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Toxicity to aquatic algae and cyanobacteria

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Reference
Endpoint:
toxicity to aquatic algae and cyanobacteria
Type of information:
experimental study
Adequacy of study:
key study
Study period:
08/04/2011 - 09/06/2011
Reliability:
1 (reliable without restriction)
Rationale for reliability incl. deficiencies:
other:
Remarks:
This study was conducted in accordance with OECD guideline 201, with analytical confirmation of exposure concentrations and GLP standards were maintained throughout.
Qualifier:
according to guideline
Guideline:
OECD Guideline 201 (Alga, Growth Inhibition Test)
Deviations:
no
Qualifier:
according to guideline
Guideline:
other: other guideline
Version / remarks:
Test method relating to new chemical substances "Alga growth inhibition test, Daphnia sp. acute immobilisation test, and fish acute toxicity test" (Japanese notification, Yakushokuhatsu 0331 No.7, Heisei 23.03.29 Seikyoku No.5, Kanpokihatsu No. 110331009)
Deviations:
no
GLP compliance:
yes
Specific details on test material used for the study:
IUPAC name: Aluminium titanium oxide
CAS No.: 12004-39-6
Molecular weight: 181.83
Purity: 99.9%
Melting point: 1860oC
Solubility in water: <50 mg/ml
Appearance: White powder
Stability: Stable for 5 years at room temperature
Analytical monitoring:
yes
Details on sampling:
The concentrations of the test substance in all test solutions and test cultures were measured at initiation (0h) and termination (72h).
Vehicle:
no
Details on test solutions:
Stock solution was prepared by adding an appropriate amount of the test material to dechlorinated dilution water, followed by ultrasonification for 30 minutes, stirring with a stirrer for 48 hours and then filtering (membrane filter, HA, 0.45um, Millipore corporation). Appropriate amounts of stock dilution were diluted to create the fnal test solutions. Nominal test concentrations used in this study were 0 (control), 10, 18, 32, 56 and 100 mg/L.

As an interaction between the test substance and constituents of the algal growth medium was anticipated that might cause an adverse impact on algal growth by reducing the bioavailability of essential nutrients, an enhanced medium that contained components at concentrations twice as high as the standard medium was also created to test algal toxicity mitigation.
Test organisms (species):
Pseudokirchneriella subcapitata (previous names: Raphidocelis subcapitata, Selenastrum capricornutum)
Details on test organisms:
Test organism: Pseudokirchneriella subcapitata (Unicellular green alga)
Strain: ATCC22662
Source: American Type Culture Collection
Test type:
static
Water media type:
freshwater
Limit test:
no
Total exposure duration:
72 h
Post exposure observation period:
None.
Test temperature:
22 ± 2oC.
pH:
7.9 in all treatments at 0h, 8.2 (in control) to 7.8 at 72hr.
Dissolved oxygen:
Not measured.
Salinity:
Not appropriate.
Nominal and measured concentrations:
Nominal Concentrations: 0(control), 10, 18, 32, 56 and 100 mg/L.
Measured based on aluminium analysis: 0, 0.0483, 0.0776, 0.127, 0.252 and 0.388 mg/L.
Measured based on titanium analysis: 0, 0.0293, 0.0528, 0.0821, 0.169 and 0.274 mg/L.
Details on test conditions:
See method below
Reference substance (positive control):
yes
Remarks:
The sensitivity of the algae was checked in a separate study performed on 2010-12-17 with the reference substance potassium dichromate.
Duration:
72 h
Dose descriptor:
other: ErC50
Effect conc.:
> 0.21 mg/L
Nominal / measured:
meas. (arithm. mean)
Conc. based on:
test mat.
Remarks:
derived from measured titanium concentration
Basis for effect:
growth rate
Remarks on result:
other: 2x concentrated algal growth medium
Duration:
72 h
Dose descriptor:
other: NOECr
Effect conc.:
0.21 mg/L
Nominal / measured:
meas. (arithm. mean)
Conc. based on:
test mat.
Remarks:
derived from measured titanium concentration
Basis for effect:
growth rate
Remarks on result:
other: 2x concentrated algal growth medium
Duration:
72 h
Dose descriptor:
other: NOECr
Effect conc.:
0.398 mg/L
Nominal / measured:
meas. (arithm. mean)
Conc. based on:
test mat.
Remarks:
derived from measured aluminium concentration
Basis for effect:
growth rate
Remarks on result:
other: 2x concentrated algal growth medium
Duration:
72 h
Dose descriptor:
other: ErC50
Effect conc.:
> 0.274 mg/L
Nominal / measured:
meas. (arithm. mean)
Conc. based on:
test mat.
Remarks:
derived from measured titanium concentration
Basis for effect:
growth rate
Remarks on result:
other: standard algal growth medium
Duration:
72 h
Dose descriptor:
other: ErC50
Effect conc.:
> 0.388 mg/L
Nominal / measured:
meas. (arithm. mean)
Conc. based on:
test mat.
Remarks:
derived from measured aluminium concentration
Basis for effect:
growth rate
Remarks on result:
other: standard algal growth medium
Details on results:
The 72hr ErC50 based on aluminium analysis was determined to be >0.388 mg/L.

The 72hr ErC50 based on titanium analysis was determined to be >0.274 mg/L in the standard growth medium.
The 72-hour EC50 was determined to be >210 mg/L in the enhanced growth medium and the NOECr was found to be 0.210 mg/L.

No unusual cell shape or agglutination was observed at any test concentration after 72 hours.

The NOEC for the standard medium occured at the 10 mg/L loading rate, while the NOEC for the enhanced medium occured at 100 mg/L. As concentrations of test substance measured were comparable in the 100 mg/L treatments of both standard and enhanced media, it was suggested that the effect of the test substance in the standard medium was due to decreased bioavailability of nutrients caused by aluminium and titanium forming complexes with components of the test media. As such, this result does not represent a toxic effect and is therefore not used for derivation of endpoints.
Results with reference substance (positive control):
The 72hr ErC50 of potassium dichromate was 0.835 mg/L (95% CL: 0.809-0.861 mg/L), compared to a 10-yr laboratory mean of 0.816+/-0.0831 mg/L (n = 22).
Reported statistics and error estimates:
The mean values of biomass for each concentration group were plotted against time to produce plots of growth curves. Based on the plots on the control group, the exponential growth of algae during the exposure period was checked. The EC50 was calculated by the least squares linear regression as straight in the concentration-inhibition curves. The NOECr values were determined by one-way ANOVA, Williams multiple comparison test, subsequent to Bartlett test for homogeneity of variances.

Table 1.         Concentrations of dialuminium titanium pentaoxide derived from measured concentrations of dissolved aluminium during a 72 hour growth inhibition test with P. subcapitata under static conditions. 

Loading rate, mg test substance/L

Measured concentration (mg dialuminium titanium pentaoxide-equiv/L), based on dissolved aluminium and titanium analysis

aluminium

titanium

0 h

72 h

mean

0 h

72 h

mean

0 (control)

< 0.04

< 0.04

-

< 0.04

< 0.04

-

10

0.0482

0.0485

0.0483

0.0306

0.0281

0.0293

18

0.0752

0.0801

0.0776

0.0550

0.0506

0.0528

32

0.124

0.131

0.127

0.0850

0.0793

0.0821

56

0.267

0.238

0.252

0.165

0.174

0.169

100

0.392

0.384

0.388

0.274

0.274

0.274

LOD:      Limit of detection (0.04 mg-equiv/L).

 

 

Table 2.         Effects on the growth rate of P. subcapitata following 72 hour exposure to dialuminium titanium pentaoxide under static conditions. 

Loading rate,

[mean measured concentration]

mg test substance/L

Meanaalgal biomass, cells/mLb

mean

growth rate μ (0 – 72 h)

%

inhibition

24 h

48 h

72 h

0 (control)

36300

286000

1240000

0.0765

-

10

[0.0483 (Al)]

[0.0293 (Ti)]

36400

275000

1130000

0.0752

1.7

18

[0.0776 (Al)]

[0.0528 (Ti)]

36700

231000

749000

0.0695**

9.1

32

[0.127 (Al)]

[0.0821 (Ti)]

38300

197000

516000

0.0642**

16.1

56

[0.252 (Al)]

[0.169 (Ti)]

36300

156000

301000

0.0569**

25.6

100

[0.388 (Al)]

[0.274 (Ti)]

34500

130000

217000

0.0523**

31.6

a             There were 6 replicates in the control group and 3 replicates in each of the test substance groups. 

b             The initial (0 h) biomass density was 5000 cells/mL in all treatments. 

[(Al)]       Mean measured concentration based on aluminium analysis. 

[(Ti)]       Mean measured concentration based on titanium analysis. 

**            Significantly (α=0.01) different from the control. 

Validity criteria fulfilled:
yes
Conclusions:
The 72hr ErC50 was determined to be >0.210 mg/l and the 72hr NOECr was found to be 0.210 mg/l based on the results from the enhanced growth medium.
Executive summary:

The inhibitory effect of dialuminium titanium pentaoxide on algal growth was determined in a 72 hour growth inhibition test withPseudokirchneriella subcapitata, conducted in accordance with OECD guideline 201. The algae were exposed to the test substance at loading rates of 0 (control), 10, 18, 32, 56 and 100 mg/L. Concentrations of dissolved aluminium and titanium were determined by ICP-MS which were then used to calculate equivalent concentrations of the test substance present in the test solutions. Test media were not renewed during the 72 hours.

The 72hr ErC50 based on aluminium analysis was determined to be >0.388 mg/L. The 72hr ErC50 based on titanium analysis was determined to be >0.274 mg/L in the standard growth medium and >0.210 mg/L in the enhanced growth medium. The 72hr NOECr based on titanium analysis was determined to be 0.210 mg/L in enhanced growth medium.

The NOECr for the standard medium occured at the 10 mg/L loading rate, while the NOEC for the enhanced medium occured at 100 mg/L. As concentrations of test substance measured were comparable in the 100 mg/L treatments of both the standard and enhanced media, it was suggested that the effect of the test substance in the standard medium was due to decreased bioavailability of nutrients caused by aluminium and titanium forming complexes with components of the test media. This is not considered to be a toxicological effect and as such is not used for derivation of NOEC of endpoints.

No unusual cell shape or agglutination was observed at any test concentration after 72 hours.

Description of key information

A key study is provided for the assessment of growth inhibition in algae. The study was conducted in accordance with the appropriate OECD test guideline, with analytical confirmation of exposure concentrations, with no deviations and in compliance with GLP. The 72-hour EC50 was found to be >0.210 mg/L and the NOEC was determined to be 0.210 mg/L.

Key value for chemical safety assessment

EC50 for freshwater algae:
0.21 mg/L
EC10 or NOEC for freshwater algae:
0.21 mg/L

Additional information

The inhibitory effect of dialuminium titanium pentaoxide on algal growth was determined in a 72 hour growth inhibition test with Pseudokirchneriella subcapitata, conducted in accordance with OECD guideline 201. The algae were exposed to the test substance at loading rates of 0 (control), 10, 18, 32, 56 and 100 mg/L. Concentrations of dissolved aluminium and titanium were determined by ICP-MS which were then used to calculate equivalent concentrations of the test substance present in the test solutions. Test media were not renewed during the 72 hours.

The 72hr ErC50 based on aluminium analysis was determined to be >0.388 mg/L. The 72hr ErC50 based on titanium analysis was determined to be >0.274 mg/L in the standard growth medium and >0.210 mg/L in the enhanced growth medium. The 72hr NOECr based on titanium analysis was determined to be 0.210 mg/L in enhanced growth medium.

The NOECr for the standard medium occurred at the 10 mg/L loading rate, while the NOEC for the enhanced medium occurred at 100 mg/L. As concentrations of test substance measured were comparable in the 100 mg/L treatments of both the standard and enhanced media, it was suggested that the effect of the test substance in the standard medium was due to decreased bioavailability of nutrients caused by aluminium and titanium forming complexes with components of the test media. This is not considered to be a toxicological effect and as such is not used for derivation of NOEC of endpoints.

No unusual cell shape or agglutination was observed at any test concentration after 72 hours.