Potassium titanium oxide (K1.88-2.13Ti8O16.94-17.07)

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:

weight of evidence

Study period:

30/06/2003-14/08/2003

Reliability:

2 (reliable with restrictions)

Rationale for reliability incl. deficiencies:

other: see 'Remark'

Remarks:

Study has been performed according to OECD and EC guidelines and according to GLP principles. The restriction is due to the use of the read across approach: the test was performed not with Terracess TF but with Terracess L, a substance which is very similar in structure and physical/chemical properties.

Qualifier:

according to guideline

Guideline:

EU Method C.3 (Algal Inhibition test)

Deviations:

no

Qualifier:

according to guideline

Guideline:

OECD Guideline 201 (Alga, Growth Inhibition Test)

Version / remarks:

(1984)

Deviations:

no

Qualifier:

according to guideline

Guideline:

ISO 8692 (Water Quality - Fresh Water Algal Growth Inhibition Test with Scenedesmus subspicatus and Selenastrum capricornutum)

Deviations:

no

GLP compliance:

yes

Analytical monitoring:

yes

Details on sampling:

- Concentrations: During the final test duplicate samples for possible analysis were taken from all test concentrations and the control at the start and the end of the test period. Additionally, duplicate samples were taken after 24 hours from the 5 µm filtered solution prepared at 100 mg/l and the control. The volume taken was 20 ml and it was sampled from the approximate centre of the vessels
- Sample storage conditions before analysis: All samples were stored in a deep-freeze until shipment to Solvias AG, Basel, Switzerland

Vehicle:

no

Details on test solutions:

As a consequence of the poor solubility, supersaturated solutions were prepared at nominally 100 mg/l and magnetically stirred for three days to achieve maximum solubility. The resulting dispersions were filtered through a paper filter (Schleicher and Schuell 604) to remove the larger undissolved test substance particles (ca. > 5µm). The filtrate was used to prepare the lower test concentrations. The control received the same treatment. All final solutions were clear and colourless.
In the combined range-finding test dilutions containing 0.1, 1.0 and 10% of the filtrate were tested

Test organisms (species):

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

Details on test organisms:

TEST ORGANISM
- Common name: Selenastrum capricornutum
- Strain: NIVA CHL 1
- Source (laboratory, culture collection): In-house laboratory culture
- Method of cultivation: Algae stock cultures were started by inoculating growth medium with algal cells from a pure culture on agar. The suspensions were continuously aerated and exposed to light in a climate room at a temperature of 23 ± 2°C

ACCLIMATION
- Acclimation period: 3 days
- Culturing media and conditions (same as test or not): Pre-culture and test medium: M2. Stock culture medium: M1.

Test type:

static

Water media type:

freshwater

Limit test:

no

Total exposure duration:

72 h

Hardness:

24 mg CaCO3/L

Test temperature:

22.6°C at the start of the test. During the exposure period the temperature measured in the incubator was maintained between 22.1 and 22.6°C

pH:

8.0-9.5

Nominal and measured concentrations:

Controls: Test medium without test substance or other additives (blank).
A 5 µm filtered solution prepared at a loading rate of 100 mg/l and dilutions containing 10, 18, 32 and 56% of the filtrate.
Analysis of the samples taken from the 5 µm filtrate prepared at a loading rate of 100 mg/l showed that measured concentrations based on Lithium at the start of the test were below the detection limit of the analytical method, i.e. below 0.1 mg/l. Those measured for Titanium equalled 0.1 mg/l in the presence of algae and were below the limit of detection in the absence of algae. The Potassium concentration could be measured and remained stable at approximately 4.5 mg/l during the 72-hour test period. Extrapolating the Potassium concentration to the concentration of the test substance TERRACESS L resulted in a concentration of 20 mg/l (using the formula: K concentration analysed / 0.76*M K )* M TERRACESS L). Extrapolating the measured Titanium concentration (0.1 mg/l) to a concentration TERRACESS L resulted in a concentration of 0.4 mg/l. Hence, actual measured TERRACESS L concentrations cannot be determined and are expected to be even lower than indicated above based on the insoluble character of this inert substance

Details on test conditions:

TEST SYSTEM
- Test vessel: 100 ml, all-glass
- Initial cells density: 1 x 10^4 cells/ml
- No. of vessels per concentration (replicates): 3 replicates of each test concentration.
6 replicates of the blank-control.
2 replicates of the highest concentration without algae for sampling purposes.
2 extra replicates of the control and highest concentration for sampling after 24 hours



GROWTH MEDIUM
- Standard medium used: yes


TEST MEDIUM / WATER PARAMETERS
-M2; according to the ISO-Standard "Algal growth inhibition test" Nov. 1989; formulated using Milli-Q water preventing precipitation and with the following composition:
NH4Cl 15 mg/l
MgCl2.6H2O 12 mg/l
CaCl2.2H2O 18 mg/l
MgSO4.7H2O 15 mg/l
KH2PO4 1.6 mg/l
FeCl3.6H2O 80 µg/l
Na2EDTA.2H2O100 µg/l
H3BO3 185 µg/l
MnCl2.4H2O 415 µg/l
ZnCl2 3 µg/l
CoCl2.6H2O 1.5 µg/l
CuCl2.2H2O 0.01 µg/l
Na2MoO4.2H2O 7 µg/l
NaHCO3 50 mg/l
Hardness (Ca+Mg) 0.24 mmol/l (24 mg CaCO3/l)
pH 8.3 ± 0.2



OTHER TEST CONDITIONS
- Sterile test conditions: no
- Adjustment of pH: no
- Photoperiod: Continuously using TLD-lamps of the type ‘Cool-white’ of 30 Watt, with a light intensity within the range of 70 to 95 uE.m-2.s-1



EFFECT PARAMETERS MEASURED (with observation intervals if applicable) :
- Determination of cell concentrations: [counting chamber; electronic particle counter; fluorimeter; spectrophotometer; colorimeter]: At the beginning of the test, cells were counted by microscope, using a counting chamber. Thereafter cell densities were determined by spectrophotometric measurement of samples at 720 nm using a Varian Cary 50 single beam spectrophotometer with immersion probe (pathlength =20 mm). Algal medium was used as blank.


TEST CONCENTRATIONS
- Spacing factor for test concentrations: 1.8
- Range finding study: yes
- Test concentrations: Algae were exposed to a blank-control and a 5 µm filtered solution prepared at a loading rate of 100 mg/l in the limit test. In the combined range-finding test dilutions containing 0.1, 1.0 and 10% of the filtrate were tested
- Results used to determine the conditions for the definitive study: no significant algal growth inhibition was observed at 10% of the filtrate and lower. The mean growth rate reduction and cell growth inhibition at 100 % filtrate were 17% and 50%, respectively. Therefore, the results showed that the EC50 for cell growth inhibition was close to or just below a 5 µm filtered solution prepared at 100 mg/l. The EC50 for growth rate reduction was above the maximum soluble concentration present in the filtered solution

Reference substance (positive control):

yes

Remarks:

potassium dichromate

Key result

Duration:

72 h

Dose descriptor:

EC50

Effect conc.:

> 4.5 mg/L

Nominal / measured:

meas. (arithm. mean)

Conc. based on:

element

Remarks:

potassium

Basis for effect:

growth rate

Remarks on result:

other: (> WSL)

Key result

Duration:

72 h

Dose descriptor:

EC50

Effect conc.:

> 26 mg/L

Nominal / measured:

meas. (arithm. mean)

Conc. based on:

test mat.

Remarks:

Terracess L

Basis for effect:

growth rate

Remarks on result:

other: (>WSL)

Key result

Duration:

72 h

Dose descriptor:

NOEC

Effect conc.:

> 6.4 mg/L

Nominal / measured:

meas. (arithm. mean)

Conc. based on:

test mat.

Remarks:

Terracess L

Basis for effect:

growth rate

Remarks on result:

other: (> WSL)

Key result

Duration:

72 h

Dose descriptor:

NOEC

Effect conc.:

> 3.6 mg/L

Nominal / measured:

meas. (arithm. mean)

Conc. based on:

test mat.

Remarks:

Terracess L

Basis for effect:

biomass

Remarks on result:

other: (> WSL)

Details on results:

- Exponential growth in the control (for algal test): yes
- Observation of abnormalities (for algal test): no

Results with reference substance (positive control):

- Results with reference substance valid? yes
The EC50 for cell growth inhibition (EBC50: 0-72h) was 0.59 mg/l with a 95 % confidence interval ranging from 0.38 to 0.92 mg/l. The ISO-Standard gives a range for the EC50 for cell growth inhibition from 0.20 to 0.75 mg/l based on 16 inter-laboratory tests. Hence, the EBC50: 0-72h for the present batch was within this range.

The EC50 for growth rate reduction (ERC50: 0-72h) was 0.89 mg/l with a 95 % confidence interval ranging from 0.56 to 1.4 mg/l. The ISO-Standard gives a range for the EC50 for cell growth inhibition from 0.60 to 1.03 mg/l based on 16 inter-laboratory tests. Hence, the ERC50: 0-72h for the present batch was within this range

Validity criteria fulfilled:

yes

Reference 2

Endpoint:

toxicity to aquatic algae and cyanobacteria

Type of information:

experimental study

Adequacy of study:

weight of evidence

Study period:

18 October 2000-31 May 2001

Reliability:

2 (reliable with restrictions)

Rationale for reliability incl. deficiencies:

other: see 'Remark'

Remarks:

Study has been performed according to the appropriate ISO and OECD and EC guidelines and according to GLP principles. The restriction is due to the use of the read across approach: the test was performed not with Terracess TF but with Terracess P, a substance which is very similar in structure and physical/chemical properties.

Qualifier:

according to guideline

Guideline:

ISO 8692 (Water Quality - Fresh Water Algal Growth Inhibition Test with Scenedesmus subspicatus and Selenastrum capricornutum)

Version / remarks:

(1989)

Deviations:

no

Qualifier:

according to guideline

Guideline:

EU Method C.3 (Algal Inhibition test)

Version / remarks:

(1992)

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

Details on sampling:

- Concentrations:
- Sampling method: samples (10 mL) were taken from each concentration before introduction of algae at the start of the exposure. After 24 h and at the end of the exposure (t=72 h), the replicates incubated with and without algae were pooled at each concentration. Subsequently, samples were taken from all pooled solutions.

- Sample storage conditions before analysis: all samples were stored in a deep freeze until shipment to Solvias AG, Basel, Switzerland.

Vehicle:

no

Details on test solutions:

Supersaturated solutions were prepared at nominally 100 mg/L and magnetically stirred for 4 days to achieve maximum solubility. The resulting solutions were filtered through a paper filter to remove the larger undissolved test substance particles (ca. > 5µm). The filtrates were used to prepare the lower test concentrations (10, 18, 32 and 56% of the filtrate). The undiluted filtrate and the concentration containing 56% of the filtrate were slightly turbid (hazy). All lower test concentrations were clear and colourless. After preparation, volumes of 50 ml were added to each replicate of the respective test concentration. Subsequently, adequate volumes of an algal suspension were added to each replicate providing a cell density of 10E4 cells/mL.

Test organisms (species):

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

Details on test organisms:

TEST ORGANISM
- Common name: Pseudokirchneriella subcapitata
- Strain: CCAP 278/4.
- Source (laboratory, culture collection): In-house laboratory culture
- Age of inoculum (at test initiation): not indicated
- Method of cultivation: Algae stock cultures were started by inoculating growth medium with algal cells from a pure culture on agar. The suspensions were continuously aerated and exposed to light (4000-9000 lux) in a climate room at a temperature of 23 ± 2°C.

ACCLIMATION
- Acclimation period: 4 days
- Culturing media and conditions (same as test or not): test medium and pre-culture medium is M2 according to the ISO standard "Algal growth inhibition test" .

Test type:

static

Water media type:

freshwater

Limit test:

no

Total exposure duration:

72 h

Post exposure observation period:

No post exposure observation method.

Hardness:

24 mg CaCO3/L

Test temperature:

20.9 - 24.0 °C.

pH:

8.1 - 10.0

Nominal and measured concentrations:

The determination of the TERRACESS P concentrations in the water phase was based on measurement of TITANIUM (Ti) concentrations. The titanium was present in samples at approximately 28%. Detailed specification of the analytical procedures was put down in a Study Part (L01-000672).
The concentrations represented as percentage (%) are the nominal ones prepared by the dilution of the 5 µm filtrate prepared at nominal 100 mg/l. The concentrations measured as µg/L represent the measured Ti concentrations in water.

Sampling time: t=0

32%: 574 µg/L
56%: 937 µg/L
100% : 321 µg/L
100% (without algae): 2331 µg/L

Sampling time: t=72h

32%: 88 µg/L
56%: 268 µg/L
100% : 298 µg/L
100% (without algae): 117 µg/L

As expected, analysis of the samples showed that concentrations decreased (significantly) during the test period as a consequence of the extremely poor solubility in water. The variation in measurements at the start of the test in the 100 mg/L solution with and without algae was probably a consequence of the inhomogeneity of the samples, i.e. undissolved test material, which was related to the in fact only indicative for the concentrations present in the test solutions. All analysed concentrations at the end of the test period (72 h) showed measured concentrations that were still above the maximum calculated solubility level  for the test substance. Hence, the conclusion can be made that these concentrations (32% to 100% of the filtrate) were inhomogeneous dispersions with undissolved test substance particles present in various amounts.     

Details on test conditions:

TEST SYSTEM
- Test vessel: 100 mL, all-glass, containing 50 mL of test solution
- Initial cells density: 1 x 104 cells/mL.
- No. of vessels per concentration (replicates): 3 replicates of each test concentration, 6 replicates of the blank control, 1 replicate of each concentration without algae, 1 extra replicate of each test concentration for sampling purposes, 2 extra replicates of the highest concentration without algae.

GROWTH MEDIUM
- Standard medium used: yes

OTHER TEST CONDITIONS
- Sterile test conditions: no
- Adjustment of pH: no
- Photoperiod: continuous
- Light intensity and quality: Cool-white TLD-lamps of 30 Watt, with a light intensity within the range of 99 to 132 µE.m-2.s-1.

EFFECT PARAMETERS MEASURED (at t=0, 24 h, 48 h and 72h) :
- Determination of cell densities: microscope/counting chamber (t=0) and spectrophotometer (t> 0)

TEST CONCENTRATIONS
- Spacing factor for test concentrations: approx. 1.8

- Range finding study
- Test concentrations: Algae were exposed to a 5 µm filtrate prepared at nominal 100 mg/l and to dilutions containing 0.1, 1.0 and 10% of the filtrate.
- Results used to determine the conditions for the definitive study: no significant inhibition of algal growth was observed at concentrations corresponding with 10% of the filtrate and lower. The undiluted filtrate inhibited algal growth by just over 50% (54.7%). Hence, the EC50 for algal cell growth inhibition was expected to be just below a 5 µm filtered solution prepared at 100 mg/L, i.e. just below the expected maximum solubility in the test medium.

Reference substance (positive control):

yes

Remarks:

Potassium dichromate

Duration:

72 h

Dose descriptor:

EC50

Effect conc.:

>= 2.185 mg/L

Nominal / measured:

meas. (arithm. mean)

Conc. based on:

other: Terraces P

Basis for effect:

growth rate

Duration:

72 h

Dose descriptor:

EC50

Effect conc.:

>= 602 µg/L

Nominal / measured:

meas. (arithm. mean)

Conc. based on:

element

Remarks:

Titanium

Basis for effect:

growth rate

Duration:

72 h

Dose descriptor:

NOEC

Effect conc.:

> 1.21 mg/L

Nominal / measured:

meas. (arithm. mean)

Conc. based on:

other: Terraces P

Basis for effect:

growth rate

Duration:

72 h

Dose descriptor:

NOEC

Effect conc.:

> 331 µg/L

Nominal / measured:

meas. (arithm. mean)

Conc. based on:

element

Remarks:

Titanium

Basis for effect:

growth rate

Details on results:

TERRACESS P inhibited growth and reduced growth rate of the fresh water algae species significantly at concentrations corresponding with 56% and higher of a 5 µm filtered solution prepared at a nominal loading of 100 mg/l. This corresponded with a measured concentration decreasing from 3.4 mg/L at the start to 0.97 mg/L Terraces P at the end of the test period, i.e. concentrations above the maximum solubility limit in water (< 36 µg/L Terraces P).

The NOEC for cell growth inhibition and growth rate reduction corresponded with 32% of a 5 µm filtered solution prepared at a nominal loading of 100 mg/l. This corresponded with a measured concentration decreasing from 2.1 mg/L at the start to 0.32 mg/l Terraces P at the end of the test period.
The concentration measured at the end of the test period was above the calculated solubility limit in water (< 36 µg/L Terraces P).

In conclusion: effects from TERRACESS P on growth of the fresh water algae species Selenastrum capricornutum only occurred at concentrations above the water solubility limit.

- Exponential growth in the control (for algal test): yes
- Observation of abnormalities (for algal test): no abnormalities observed
- Any stimulation of growth found in any treatment: no

Results with reference substance (positive control):

The EC50 for cell growth inhibition (EBC50:0-72h) was 0.59 mg/L with a 95 % confidence interval ranging from 0.32 to 1.1 mg/L.

The EC50 for growth rate reduction (ERC50:0-72h) was 1.1 mg/L with a 95 % confidence interval ranging from 0.71 to 1.7 mg/L.

Reported statistics and error estimates:

The ISO-Standard gives a range for the EC50 for cell growth inhibition between 0.20 to 0.75 mg/L based on 16 inter-laboratory tests. Hence, the EBC50:0-72h for the present batch corresponds with this range.
The ISO-Standard gives a range for the EC50 for cell growth inhibition between 0.60 to 1.03 mg/L based on 16 inter-laboratory tests. Hence, the ERC50:0-72h for the present batch was slightly higher than the upper limit of the ISO standard range. However, the lower limit of the 95% confidence interval was still within the ISO standard range.

Validity criteria fulfilled:

yes

Reference 3

Endpoint:

toxicity to aquatic algae and cyanobacteria

Type of information:

read-across based on grouping of substances (category approach)

Adequacy of study:

key study

Justification for type of information:

1. HYPOTHESIS FOR THE CATEGORY APPROACH (ENDPOINT LEVEL)
Please see attached justification.
2. CATEGORY APPROACH JUSTIFICATION (ENDPOINT LEVEL
Please see attached justification.

Reason / purpose for cross-reference:

read-across source

Reason / purpose for cross-reference:

read-across source

Key result

Duration:

72 h

Dose descriptor:

EC50

Effect conc.:

> 26 mg/L

Nominal / measured:

meas. (arithm. mean)

Conc. based on:

test mat.

Basis for effect:

growth rate

Remarks on result:

other: Results are based on the read across substance Terracess L.

Key result

Duration:

72 h

Dose descriptor:

NOEC

Effect conc.:

> 6.4 mg/L

Nominal / measured:

meas. (arithm. mean)

Conc. based on:

test mat.

Basis for effect:

growth rate

Remarks on result:

other: Results are based on the read across substance Terracess L.

Key result

Duration:

72 h

Dose descriptor:

NOEC

Effect conc.:

> 3.6 mg/L

Nominal / measured:

meas. (arithm. mean)

Conc. based on:

test mat.

Basis for effect:

biomass

Remarks on result:

other: Results are based on the read across substance Terracess L.

Key result

Duration:

72 h

Dose descriptor:

EC50

Effect conc.:

>= 2.185 mg/L

Nominal / measured:

meas. (arithm. mean)

Conc. based on:

test mat.

Basis for effect:

growth rate

Remarks on result:

other: Results based on read across substance Terracess P.

Key result

Duration:

72 h

Dose descriptor:

NOEC

Effect conc.:

> 1.21 mg/L

Nominal / measured:

meas. (arithm. mean)

Conc. based on:

test mat.

Basis for effect:

growth rate

Remarks on result:

other: Results based on read across substance Terracess P.

Description of key information

The ErC50  for algae exceeds the maximum solubility of the substance in water in studies on two similar substances. Therefore the weight of evidence shows that Terracess TF will not be toxic to algae.

Additional information

Under the conditions of the present study with Selenastrum capricornutum exposed to TERRACESS L, the ErC50 was calculated to be above the water solubility.

Effects from Terracess P on growth of the fresh water algae species Selenastrum capricornutum only occurred at concentrations above the water solubility limit which calculated to be 36 µg/L Terraces P.