2,4,8,10-tetra(tert-butyl)-6-hydoroxy-12H-dibenzo[d,g] [1,3,2]dioxaphosphocin, 6-oxide

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:

From 2017-11-06 to 2017-11-09

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)

Version / remarks:

2006

Deviations:

no

GLP compliance:

yes

Specific details on test material used for the study:

Purity: > 99%
Batch No.: 101Z4

Analytical monitoring:

yes

Details on sampling:

Cell growth
- Concentrations: stock solution content of 100, 35.7, 12.8, 4.56 and 1.63%
- Sampling method: every 24 hours after the start of exposure

Vehicle:

no

Details on test solutions:

PREPARATION AND APPLICATION OF TEST SOLUTION (especially for difficult test substances)
- Method: The test sample (0.100 g) and medium (1000 mL) were mixed to prepare the nominal concentration of 100 mg/L, and they were stirred with a magnetic stirrer for 48 hours. Then, the suspension was filtered with a glass fiber filter by suction. The filtrate was used as stock solution. The test solution was prepared in a preparation container by mixing required volume of the stock solution and the medium processing similar to the control and divided into each test vessel. In 100% level, the stock solution was divided into test vessel.
- Controls: The medium without the test item, which was treated in the same manner as stock solution.

Test organisms (species):

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

Details on test organisms:

TEST ORGANISM
- Common name: Pseudokirchneriella subcapiitata
- Source (laboratory, culture collection): American Type Culture Collection

ACCLIMATION
- Acclimation period: 3 days
- Culturing media and conditions (same as test or not): yes

Test type:

static

Water media type:

freshwater

Limit test:

no

Total exposure duration:

72 h

Test temperature:

23.2-23.3 °C

pH:

7.1-7.7

Nominal and measured concentrations:

Nominal: Stock solution content of 100, 35.7, 12.8, 4.56 and 1.63%
Measured: 80.4, 28.2, 9.85, 3.51 and 1.16 mg/L

Details on test conditions:

TEST SYSTEM
- Test vessel: Sterilized 300mL Erlenmeyer fllask
- Initial cells density: 0.54E+04 cells / mL
- No. of vessels per concentration (replicates): 3
- No. of vessels per control (replicates): 6

GROWTH MEDIUM
- Standard medium used: yes

OTHER TEST CONDITIONS
- Sterile test conditions: yes
- Light intensity and quality:
Nominal: 90 μmol.m-2.s-1 in wavelength range of 400-700 nm within ± 20% of nominal (within ± 15% from the average light intensity)
Measured: 87 - 91 μmol.m-2.s-1, Continuous illumination provided by alternative LED light to fluorescent lamp

EFFECT PARAMETERS MEASURED (with observation intervals if applicable) :
- Determination of cell concentrations: particle counter

TEST CONCENTRATIONS
- Spacing factor for test concentrations: 2.8
- Range finding study
- Test concentrations: 1, 3.2, 10.0, 32.0, 100% stock solution content
- Results used to determine the conditions for the definitive study: Growth inhibition rates based on growth rate (0-3 d) of 1% stock solution content on two test organisms were -2.4% and -2.0%, which of 100% stock solution content were 89%, 87%.

Reference substance (positive control):

yes

Remarks:

Potassium dichromate

Key result

Duration:

72 h

Dose descriptor:

EC50

Effect conc.:

20 mg/L

Nominal / measured:

meas. (geom. mean)

Conc. based on:

test mat.

Basis for effect:

growth rate

Key result

Duration:

72 h

Dose descriptor:

NOEC

Effect conc.:

3.51 mg/L

Nominal / measured:

meas. (geom. mean)

Conc. based on:

test mat.

Basis for effect:

growth rate

Remarks on result:

other: growth rate 0-3 d

Details on results:

- Observation of abnormalities (for algal test): In 80.4 mg/L levels, some bloated cells and small cells were observed. In 28.2 mg/L level, some cells were bloated. In 9.85 mg/L level, some small cells were observed.
- Growth inhibition: In 80.4 mg/L levels, growth inhibition was observed remarkably during exposure. In 28.2 mg/L level, algae grew until 24 hours after exposure, and growth inhibition was found remarkably after that. The algal growth in 9.85 mg/L level was logarithmic, although a little growth inhibition was found. The algal growths in 3.51 and 1.16 mg/L levels were same as control.
- Appearance of the solution: At the start of exposure, test solutions of all exposure levels were colorless and clear. At the end of exposure, the appearance of test solution in 80.4 mg/L level was colorless and clear, that in 28.2 mg/L level was slightly light green, that in 9.85 mg/L level was a little light green and those in other exposure levels were green, due to the algal growth. The solution of control was colorless and clear at the start of exposure, and it was green at the end of exposure due to the algal growth.
- Concentration of test item in test solution: The measured concentrations of test item in the test solutions at the start of exposure were 1.23-80.1 mg/L and those at the end of exposure were 1.10-80.6 mg/L, which were 89.7-102% of the concentrations at the start of exposure.

Results with reference substance (positive control):

The EC50 (0-3 d) of the reference substance was 1.1 mg/L. This value was within the stipulated range (mean ± 2S.D.) [mean ± S.D.: 1.00 ± 0.20 mg/L (n=31, fluorescent lights)] to background data in this laboratory.

Validity criteria fulfilled:

yes

Conclusions:

ErC50 of the test item based on the growth rate was 20.0 mg/L (95% confidence limits: 19.4-20.6 mg/L). NOEC based on the growth rate (0-3 d) was estimated at 3.51 mg/L .

Executive summary:

The objective of this study is to determine the 0-72-hour median effective concentration (EC50) and no observed effect concentration (NOEC) by conducting an algae growth inhibition test of the test item in Pseudokirchneriella subcapiitata according to OECD Guideline 201 (Freshwater Alga and Cyanobacteria, Growth Inhibition Test).

Algal cultures of three replicates were exposed to the substance at 100, 35.7, 12.8, 4.56, 1.63% stock solution content and incubated for 72 hours. Six replicate control cultures were also incubated in the medium without the test item.

 

In 80.4 mg/L levels, growth inhibition was observed remarkably during exposure. In 28.2 mg/L level, algae grew until 24 hours after exposure, and growth inhibition was found remarkably after that. The algal growth in 9.85 mg/L level was logarithmic, although a little growth inhibition was found. The algal growths in 3.51 and 1.16 mg/L levels were same as control.

In 80.4 mg/L levels, some bloated cells and small cells were observed. In 28.2 mg/L level, some cells were bloated. In 9.85 mg/L level, some small cells were observed. The conditions of cells in other exposure were same as the control. In the control, the conditions of cells was not abnormal.

At the start of exposure, test solutions of all exposure levels were colorless and clear. At the end of exposure, the appearance of test solution in 80.4 mg/L level was colorless and clear, that in 28.2 mg/L level was slightly light green, that in 9.85 mg/L level was a little light green and those in other exposure levels were green, due to the algal growth. The solution of control was colorless and clear at the start of exposure, and it was green at the end of exposure due to the algal growth.

 

ErC50 of the test item based on the growth rate was 20.0 mg/L (95% confidence limits: 19.4-20.6 mg/L). NOEC based on the growth rate (0-3 d) was estimated at 3.51 mg/L .

Description of key information

ErC50 of the test item based on the growth rate was 20.0 mg/L (95% confidence limits: 19.4-20.6 mg/L). NOEC based on the growth rate (0-3 d) was estimated at 3.51 mg/L .

Key value for chemical safety assessment

EC50 for freshwater algae:

20 mg/L

EC10 or NOEC for freshwater algae:

3.51 mg/L

Additional information

The objective of this study is to determine the 0-72-hour median effective concentration (EC50) and no observed effect concentration (NOEC) by conducting an algae growth inhibition test of the test item in Pseudokirchneriella subcapiitata according to OECD Guideline 201 (Freshwater Alga and Cyanobacteria, Growth Inhibition Test).

Algal cultures of three replicates were exposed to the substance at 100, 35.7, 12.8, 4.56, 1.63% stock solution content and incubated for 72 hours. Six replicate control cultures were also incubated in the medium without the test item.

 

In 80.4 mg/L levels, growth inhibition was observed remarkably during exposure. In 28.2 mg/L level, algae grew until 24 hours after exposure, and growth inhibition was found remarkably after that. The algal growth in 9.85 mg/L level was logarithmic, although a little growth inhibition was found. The algal growths in 3.51 and 1.16 mg/L levels were same as control.

In 80.4 mg/L levels, some bloated cells and small cells were observed. In 28.2 mg/L level, some cells were bloated. In 9.85 mg/L level, some small cells were observed. The conditions of cells in other exposure were same as the control. In the control, the conditions of cells was not abnormal.

At the start of exposure, test solutions of all exposure levels were colorless and clear. At the end of exposure, the appearance of test solution in 80.4 mg/L level was colorless and clear, that in 28.2 mg/L level was slightly light green, that in 9.85 mg/L level was a little light green and those in other exposure levels were green, due to the algal growth. The solution of control was colorless and clear at the start of exposure, and it was green at the end of exposure due to the algal growth.

 

ErC50 of the test item based on the growth rate was 20.0 mg/L (95% confidence limits: 19.4-20.6 mg/L). NOEC based on the growth rate (0-3 d) was estimated at 3.51 mg/L .