TETRASODIUM ETHYLENE 1,1-DIPHOSPHONATE

Toxicity to aquatic algae and cyanobacteria

Administrative data

Endpoint:

toxicity to aquatic algae and cyanobacteria

Type of information:

experimental study

Adequacy of study:

key study

Study period:

2002-05-20 - 2002-07-05

Reliability:

1 (reliable without restriction)

Rationale for reliability incl. deficiencies:

guideline study

Data source

Reference

Reference Type:

study report

Title:

Unnamed

Year:

2002

Report date:

2002

Materials and methods

Test guideline

Test guideline 1

Qualifier:

according to guideline

Guideline:

OECD Guideline 201 (Alga, Growth Inhibition Test)

Test guideline 2

Qualifier:

according to guideline

Guideline:

EU Method C.3 (Algal Inhibition test)

GLP compliance:

yes (incl. QA statement)

Remarks:

2000-04-26

Test material

Test material information

Constituent 1

Reference substance name:

-

EC Number:

443-520-7

EC Name:

-

Cas Number:

33016-77-2

Molecular formula:

C2H4Na4O6P2

IUPAC Name:

tetrasodium (1-phosphonatoethenyl)phosphonate

Test material form:

solid

Specific details on test material used for the study:

- Synonyms: ITC 908, vinylidenephosphonic acid tetrasodium salt (VDPA tetrasodium salt)
- Lot/batch No.of test material: 562CH/646
- Purity: 87.33%
- Storage condition of test material: Room temperature, in the dark

Sampling and analysis

Analytical monitoring:

yes

Details on sampling:

The concentration and stability of the test material in the test preparations were verified by chemical analysis at 0 and 96 hours.

Test solutions

Vehicle:

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: Liquid cultures of P. subcapitata were obtained from the Culture Collection of Algae and Protozoa (CCAP), Institute of Freshwater Ecology, The Ferry House, Far Sawrey, Ambleside, Cumbria.

ACCLIMATION
- Culturing media and conditions: The culture was maintained in the laboratory at a temperature of 21 °C under continunous illumination (intensity approximately 7000 lux) and constant aeration. The culture medium used is the same as that used in the range-finding and definitive tests.

Study design

Test type:

static

Water media type:

freshwater

Limit test:

no

Total exposure duration:

96 h

Test conditions

Test temperature:

24 °C

pH:

7.5 - 10.3 (0h)
7.8 - 9.3 (96h)

Nominal and measured concentrations:

Range-finding test: 1.0, 10 and 100 mg/L
Definitive test: 10, 20, 40, 80 and 160 mg/L

Details on test conditions:

TEST SYSTEM
- Test vessel: Glass conical flasks
- Type: closed
- Fill volume: 250 ml
- Initial cells density: 10E4 cells per ml
- Control end cells density: 3.82x10E6 cells per ml (96h)
- No. of vessels per concentration (replicates): 3
- No. of vessels per control (replicates): 3

GROWTH MEDIUM
- Standard medium used: yes

OTHER TEST CONDITIONS
- Adjustment of pH: No
- Photoperiod: Continuous illumination
- Light intensity and quality: approximately 7000 lux

EFFECT PARAMETERS MEASURED (with observation intervals if applicable) :
- Determination of cell concentrations: Coulter Multisizer II Particle Counter; 0 and 96h in the range-finding test; 0, 24, 48, 72 and 96h in the definitive test

TEST CONCENTRATIONS
Range finding study
- Test concentrations: 1.0, 10 and 100 mg/L
- Results used to determine the conditions for the definitive study: The results showed no effect on growth at the test concentrations of 1.0 and 10 mg/L. However, the growth was observed to be reduced at 100 mg/L.

Reference substance (positive control):

yes

Remarks:

Zinc chloride: 0.050, 0.10, 0.20, 0.40 and 0.80 mg/L

Results and discussion

Effect concentrations

Effect concentrations 1

Duration:

96 h

Dose descriptor:

EC50

Effect conc.:

220 mg/L

Nominal / measured:

meas. (geom. mean)

Conc. based on:

test mat.

Basis for effect:

growth rate

Effect concentrations 2

Duration:

96 h

Dose descriptor:

EC10

Effect conc.:

29 mg/L

Nominal / measured:

meas. (geom. mean)

Conc. based on:

test mat.

Basis for effect:

growth rate

Details on results:

There were no abnormalities detected in any of the control or test cultures.

A re-growth experiment was performed after 96 hours exposure to the test material to determine the algicidal or algistatic effect of the test material. Re-growth occurred in the control, 10 and 20 mg/L test cultures after 48 hours, in the 40 and 80 mg/L test cultures after 72 hours and in the 160 mg/L test culture after 96 hours. These results indicated that the test material was algistatic in effect.

Analysis of the test preparations at 0 hours showed measured test concentrations to range from 80 to 107% of nominal. Analysis of the test preparations at 96 hours showed measured test concentrations to be near nominal with the exception of the 10 mg/L test concentration which showed a slight decline in measured test concentrations to 73% of nominal. This decline in measured test concentration in the 10 mg/L test group was in line with both the preliminary recovery analyses conducted which indicated that the test material was possibly unstable in culture medium over the 96-hour test period at the bottom concentration. Adsorption was not a factor in the preliminary stability analysiss since no algal cells were present.
Given this decline in measured test concentrations it was considered justifiable to base the results on the mean measured test concentrations.

The coefficient of variation in the daily growth rates of the control cultures was 21% and hence satisfied the validation criterion whereby the coefficient of variation must not exceed 35%
The coefficient of variation of average growth rate in replicate control and test cultures was in the range 0% to 7% and hence satisfied the validation criterion whereby the coefficient of variation must not exceed 15%.

Results with reference substance (positive control):

EbC50 (72h): 0.18 mg/L
EbC50 (96h): 0.16 mg/L
ErC50 (96h): 0.26 mg/L
NOEC: 0.050 mg/L

Reported statistics and error estimates:

One way analysis of variance incorporating Bartlett's test for homogeneity of variance and Dunnett's multiple comparison procedure for comparing several treatments with a control test was carried out on the area under the growth curve data at 96 hours for the control and all test concentrations to determine any statistically significant differences between the test and control groups. All statistical analyses were performed using the SAS computer software package.

Applicant's summary and conclusion

Validity criteria fulfilled:

yes

Conclusions:

The effect of the test material on the growth of Pseudokirchneriella subcapitata has been investigated over a 96-hour period and gave an ErC50 (96h) value of 220 mg/L and an ErC10 (96h) value of 29 mg/L.

Executive summary:

The effect of the test material on the growth of Pseudokirchneriella subcapitata has been investigated following OECD TG 201 and under GLP conditions over a 96-hour period and gave an ErC50 (96h) value of 220 mg/L and an ErC10 (96h) value of 29 mg/L.