Bis(acetato-O)hydroxyaluminium

Toxicity to aquatic algae and cyanobacteria

Link to relevant study record(s)

Reference

Endpoint:

toxicity to aquatic algae and cyanobacteria

Type of information:

experimental study

Adequacy of study:

key study

Study period:

2017-08-29 to 2017-09-02

Reliability:

1 (reliable without restriction)

Rationale for reliability incl. deficiencies:

guideline study

Qualifier:

according to guideline

Guideline:

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

Deviations:

no

Qualifier:

according to guideline

Guideline:

EPA OPPTS 850.5400 (Algal Toxicity, Tiers I and II) (January 2012)

Deviations:

no

Qualifier:

according to guideline

Guideline:

EU Method C.3 (Algal Inhibition test)

Deviations:

no

GLP compliance:

yes (incl. QA statement)

Analytical monitoring:

yes

Details on sampling:

Five test concentrations in a geometric series (with a spacing factor of 2) as a dilution series of a saturated stock solution were investigated in the main test and analysed by ICP-OES method at the start and at the end of the test. A volume of 50 mL of the sample was concentrated to dryness by evaporation of water. Then it was reconstituted with 10 mL of 2% nitric acid and analysed by the ICP-OES method.
At the start of the test one replicate of the control (60 mL), on replicate of the test item concentration 6.25, 12.5 and 25 mg/L (50 mL) and three replicates of the test item concentration 50 an 100 mg/L were analysed. At the end (72 hours) three replicates of each test concentration and control, 60 mL each, were analysed.

Vehicle:

no

Details on test solutions:

The test solutions used in the test was prepared by mechanical dispersion without using any solubilising agents. The test item is poorly water soluble material. For preparation of the test solutions a supersaturated solution (100 mg/L nominal loading) was first prepared by dispersing/dissolving an amount of 0.1 g test item into 1000 mL test medium (OECD medium). This solution was agitated by orbital shaker overnight and then the non-dissolved test material was separated by filtration through a membrane filter (0.22 µm) in order to obtain the saturated test solution (i.e. 100 % v/v saturated solution). The pH of this stock solution was adjusted to that of the culture medium using 1N NaOH. The test solutions of subsequent lower test concentrations were prepared by appropriate diluting of this stock solution.

Test organisms (species):

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

Details on test organisms:

TEST ORGANISM
- Common name: Raphidocelis subcapitata (Printz-Starr)
- Strain: 61.81 SAG (identical strains: CCAP 278/4; UTEX 1648; ATCC 22662)
- Source: SAG: Collection of Algal Cultures, Inst. Plant Physiology, University of Göttingen, Untere Karspüle 2, D-37073 Göttingen, Germany
- Method of cultivation: Small algal cultures that are regularly planted on agar are transferred to fresh medium at least once every two months under standardised conditions according to the test guidelines. Pre-culture was prepared with Algal Mineral Salts Culture Medium, incubated under the conditions of the test and used when still exponentially growing, normally after an incubation period of 2-4 days.

Test type:

static

Water media type:

freshwater

Limit test:

no

Total exposure duration:

72 h

Hardness:

not specified

Test temperature:

The temperature in the flask was in the range of 22.3-22.8 °C and within the climate chamber between 22.2 and 23.6 °C.

pH:

7.52 - 8.67 (Control: 8.30 - 8.67)

Dissolved oxygen:

not specified

Salinity:

not applicable

Conductivity:

not specified

Nominal and measured concentrations:

The following nominal concentrations were tested: 6.25, 12.5, 25, 50 and 100 % v/v saturated solution. The corresponding calculated geometric mean concentrations were: 0.34, 0.42, 0.76, 0.84 and 1.30 mg/L.

Details on test conditions:

TEST SYSTEM
- Test vessel: 250 mL Erlenmeyer flask
- Type: covered with air-permeable stoppers
- Medium volume per flask: 100 mL
- Initial cells density: 10^4 cells/mL
- Control end cells density: 104×10^4 cells/mL (mean value)
- No. of vessels per concentration (replicates): 3
- No. of vessels per control (replicates): 6

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

OTHER TEST CONDITIONS
- Adjustment of pH: pH of stock solution was adjusted to that of the culture medium using 1N NaOH.
- Photoperiod: continuously
- Light intensity and quality: 8011 lux, fluorescent lamps

EFFECT PARAMETERS MEASURED:
- Determination of cell concentrations: counting chamber
- Morphological Changes of Algal Cells: microscopy

TEST CONCENTRATIONS
- Spacing factor for test concentrations: 2
- Range finding study: yes
- Test concentrations: 0.1, 1, 10 and 100 mg/L plus a control
- Results used to determine the conditions for the definitive study: 32.7% growth inhibition at the highest concentration, no growth inhibition at 10 mg/L

Reference substance (positive control):

yes

Remarks:

Potassium dichromate, test period: 07–10 March 2017

Duration:

72 h

Dose descriptor:

EC50

Effect conc.:

1.02 mg/L

Nominal / measured:

meas. (geom. mean)

Conc. based on:

test mat.

Basis for effect:

growth rate

Key result

Duration:

72 h

Dose descriptor:

EC50

Effect conc.:

0.92 mg/L

Nominal / measured:

meas. (geom. mean)

Conc. based on:

other: calculated value for anhydrous substance

Basis for effect:

growth rate

Duration:

72 h

Dose descriptor:

EC50

Effect conc.:

0.9 mg/L

Nominal / measured:

meas. (geom. mean)

Conc. based on:

test mat.

Basis for effect:

biomass

Duration:

72 h

Dose descriptor:

NOEC

Effect conc.:

0.76 mg/L

Nominal / measured:

meas. (geom. mean)

Conc. based on:

test mat.

Basis for effect:

growth rate

Duration:

72 h

Dose descriptor:

NOEC

Effect conc.:

0.76 mg/L

Nominal / measured:

meas. (geom. mean)

Conc. based on:

test mat.

Basis for effect:

biomass

Duration:

72 h

Dose descriptor:

LOEC

Effect conc.:

0.84 mg/L

Nominal / measured:

meas. (geom. mean)

Conc. based on:

test mat.

Basis for effect:

growth rate

Duration:

72 h

Dose descriptor:

LOEC

Effect conc.:

0.84 mg/L

Nominal / measured:

meas. (geom. mean)

Conc. based on:

test mat.

Basis for effect:

biomass

Duration:

72 h

Dose descriptor:

EC10

Effect conc.:

0.73 mg/L

Nominal / measured:

meas. (geom. mean)

Conc. based on:

test mat.

Basis for effect:

growth rate

Key result

Duration:

72 h

Dose descriptor:

EC10

Effect conc.:

0.66 mg/L

Nominal / measured:

meas. (geom. mean)

Conc. based on:

other: calculated value for anhydrous substance

Basis for effect:

growth rate

Duration:

72 h

Dose descriptor:

EC10

Effect conc.:

0.44 mg/L

Nominal / measured:

meas. (geom. mean)

Conc. based on:

test mat.

Basis for effect:

biomass

Details on results:

No morphological abnormalities were observed in the control and in the lowest concentration of 0.34 mg/L (measured) during the test. Some swollen cells were observed together with the normal cells in the concentrations of 0.42, 0.76 and 0.84 mg/L (measured) at the 48-h and 72-h observation period. In the highest concentration of 1.30 mg/L (measured) shape of algal cells could not be evaluated due to the absence of cells. The cell density in the control has increased from 1×10^4 cells/mL at the start of the test (0 hours) to 94.0×10^4 cells/mL (mean value) after 72 hours in the in the control.

Results with reference substance (positive control):

The 72h ErC50: 0.99 mg/L (95 % confidence limits: 0.68 – 1.49 mg/L). The 72h EyC50: 0.59 mg/L (95 % confidence limits: 0.47 – 0.75 mg/L)

Reported statistics and error estimates:

For the determination of the LOEC and NOEC, ANOVA and Dunnett’s test (α = 0.05; 2-sided) was used and for the determination of the ECx values Probit analysis was used.

Validity criteria fulfilled:

yes

Conclusions:

In this 72-h algal growth inhibition test the effects of Aluminium hydroxide diacetate hydrate on the growth of Raphidocelis subcapitata was determined. The 72-h EC10, EC20, EC50 for growth were determined to be 0.73, 0.83 and 1.02 mg/L (hydrated substance), and 0.66, 0,75 and 0.92 mg/L (calculated for anhydrous substance), respectively. The 72-h EC10, EC20, EC50 for yield were determined to be 0.44, 0.60 and 0.90 mg/L (hydrated substance), and 0.40, 0.54 and 0.81 mg/L (calculated for anhydrous substance), respectively. The 72-h NOEC for growth and yield was determined to be 0.76 mg/L (hydrated substance) and 0.68 mg/L (calculated for anhydrous substance). The results are based on measured geometric mean test item concentrations.

Executive summary:

The purpose of this study was to determine the effect of the test item Aluminium hydroxide diacetate hydrate on the growth of a unicellular green algal species Raphidocelis subcapitata. Exponentially growing cultures of Raphidocelis subcapitata were exposed to test item concentrations of 6.25, 12.5, 25, 50 and 100 % v/v saturated solution (corresponding to 0.34, 0.42, 0.76, 0.84 and 1.30 mg/L (calculated geometric mean measured concentrations)). The algal growth in relation to a control culture was determined over a fixed test period of 72 hours. The 72-h EC10, EC20, EC50 for growth were determined to be 0.73, 0.83 and 1.02 mg/L (hydrated substance), and 0.66, 0.75 and 0.92 mg/L (calculated for anhydrous substance), respectively. The 72-h EC10, EC20, EC50 for yield were determined to be 0.44, 0.60 and 0.90 mg/L (hydrated substance), and 0.40, 0.54 and 0.81 mg/L (calculated for anhydrous substance), respectively. The 72-h NOEC for growth and yield was determined to be 0.76 mg/L (hydrated substance) and 0.68 mg/L (calculated for anhydrous substance). The results are based on measured geometric mean test item concentrations.

Description of key information

The effect of the test item on the growth of a unicellular green algal species Raphidocelis subcapitata was determined according to OECD 201, Regulation (EC) No. 226/2016 Method C.3 and EPA OPPTS 850.5400 and under GLP (reference 6.1.5-1). The study was conducted under static conditions with an initial cell density of 1x10^4 cells/mL for a period of 72 hours. The EC50 and EC10 based on growth rate were determined to be 1.02 mg/L and 0.73 mg/L (hydrated substance, mean measured), and 0.92 mg/L and 0.66 mg/L (calculated value for anhydrous substance), respectively.

Key value for chemical safety assessment

EC50 for freshwater algae:

0.92 mg/L

EC10 or NOEC for freshwater algae:

0.66 mg/L

Additional information

The effect of the test item on the growth of a unicellular green algal species Raphidocelis subcapitata was determined according to OECD 201, Regulation (EC) No. 226/2016 Method C.3 and EPA OPPTS 850.5400 and under GLP (reference 6.1.5-1). The study was conducted under static conditions with an initial cell density of 1x10^4 cells/mL for a period of 72 hours. A stock solution of 100 mg/L was freshly prepared with OECD medium and shaken overnight and the non-dissolved test material was separated by filtration. Six concentrations were tested in a geometrical series with a dilution factor of 2: 6.25, 12.5, 25, 50 and 100 % v/v saturated solution (corresponding to 0.34, 0.42, 0.76, 0.84 and 1.30 mg/L (calculated geometric mean concentrations)).The EC50 values for inhibition of growth rate (ErC50) was 1.02 mg/L (hydrated substance) and 0.92 mg/L (calculated value for anhydrous substance) and for inhibition of yield (EyC50) 0.90 mg/L (hydrated substance) and 0.81 mg/L (calculated value for anhydrous substance). The EC10 value was determined to be 0.73 mg/L (hydrated substance) and 0.66 mg/L (calculated value for anhydrous substance) based on growth rate (ErC10) and 0.44 mg/L (hydrated substance) and 0.40 mg/L (calculated value for anhydrous substance) based on yield (EyC10).