2-(2H-benzotriazol-2-yl)-4,6-bis(1-methyl-1-phenylethyl)phenol

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:

1993-09-03 to 1993-09-06

Reliability:

2 (reliable with restrictions)

Rationale for reliability incl. deficiencies:

other: see 'Remark'

Remarks:

Non-GLP-guideline study. Deviations to the updated OECD Guideline (adopted 2006): The coefficient of analysis of the daily growth rate in the control replicates and the coefficient of analysis of the average specific growth rates in the control replicates were not reported. No analytical confirmation.

Qualifier:

according to guideline

Guideline:

OECD Guideline 201 (Alga, Growth Inhibition Test)

Deviations:

yes

Remarks:

no analytical dose verification

GLP compliance:

yes

Analytical monitoring:

no

Vehicle:

yes

Details on test solutions:

PREPARATION AND APPLICATION OF TEST SOLUTION (especially for difficult test substances)
- Method: 1, 10 and 100 mg/L working standards were prepared by quantitatively transferring 0.0005, 0.005 and 0.05 g, respectively, of test substance into 500 mL volumetric flasks, and each was brought to volume with sterile nutrient media. After each working standard was brought to volume, 0.05 mL of DMF was injected into each. Each working standard was then sonicated for 30 minutes and then magnetically stirred for 30 minutes. Vehicle blank was prepared by injecting a 0.01 mL aliquotof DMF into each vehicle blank replicate containing 100 mL of algal nutrient media.
- Chemical name of vehicle (organic solvent, emulsifier or dispersant): DMF
- Concentration of vehicle in test medium: 0.01%
- Evidence of undissolved material (e.g. precipitate, surface film, etc): after preparation, the 1 mg/L solution was clear, the 10 mg/L soultion was clear with a precipitate and the 100 mg/L solution was cloudy wiht a precipitate

Test organisms (species):

Desmodesmus subspicatus (previous name: Scenedesmus subspicatus)

Details on test organisms:

TEST ORGANISM
- Common name: Green Algae
- Source (laboratory, culture collection): Department of Botany, Culture collection of Algae, University of Texas at Austin
- Age of inoculum (at test initiation): 3 days old
- Method of cultivation: in 250 mL Erlenmeyer flasks containing 100 mL of sterile algae nutrient media. Flasks were incubated in environmental chamber at 24 ± 2°C. Periodically, new cultures were initiated using a lot of parent stock or cloned from an existing culture derived from the parent stock in 100 mL of sterile culture medium.

ACCLIMATION
- Acclimation period: 3 days under test conditions.
- Culturing media and conditions (same as test or not): same as test
- Any deformed or abnormal cells observed: none reported

Test type:

static

Water media type:

freshwater

Limit test:

no

Total exposure duration:

72 h

Hardness:

no data applicable

Test temperature:

23°C

pH:

7.4 -7.6

Dissolved oxygen:

7.4 mg/L

Salinity:

no data applicable

Nominal and measured concentrations:

1, 10 and 100 mg test substance/L (nominal)

Details on test conditions:

TEST SYSTEM
- Test vessel: 250 mL Erlenmeyer flasks, constant rotary agitation (100 rpm), containing 100 mL of algae nutrient media or test solution
- - Material, size, headspace, fill volume: 250 mL filled with 100 mL test solution
- Initial cells density: approx. 10,000 cells/mL (measured 9300 cells/mL for control, 8500 cells/mL for vehicle blank)
- Control end cells density: 690000 cells/mL
- No. of vessels per concentration (replicates): 3
- No. of vessels per control (replicates): 3
- No. of vessels per vehicle control (replicates): 3

GROWTH MEDIUM
- Standard medium used: yes

OTHER TEST CONDITIONS
- Sterile test conditions: no
- Adjustment of pH: none
- Photoperiod: continuous
- Light intensity and quality: cool white fluorescent light, 800 ± 10% ft-c (approx. 8600 lux)

EFFECT PARAMETERS MEASURED : cell density
- Determination of cell concentrations: Cell densities were measured at 24, 48 and 72 hours exposure by hemacytometer and using an Olympus Model BH-2 microscope
- Other: Temperature was continuously measured and maintained at 23 + 0.5°C. pH was measured at 0h exposure

TEST CONCENTRATIONS
- Spacing factor for test concentrations: 10
- Test concentrations: 1, 10 and 100 mg test substance/L

Reference substance (positive control):

no

Duration:

72 h

Dose descriptor:

EC50

Effect conc.:

> 100 mg/L

Nominal / measured:

nominal

Conc. based on:

test mat.

Remarks:

Values are based on nominal concentrations

Basis for effect:

growth rate

Duration:

72 h

Dose descriptor:

NOEC

Effect conc.:

100 mg/L

Nominal / measured:

nominal

Conc. based on:

test mat.

Remarks:

Values are based on nominal concentrations

Basis for effect:

growth rate

Remarks on result:

other: 5% level

Details on results:

- Exponential growth in the control (for algal test): yes (74-fold increase in control, 80-fold increase in vehicle blank)
- No toxicity observed in the range of solubility.
- Any stimulation of growth found in any treatment: after 72 hours, algal cell counts for 1, 10 and 100 mg/L were 104, 94 and 94% of the pooled control population, respectively.
- Any observations (e.g. precipitation) that might cause a difference between measured and nominal values: a precipitate was observed in the 10 and 100 mg/L test replicates at 72 hours

Reported statistics and error estimates:

Student t-test
ANOVA and Dunnett's multiple means Test

Description of key information

With high probability acutely not harmful to aquatic algae and cyanobacteria. No toxic effects occur at the range of solubility.

Additional information

In a 72h non-GLP guideline study using Scenedesmus subspicatus, an EC50 of > 100 mg/L for biomass was determined according to OECD 201.

After 72 hours, algal cell counts for the 1.0 , 10, and 100 mg/L test levels were 104, 94, and 94% of the pooled control population, respectively. The NOEC based on cell number was > 100 mg/L. DMF was used as a solvent with a concentration of 0.01% in test medium.

The effect concentration is well above the solubility limit of the test substances (water solubility < 0.005 mg/L at 20°C, see chapter 4.8 in IUCLID). Therefore, it represents a worst case assumption where water solubility is achieved by use of solvent. In conclusion, the substance shows no effect in the range of water solubility.