were:

- No observed effect (P=0.05) concentration (NOEC) = 0.47 mg/L

- Lowest significant effect (P=0.05) concentration (LOEC) = 0.85 mg/L

- Median effective concentration, biomass (EbC50) = 18 mg/L; 95% confidence limits = 0.27 to >32 mg/L.

  

Physical data

- At the start of the test the pH ranged from 8.3 to 8.4 and at the end of the test the range was from 8.2 to 8.9

- The daily temperature measurements recorded (by thermometer reading) in the incubator ranged from 19.8 to 20.7°C.

- The light intensity, measured once during the study, was 4800 Lux (by cosine receptor). This was also measured in terms of quantum response and was 63.9 microeinsteins/m2/sec.

 

Analytical results for the concentrations of the test solutions

Nominal conc. of test item	Exposure time (h)	Conc. Determined (mg/L)	% of nominal conc.
Control	0	<0.046	-
	96	<0.048	-
	mean	<0.048	-
0.25	0	0.17	68
	96	0.26	104
	mean	0.22	88
0.5	0	0.46	92
	96	0.48	96
	mean	0.47	94
1	0	0.87	87
	96	0.82	82
	mean	0.85	85
2	0	2.1	105
	96	2	100
	mean	2.1	105
4	0	4.2	105
	96	4.3	18
	mean	4.3	108
8	0	7.6	95
	96	7.7	96
	mean	7.7	96
16	0	15	94
	96	17	106
	mean	16	100
32	0	32	100
	96	32	100
	mean	32	100

Calculation of key result

The doses of the test substance were expressed in technical test material, which relates to an aqueous solution of the registered substance. The effect values are calculated by correction for the amount of water:

- Purity of the test item: (100/72.4 * 32.7) % = 45.16 %

- EC50: 0.4516 x 18 mg technical test material/L = 8.13 mg pure test substance/L.

- NOEC: 0.4516 x 0.47 mg technical test material/L = 0.21 mg pure test substance/L.

Validity criteria fulfilled:

not specified

Conclusions:

On a FIFRA 123-2 study on marine alga (skeletoma costatum), based on measured concentrations, and on the logarithmic growth rate over the test period, the results obtained were: NOEC = 0.47 mg/L and ErC50 = 18 mg/L. After recalculating the effect values, ErC50 is equal to 8.13 mg pure test substance/L and the NOEC is equal to 0.21 mg pure test substance/L.

Executive summary:

The toxicity to aquatic algae was determined in a study according FIFRA 123-2 and in compliance with GLP criteria. In this study, the test item was tested on the marine alga Skeletonema costatum were exposed to the test substance at nominal concentrations 0 (control) and nominal concentrations of 0.25, 0.50, 1.0, 2.0, 4.0, 8.0, 16 and 32 mg of test substance/L for 96 hours. Test concentrations were analytical verified at the start and the end of the test. The determined concentrations at the start of the test ranged from 68 to 105% of the nominal values. The determined concentrations in the blanks after 96 hours ranged from 82 to 108% of the nominal values (based on test substance as received). The mean determined concentrations for the nominal concentrations of 0.25, 0.50, 1.0, 2.0, 4.0, 8.0, 16 and 32 mg of paraquat dichloride technical per litre were 0.22, 0.47, 0.85, 2.1, 4.3, 7.7, 16 and 32 mg per litre respectively. Cell densities were measured throughout the test and growth rate and biomass (as Areas Under the Curve) determined. No significant (dose-response) inhibition in growth rate or biomass was observed in all treatments throughout the test. Based on areas under the growth curve the results obtained mean of determined were: (NOEbC)= 0.22 mg/L and EbC50 = 2.2 mg/L. Based on logarithmic growth rate over the test period, the results obtained were: NOErC =0.47 mg/L and ErC50 = 18 mg/L. After recalculating the effect values, ErC50 is equal to 8.13 mg pure test substance/L and the NOErC is equal to 0.21 mg pure test substance/L.

Description of key information

The NOEC and EC50 for growth rate were determined to be 0.29 and 0.47 µg pure substance/L (recalculated), respectively, in freshwater algae (Navicula pelliculosa), OECD 201, Smyth 1992

The NOEC and EC50 for growth rate were determined to be 0.21 und 8.13 mg pure substance/L (recalculated), respectively, in marine algae (Skeletonema costatum), FIFRA 123 -2, Smyth 1992

Key value for chemical safety assessment

EC50 for freshwater algae:

0.47 µg/L

EC50 for marine water algae:

8.13 mg/L

EC10 or NOEC for freshwater algae:

0.29 µg/L

EC10 or NOEC for marine water algae:

0.21 mg/L

Additional information

Table: Overview of aquatic algae toxicity data available for the substance:

Species	Guideline/ GLP	Endpoint	Effect value	Comment	Reference
Navicula pelliculosa (freshwater)	OECD 201/ GLP	96-h ErC50 96-h NOErC	0.47 µg/L 0.29 µg/L	Based on nominal concentrations and growth rates	Smyth 1992
Skeletonema costatum (saltwater)	FIFRA 123-2	96-h ErC50 96-h NOErC	8.13 mg/L 0.21 mg/L	Based on nominal concentrations and growth rates	Smyth 1992