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Ecotoxicological information

Toxicity to aquatic algae and cyanobacteria

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Administrative data

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:
other: Well conducted Guideline study conducted under GLP

Data source

Reference
Reference Type:
study report
Title:
Unnamed
Year:
1992
Report Date:
1992

Materials and methods

Test guideline
Qualifier:
according to
Guideline:
OECD Guideline 201 (Alga, Growth Inhibition Test)
Deviations:
no
GLP compliance:
yes

Test material

Reference
Name:
Unnamed
Type:
Constituent
Type:
Constituent
Details on test material:
CC(C)C(=O)OCC(C)(C)C(O)C(C)C
CC(C)C(=O)OC(C(C)(C))C(C)(C)CO

Sampling and analysis

Analytical monitoring:
yes
Details on sampling:
- Concentrations: 2.5, 5, 10, 20, 40 and 80 mg/L nominal
- Sampling method: Samples were analyzed for the actual concentration of test substance present in the test solutions at 0 hours and 72 hours. Samples for the 0 hour analyses were portions of the test treatments used to begin the test. At test termination, the contents of the replicate flasks were combined, the pH recorded, and the algae were removed by centrifugation at 3700 rpm for four minutes. The supernatants were then submitted for analyses.

Test solutions

Vehicle:
no

Test organisms

Test organisms (species):
Pseudokirchneriella subcapitata (previous names: Raphidocelis subcapitata, Selenastrum capricornutum)
Details on test organisms:
TEST ORGANISM
- Source (laboratory, culture collection): Lab cultures originating fromthe University of Texas Culture Collection (UTEX #1648), Austin, TX.
- Age of inoculum (at test initiation): The culture used to prepare the inoculum for testing was actively growing at the test conditions for at least two transfers prior to the start of the definitive test.
- Method of cultivation: liquid algal medium


ACCLIMATION
- Acclimation period: Several previous passages were performed in order to confirm expotential growth under the conditions of the test.
- Culturing media and conditions (same as test or not): same
- Any deformed or abnormal cells observed: none reported

Study design

Test type:
static
Water media type:
freshwater
Limit test:
no
Total exposure duration:
72 h

Test conditions

Test temperature:
24 +or-2 C
pH:
Conc. Initial Final
Control 7.45 7.63
2.5 7.47 7.79
5.0 7.46 7.82
10 7.46 7.81
20 7.39 7.78
40 7.40 7.75
80 7.38 7.78
Nominal and measured concentrations:
Nominal Measured (mean)
2.5 1.10
5 3.28
10 7.28
20 14.4
40 29.1
80 57.0
Details on test conditions:
TEST SYSTEM
- Test vessel:
- Type (delete if not applicable): open (with foam stoppers)
- Material, size, headspace, fill volume: 125 mL glass Erlenmeyer flasks, ~25 mL of algal growth medium
- Initial cells density: 1e4 cells/mL
- Control end cells density: 2.44e6 cells/mL
- No. of vessels per concentration (replicates): 3
- No. of vessels per control (replicates): 3



GROWTH MEDIUM
- Standard medium used: yes



TEST MEDIUM / WATER PARAMETERS
- Source/preparation of dilution water: Type I water
- Intervals of water quality measurement: intial and final test solutions


OTHER TEST CONDITIONS
- Sterile test conditions: yes
- Adjustment of pH: none
- Photoperiod: continuous light
- Light intensity and quality: 8000 ± 1200 lumens/m2



EFFECT PARAMETERS MEASURED (with observation intervals if applicable) : Biomass @ 24, 48 and 72 hours
- Determination of cell concentrations: Coulter Counter



TEST CONCENTRATIONS
- Spacing factor for test concentrations: 0.5x

Results and discussion

Effect concentrationsopen allclose all
Duration:
72 h
Dose descriptor:
EC50
Effect conc.:
> 57 mg/L
Nominal / measured:
meas. (arithm. mean)
Conc. based on:
test mat.
Basis for effect:
growth rate
Duration:
72 h
Dose descriptor:
NOEC
Effect conc.:
7.28 mg/L
Nominal / measured:
meas. (arithm. mean)
Conc. based on:
test mat.
Basis for effect:
growth rate
Duration:
72 h
Dose descriptor:
EC50
Effect conc.:
18.4 mg/L
Nominal / measured:
meas. (arithm. mean)
Conc. based on:
test mat.
Basis for effect:
biomass
Remarks on result:
other: 14.6-23.3 mg/L
Duration:
72 h
Dose descriptor:
NOEC
Effect conc.:
3.28 mg/L
Nominal / measured:
meas. (arithm. mean)
Conc. based on:
test mat.
Basis for effect:
biomass
Details on results:
- Exponential growth in the control (for algal test): yes
- Observation of abnormalities (for algal test): none
- Any stimulation of growth found in any treatment: none
- Any observations (e.g. precipitation) that might cause a difference between measured and nominal values: no
- Effect concentrations exceeding solubility of substance in test medium: no
Reported statistics and error estimates:
The concentration-effect relationship was assessed by comparison of the areas under the growth curves. The mean values of the cell counts for each test concentration and for the control is plotted against time to produce growth curves. The percent inhibition is calculated as the difference between the area under the control growth curve and the area under the growth curve at each test concentration. To determine the EbC50 value and associated 95% confidence limits, weighted least squares nonlinear regression of the log of test concentration against the area under the growth curve was performed. The NOEC was determined, based upon the mean cell counts at 72 hours, from an analysis of variance (ANOVA) and Dunnett's test. Statistical analyses were performed using SAS Software (SAS Institute, Cary, NC). The level of significance was at alpha = 0.05. The average specific growth rate is derived from the slope of the regression line of a plot of the natural log of the cell counts versus time. The percent inhibition is calculated as the difference between the growth rate of the controls and the growth rate for each test concentration. Statistical analyses were not performed to estimate the ErC50 value because greater than 50% inhibition in growth rate was not measured in this study. The NOEC for growth rate was estimated using one-sided t-test and calculated using the Microsoft Excel statistical tool package. The level of significance was at alpha = 0.05.

Any other information on results incl. tables

 Table 1: Data used for Calculation of Specific Growth Rate and Biomass

 Cell counts (cells/mL) during test

       Measured Concentrations (mg/L)
Time 1.10 3.28 7.28 14.40 29.10 57.00 Control
0 10000 10000 10000 10000 10000 10000 10000
24 41637 33253 33923 38153 38613 27290 35650
48 453147 406600 312833 219020 179407 113823 466340
72 2602133 2143467 1876333 1468000 1019200 559733 2445333

  Calculation of ln (cells/mL)

Measured Concentrations (mg/L)        
Time 1.1 mg/L 3.28 mg/L 7.28 mg/L 14.4 mg/L 29.1 mg/L 57 mg/L Control
0 9.2 9.21 9.21 9.21 9.21 9.21 9.21
1 10.64 10.41 10.43 10.55 10.56 10.21 10.48
2 13.02 12.92 12.65 12.30 12.10 11.64 13.05
3 14.77 14.58 14.44 14.20 13.83 13.24 14.71

Calculated Specific Growth Rate (u) & Percent Inhibition

Concentration (mg/L) Growth Rate Percent Inhibition
Control 1.90690  
1.1 1.90720 -0.02
3.28 1.86060 2.43
7.28 1.79250 6.00
14.4 1.67150 12.34
29.1 1.54090 19.19
57 1.35030 29.19

ErC50 (0 -72 h) > 57 mg/L

NOEC (growth rate) = 7.28 mg/L

Applicant's summary and conclusion

Validity criteria fulfilled:
yes
Executive summary:

An algal growth inhibition test was conducted to determine the EbC50 value, and the NOEC, if possible, of Texanol Ester-Alcohol for the green alga, Selenastmm capricornutum. Selenastmm capricornutum was exposed over a 72-hour period to six concentrations (2.5 to 80 mg/L nominal). Results are based upon the mean measured concentrations, which ranged from 1.10 to 57.0 mg/L. Biomass was determined by cell counts at 24, 48 and 72 hours. The EbC50 values were determined by weighted least squares nonlinear regression of the log of concentration against the area under the growth curves for each time interval. The 72-hour EbC50 was determined to be 18.4 mg/L (95% confidence limits 14.6 - 23.3 mg/L). The no observed effect concentration (NOEC), based upon the mean cell count values at 72 hours, was determined by an analysis of variance and Dunnett's test to be 3.28 mg/L. A subsequent evalaution of the data was performed to determine the ErC50 and NOEC for growth rate. The ErC50 was determined to be > 57 mg/L (the highest concentration tested), and the NOEC for growth rate was determined to be 7.28 mg/L.