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Toxicity to aquatic algae and cyanobacteria

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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:
according to guideline
Guideline:
OECD Guideline 201 (Alga, Growth Inhibition Test)
Deviations:
yes
Remarks:
A modified ISO growth medium was used as a medium due to chelating agents found in the standard media that can interfere with the test substance
Principles of method if other than guideline:
Culture medium used was a modified ISO growth medium prepared as suggested in the difficult substances guidance document. The deviation from the guideline was required due to chelating agents found in the standard media that can interfere with the test substance.
GLP compliance:
yes
Analytical monitoring:
yes
Details on sampling:
Concentration of gold in all test concentrations and the control determined at 0 and 72 hours for the definitive test.At the start of the test samples were taken from excess test solutions and at the end of the test a sample was taken from a single test replicate solution.
Vehicle:
no
Details on test solutions:
Definitve test: The purity of the aqueous solution was corrected for during preparation of the test solutions A stock solution of 100 mg/L was prepared and stirred for 15 minutes, resulting in a clear and pale yelow solution. Test solutions were prepared by stock solution to culture medium. Solutions were clear and colourless.
Test organisms (species):
Pseudokirchneriella subcapitata (previous names: Raphidocelis subcapitata, Selenastrum capricornutum)
Details on test organisms:
Pseudokirchneriella subcapitata strain CCAP 278/4 from laboratory cultures maintained under axenic conditions. Algae in the exponential growth phase used as the inoculum in the range finder and definitive tests. The culture used in the definitve test was grown under the environmental conditions and in the media from the test.
Test type:
static
Water media type:
freshwater
Limit test:
no
Total exposure duration:
72 h
Test temperature:
22 +- 2°C
pH:
Range finder test: start of test 7.42 - 8.01; end of test 8.24 - 8.35
Definitive test: start of test 7.81 - 8.10; end of test 7.99 - 8.08
Nominal and measured concentrations:
Range finder: Control, 0.001, 0.01, 0.1, 1.0 and 10 mg/L (nominal)
Definitive: Control, 0.095, 0.31, 0.98, 3.1 and 10 mg/L (nominal)
Details on test conditions:
Range finder test: Duplicate cultures of each test concentration and triplicate cultures of the control were used. One blank ( no algae) was used per test concentration and control. Algal cell densities were determined by electronic particle counting. After 72 hours samples from the test concentration and blank were taken and the blank particle count subtracted from the test culture to obtain the cell density.

Definitive test: Six replicate cultures of the control and triplicate cultures of each test concentration were used. Test vessel position within the incubator was randomised daily. One blank was set up per test concentration and control. Algal cell densities were determined by electronic particle counting using a Coulter counter. Samples were removed from each each test concentration and the blank at 24, 48 and 72 hours. Particle counts from blank vessels showed high variability and therefore particle counts from the test vessels were used directly andf the blank was not subtracted.
Reference substance (positive control):
no
Duration:
72 h
Dose descriptor:
EC50
Effect conc.:
2.6 mg/L
Nominal / measured:
meas. (arithm. mean)
Conc. based on:
test mat.
Basis for effect:
biomass
Remarks:
yield
Remarks on result:
other: Tetrachloroauric acid concentration calculated from measured gold concentration. 95% confidence intervals: 2.5 - 2.8 mg/L
Duration:
72 h
Dose descriptor:
EC50
Effect conc.:
2.8 mg/L
Nominal / measured:
nominal
Conc. based on:
test mat.
Basis for effect:
biomass
Remarks:
yield
Remarks on result:
other: 95% confidence intervals: 2.7 - 3.0 mg/L
Duration:
72 h
Dose descriptor:
NOEC
Effect conc.:
0.9 mg/L
Nominal / measured:
meas. (arithm. mean)
Conc. based on:
test mat.
Basis for effect:
biomass
Remarks:
yield
Remarks on result:
other: Tetrachloroauric acid concentration calculated from measured gold concentration.
Duration:
72 h
Dose descriptor:
NOEC
Effect conc.:
0.98 mg/L
Nominal / measured:
nominal
Conc. based on:
test mat.
Basis for effect:
biomass
Remarks:
yield
Duration:
72 h
Dose descriptor:
EC50
Effect conc.:
> 9 mg/L
Nominal / measured:
meas. (arithm. mean)
Conc. based on:
test mat.
Basis for effect:
growth rate
Remarks on result:
other: Tetrachloroauric acid concentration calculated from measured gold concentration.
Duration:
72 h
Dose descriptor:
EC50
Effect conc.:
> 10 mg/L
Nominal / measured:
nominal
Conc. based on:
test mat.
Basis for effect:
growth rate
Duration:
72 h
Dose descriptor:
NOEC
Effect conc.:
0.9 mg/L
Nominal / measured:
meas. (arithm. mean)
Conc. based on:
test mat.
Basis for effect:
growth rate
Remarks on result:
other: Tetrachloroauric acid concentration calculated from measured gold concentration.
Duration:
72 h
Dose descriptor:
NOEC
Effect conc.:
0.98 mg/L
Nominal / measured:
nominal
Conc. based on:
test mat.
Basis for effect:
growth rate
Validity criteria fulfilled:
yes
Conclusions:
The EC50 for tetrachloroauric acid is reported to be >9.0 mg/L (measured concentration) based on growth rate and the NOEC is 0.9 mg/L.
Executive summary:

Data are available from a GLP-compliant study following OECD guideline 201, with a minor deviation in test media used due to potential for the standard media to interfere with the test item. A range finding and definitive test were performed using 5 concentrations of test item. In the definitive test 3 replicates per concentration and 6 replicates for the control were tested. Results are reported based on measured and nominal concentrations. This study is considered to be suitable for use as the key study for this endpoint. The EC50 for tetrachloroauric acid is reported to be >9.0 mg/L (measured concentration)  based on growth rate and the NOEC is 0.9 mg/L.

Endpoint:
toxicity to aquatic algae and cyanobacteria
Type of information:
experimental study
Adequacy of study:
supporting study
Reliability:
2 (reliable with restrictions)
Rationale for reliability incl. deficiencies:
guideline study with acceptable restrictions
Remarks:
Toxicity tests conducted with the gold (III) substance gold chloride (AuCl3). Tests conducted according to standard guidelines with minor limitations in reporting. GLP compliance is not reported. Effect concentrations have been converted to concentrations of tetrachloroauric acid based on molecular weight.
Qualifier:
according to guideline
Guideline:
OECD Guideline 201 (Alga, Growth Inhibition Test)
Principles of method if other than guideline:
Study also performed according to Blaise and Vasseur 2005.
GLP compliance:
no
Analytical monitoring:
yes
Vehicle:
no
Details on test solutions:
A stock solution of 1000 mg Au(III)/L was prepared in sterile deionised water and the stock solution was diluted with OECD algal medium to obtain the treatment solutions for the algal assay and with tris-acetate-phosphate (TAP) medium for the euglena assays.
Test organisms (species):
other: Pseudokirchnierella subcapitata and Euglena gracilis
Details on test organisms:
- Test organisms: Pseudokirchnierella subcapitata and Euglena gracilis
- Source: P. subcapitata obtained fro UTEX (University of Texas, Austin, USA) and E. gracilis obtained from the Korea Marine Microalgae Culture Center (KMCC, Korea)
Test type:
static
Water media type:
freshwater
Limit test:
no
Total exposure duration:
72 h
Test temperature:
24 +/- 1 C
Nominal and measured concentrations:
Nominal concentrations of 0, 0.1, 0.5, 1, 5 and 10 mg Au(III)/L.
The maximum concentration of chloride used as gold salt was 1.8 mg Cl/L.
Details on test conditions:
- Medium: P. subcapitata cultured in OECD algal medium and E. gracilis cultured in tris-acetate-phosphate medium.
- Test vessel: 48-well microplate, containing 0.9 mL of test solution and 0.1 mL of test organism per well.
- Initial cell density: 5 x 10E4 cells/mL
- Replicates: 6 to 11
- Photoperiod: 16:8 h light:dark photoperiod
- Incubation: Shaken at 100 rpm in an algal growth chamber
Reference substance (positive control):
no
Duration:
72 h
Dose descriptor:
EC10
Effect conc.:
1.02 mg/L
Nominal / measured:
nominal
Conc. based on:
test mat.
Remarks:
tetrachloroauric acid
Basis for effect:
biomass
Remarks on result:
other: P. subcapitata; effect concentrations reported for Au(III), converted to tetrachloroauric acid, based on molecular weight.
Duration:
72 h
Dose descriptor:
NOEC
Effect conc.:
1.73 mg/L
Nominal / measured:
nominal
Conc. based on:
test mat.
Remarks:
tetrachloroauric acid
Basis for effect:
biomass
Remarks on result:
other: P. subcapitata; effect concentrations reported for Au(III), converted to tetrachloroauric acid, based on molecular weight.
Duration:
72 h
Dose descriptor:
EC10
Effect conc.:
0.002 mg/L
Nominal / measured:
nominal
Conc. based on:
test mat.
Remarks:
tetrachloroauric acid
Basis for effect:
biomass
Remarks on result:
other: E. gracilis; effect concentrations reported for Au(III), converted to tetrachloroauric acid, based on molecular weight.
Details on results:
The concentrations of Au(III) after the exposure duration were measured as 9.30 +/- 0.35 mg/L at 72 h for 10 mg/L dissolved in OECD algal media and 9.78 +/- 0.40 mg/L at 72 h for 10 mg/L dissolved in TAP media. Measured concentrations for OECD algal media (7.01%) and TAP media (2.24%), ERS (19.6%) were decreased compared to nominal concentrations. Nominal concentrations were used for the calculation and reporting of results.

Effect concentrations based on concentrations of Au(III):
48-h EC50 (P. subcapitata, biomass): 1.6 mg/L (95% CL: 1.35-1.89 mg/L)
48-h EC10 (P. subcapitata, biomass): 0.34 mg/L (95% CL: 0-0.97 mg/L)
72-h EC50 (P. subcapitata, biomass): 2.04 mg/L (95% CL: 1.79-2.34 mg/L)
72-h EC10 (P. subcapitata, biomass): 0.59 mg/L
48-h NOEC (P. subcapitata, biomass): 0.5 mg/L
48-h LOEC (P. subcapitata, biomass): 1 mg/L
72-h NOEC (P. subcapitata, biomass): 1 mg/L
72-h LOEC (P. subcapitata, biomass): 5 mg/L
48-h EC50 (E. graciliis, biomass): < 0.1 mg/L (lowest concentration tested)
48-h EC10 (E. graciliis, biomass): < 0.1 mg/L (lowest concentration tested)
72-h EC50 (E. graciliis, biomass): 1.01 mg/L (95% CL: 0.05-2.01 mg/L)
72-h EC10 (E. graciliis, biomass): 0.001 mg/L (95% CL: 0-0.007 mg/L)
72-h LOEC (E. graciliis, biomass): 0.1 mg/L
Reported statistics and error estimates:
The growth inhibition data of alga and euglena were estimated as yield values, which were the biomass at the end of the test minus the starting biomass. The percentage inhibition yield at each concentration was calculated for the control, and expressed as growth yield. L(E)C50 values and corresponding 95% confidence limits for each species were estimated using the Trimmed-Spearman-Karber (TSK) Program. EC10 values and corresponding 95% confidence limits were estimated using the Probit program. NOEC values were calculated using Dunnett’s Program.
Validity criteria fulfilled:
not specified
Conclusions:
Results reported in the study have been converted to concentrations of tetrachloroauric acid based on molecular weight. The 72-hour IC50 and IC10 based on biomass were determined to be 3.52 mg/L tetrachloroauric acid (2.04 mg Au(III)/L) and 1.02 mg/L tetrachloroauric acid (0.59 mg Au(III)/L), respectively, for P. subcapitata. The NOEC for P. subcapitata was 1.73 mg/L tetrachloroauric acid. The IC50 for E. gracilis was 1.74 mg/L tetrachloroauric acid (1.01 mg Au(III)/L), based on biomass.
Executive summary:

The acute toxicity of gold (III) chloride was assessed for Pseudokirchnierella subcapitata and Euglena gracilis. Organisms were exposed to nominal concentrations of 0, 0.1, 0.5, 1, 5 and 10 mg Au(III)/L for 72 hours in OECD algal medium for P. subcapitata and tris-acetate-phosphate for E. gracilis. This study is reliable with restrictions as it was conducted according to a standard guideline and was well-reported, but no information on GLP compliance was reported. Results reported in the study have been converted to concentrations of tetrachloroauric acid based on molecular weight. The 72-hour IC50 and IC10 based on biomass were determined to be 3.52 mg/L tetrachloroauric acid (2.04 mg Au(III)/L) and 1.02 mg/L tetrachloroauric acid (0.59 mg Au(III)/L), respectively, for P. subcapitata. The NOEC for P. subcapitata was 1.73 mg/L tetrachloroauric acid. The IC50 for E. gracilis was 1.74 mg/L tetrachloroauric acid (1.01 mg Au(III)/L), based on biomass. Effect concentrations were converted to concentrations of tetrachloroauric acid, based on molecular weight.

Endpoint:
toxicity to aquatic algae and cyanobacteria
Type of information:
experimental study
Adequacy of study:
supporting study
Reliability:
2 (reliable with restrictions)
Rationale for reliability incl. deficiencies:
guideline study with acceptable restrictions
Remarks:
Toxicity test conducted with the test item, gold (III) chloride trihydrate. Test conducted according to standard guidelines with minor deviation from the guideline and limitations in reporting. GLP compliance is not reported.
Qualifier:
according to guideline
Guideline:
OECD Guideline 201 (Alga, Growth Inhibition Test)
Deviations:
yes
Remarks:
Beakers volume reduced to 20 mL in order to reduce the light path through the coloured solution. The cell concentration determined by fluorescence.
GLP compliance:
no
Remarks:
Study carried under UN-EN ISO 17025.
Analytical monitoring:
no
Vehicle:
no
Details on test solutions:
PREPARATION AND APPLICATION OF TEST SOLUTION (especially for difficult test substances)
- Method: Stock solution prepared by dissolving test material in deionised water. The stock solution was further dissolved in test media to achieve test concentrations.
- Controls: yes, positive control
Test organisms (species):
Desmodesmus subspicatus (previous name: Scenedesmus subspicatus)
Details on test organisms:
TEST ORGANISM
- Common name: freshwater algae
- Strain: not reported
- Source (laboratory, culture collection): not reported

ACCLIMATION
- Acclimation period: not reported
Test type:
static
Water media type:
freshwater
Limit test:
no
Total exposure duration:
72 h
Post exposure observation period:
No.
Hardness:
Not reported.
Test temperature:
Not reported.
pH:
Not reported.
Dissolved oxygen:
Not reported.
Salinity:
Not applicable.
Nominal and measured concentrations:
0.5 - 5.0 mg/L
Details on test conditions:
TEST SYSTEM
- Test vessel: beakers
- Material, size, headspace, fill volume: test beakers were filled with 20 mL of the test solution. The volume of the test beakers was reduced from 100 mL to 20 mL in order to reduce the light path through the coloured solution.

GROWTH MEDIUM
- Standard medium used: yes

EFFECT PARAMETERS MEASURED (with observation intervals if applicable) : cell concentration determined by fluorescence every 24 h with a reader Tecan Infinite M200 and correlated to the cell concentration with the following parameters: ʎ excitation on 437 and ʎ emission of 686 nm.
Reference substance (positive control):
yes
Remarks:
potassium dichromate
Duration:
72 h
Dose descriptor:
EC50
Effect conc.:
1.91 mg/L
Nominal / measured:
not specified
Conc. based on:
test mat.
Basis for effect:
biomass
Remarks on result:
other: 95% IC: 1.30–2.83
Details on results:
The test item, gold (III) chloride trihydrate, was shown to be toxic to freshwater algae.
Results with reference substance (positive control):
Positive control group containing potassium dichromate showed EC50 values within the range of the historical control data, ensuring the quality of the test.
Validity criteria fulfilled:
not specified
Conclusions:
The 72-h EC50 was estimated to be 1.91 mg/L (1.30–2.83, 95% IC) for Scenedesmus subspicatus.
Executive summary:

The acute toxicity of gold (III) chloride trihydrate was assessed for the algae Scenedesmus subspicatus following the OECD 201 guideline. Organisms were exposed to the test item at a range of test concentrations between 0.5 and 5 mg/L for 72 hours. This study is reliable with restrictions as it was conducted according to standard guideline, however there are some deficiencies in methods description and in reporting of results.

The 72-h EC50 was estimated to be 1.91 mg/L (1.30–2.83, 95% IC) for Scenedesmus subspicatus.

Description of key information

The EC50 for tetrachloroauric acid is reported to be >9.0 mg/L (measured concentration) based on growth rate.
The NOEC for tetrachloroauric acid is 0.9 mg/L (measured concentration) based on growth rate.

Key value for chemical safety assessment

EC50 for freshwater algae:
9 mg/L
EC10 or NOEC for freshwater algae:
0.9 mg/L

Additional information

Data are available from a GLP-compliant study following OECD guideline 201, with a minor deviation in test media used due to potential for the standard media to interfere with the test item. A range finding and definitive test were performed using 5 concentrations of test item. In the definitive test 3 replicates per concentration and 6 replicates for the control were tested. Results are reported based on measured and nominal concentrations (Brixham 2012). This study is considered to be suitable for use as the key study for this endpoint. The EC50 for tetrachloroauric acid is reported to be >9.0 mg/L (measured concentration)  based on growth rate and the NOEC is 0.9 mg/L.

Supporting data are available from a published guideline study for gold (III) chloride. Tests were conducted on two algae species with 5 test concentrations for 72 hours. Each treatment consisted of 6 to 11 replicates and results were reported as nominal concentrations (Nam et al. 2014). Effect concentrations were converted to tetrachloroauric acid concentrations on the basis of molecular weight. This study is considered to be suitable for use as a supporting study for this endpoint. The 72-hour IC50 and IC10 based on biomass were determined to be 3.52 mg/L tetrachloroauric acid (2.04 mg Au(III)/L) and 1.02 mg/L tetrachloroauric acid (0.59 mg Au(III)/L), respectively, for P. subcapitata. The NOEC for P. subcapitata was 1.73 mg/L tetrachloroauric acid. The IC50 for E. gracilis was 1.74 mg/L tetrachloroauric acid (1.01 mg Au(III)/L), based on biomass.

The acute toxicity of gold (III) chloride trihydrate to algae was also tested in a study according to the OECD 201 guideline (Garcia-Cambero 2013). The test was conducted with the test item at a range of test concentrations between 0.5 and 5 mg/L for 72 hours. This study is considered to be suitable for use as a supporting study for this endpoint. The 72-h EC50 was estimated to be 1.91 mg/L (1.30–2.83, 95% IC) for Scenedesmus subspicatus.