N-butylbenzenesulphonamide

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:

01/01/2001-12/02/2001

Reliability:

2 (reliable with restrictions)

Rationale for reliability incl. deficiencies:

guideline study with acceptable restrictions

Qualifier:

according to guideline

Guideline:

OECD Guideline 201 (Alga, Growth Inhibition Test)

Qualifier:

according to guideline

Guideline:

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

GLP compliance:

yes

Analytical monitoring:

no

Vehicle:

no

Details on test solutions:

PREPARATION AND APPLICATION OF TEST SOLUTION (especially for difficult test substances)
- Method: Preparation of test solution started with a stock solution of 100 mg/l. The stock was treated with ultrasonic waves for approximately 10 minutes to facilitate dissolution. Followed by a short 5-minute stirring period.
- Eluate: /
- Differential loading: /
- Controls: test medium without test substance or other additives.
- Chemical name of vehicle (organic solvent, emulsifier or dispersant): /
- Concentration of vehicle in test medium (stock solution and final test solution(s) including control(s)): /
- Evidence of undissolved material (e.g. precipitate, surface film, etc): no

Test organisms (species):

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

Details on test organisms:

TEST ORGANISM
- Common name: Fresh Water Algal - Selenastrum capricornutum
- Strain: CCAP 278/4
- Source (laboratory, culture collection): 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.
- Age of inoculum (at test initiation): 4 days before the start of the test, cells from the algal stock culture were inoculated in culture medium at a cell density of 2.10E4 cells/ml. The pre-culture was maintained under the same conditions as used in the test. The cell density was measured immediately before use.
- Method of cultivation:


ACCLIMATION
- Acclimation period:4 days
- Culturing media and conditions (same as test
- Any deformed or abnormal cells observed:no

Test type:

static

Water media type:

freshwater

Limit test:

no

Total exposure duration:

72 h

Post exposure observation period:

During incubation the algal cells were kept in suspension by continuous shaking.

Hardness:

24 mg CaCO3 /L

Test temperature:

22,3 - 23,5°c

pH:

8,0 - 8,3

Dissolved oxygen:

no data

Nominal and measured concentrations:

2.2,4.6,10,22,46,and 100 mg/L nominal , no measured concentrations

Details on test conditions:

TEST SYSTEM
- Test vessel: 100 ml, all-glass
- Type (delete if not applicable): open
- Material, size, headspace, fill volume: /
- Aeration: /
- Type of flow-through (e.g. peristaltic or proportional diluter): /
- Renewal rate of test solution (frequency/flow rate):/
- Initial cells density: 1*10E4 cells/ml
- Control end cells density: 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.
- No. of organisms per vessel:/
- No. of vessels per concentration (replicates):/
- No. of vessels per control (replicates):/
- No. of vessels per vehicle control (replicates):/


GROWTH MEDIUM
- Standard medium used: yes
- Detailed composition if non-standard medium was used:


TEST MEDIUM / WATER PARAMETERS
- Source/preparation of dilution water:tap water purified by reverse osmosis and then passed over activated carbon and ion-exchange cartridges.
- Total organic carbon:/
- Particulate matter:/
- Metals:/
- Pesticides:/
- Chlorine:/
- Alkalinity:/
- Ca/mg ratio: hardness: 0.24 mmol/l
- Conductivity:/
- Culture medium different from test medium:/
- Intervals of water quality measurement: /


OTHER TEST CONDITIONS
- Sterile test conditions: yes/
- Adjustment of pH:/
- Photoperiod:continuously
- Light intensity and quality: TLD-lamps of the type 'cool-white' of 30 watt (Sylvania), with a light intensity within the range of 60 to 120 µE.m-2.s-1, not varying by more than 202%.
- Salinity (for marine algae):/


EFFECT PARAMETERS MEASURED (with observation intervals if applicable) :
- Determination of cell concentrations: [counting chamber
- Chlorophyll measurement:/
- Other: 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.2H2O: 100 µg/l
MnCl2.4H2O: 415 µg/l
ZnCl2: 3µg/l
CoCl2.6H2O: 1,5 µg/l
CuCl2.2H2O: 0,01 µg/l
Na2MoO4.2H2O: 7µg/l
NaHCO3: 50 mg/l


TEST CONCENTRATIONS
- Spacing factor for test concentrations:
- Justification for using less concentrations than requested by guideline: /
- Range finding study
- Test concentrations: a range of 0,1 to 100 mg/l, increasing by a factor 10.
- Results used to determine the conditions for the definitive study:based on the result of the range-finding test the EC50 for effects on algal growth was expected to lie between 10 and 100 mg/l.

Reference substance (positive control):

yes

Remarks:

K2Cr2O7

Duration:

72 h

Dose descriptor:

EC50

Effect conc.:

49 mg/L

Nominal / measured:

nominal

Conc. based on:

test mat.

Basis for effect:

cell number

Remarks:

cell growth inhibition

Remarks on result:

other: 16 to 82 mg/l

Key result

Duration:

72 h

Dose descriptor:

EC50

Effect conc.:

83 mg/L

Nominal / measured:

nominal

Conc. based on:

test mat.

Basis for effect:

growth rate

Remarks:

growth rate reduction

Remarks on result:

other: 69 to 97 mg/l

Duration:

72 h

Dose descriptor:

NOEC

Effect conc.:

22 mg/L

Nominal / measured:

nominal

Conc. based on:

test mat.

Basis for effect:

cell number

Remarks:

cell growth inhibition

Duration:

72 h

Dose descriptor:

NOEC

Effect conc.:

10 mg/L

Nominal / measured:

nominal

Conc. based on:

test mat.

Basis for effect:

growth rate

Remarks:

growth rate reduction

Duration:

72 h

Dose descriptor:

EC10

Effect conc.:

9.1 mg/L

Nominal / measured:

nominal

Conc. based on:

test mat.

Basis for effect:

cell number

Remarks:

cell growth inhibition

Key result

Duration:

72 h

Dose descriptor:

EC10

Effect conc.:

20 mg/L

Nominal / measured:

nominal

Conc. based on:

test mat.

Basis for effect:

growth rate

Remarks:

growth rate reduction

Details on results:

- Exponential growth in the control (for algal test): yes
- Observation of abnormalities (for algal test): no
- Unusual cell shape:
- Colour differences:
- Flocculation:
- Adherence to test vessels:
- Aggregation of algal cells:
- Other:
- Any stimulation of growth found in any treatment:
- 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

Results with reference substance (positive control):

EbC50 :0.59mg/L and ErC50:1.1mg/L

Reported statistics and error estimates:

An effect was considered to be significant if statistical analysis of the data obtained for the test concentration compared with those obtained in the negative control revealed significant reduction of growth rate( ANOVA, tukey test, bonferroni t-test, TOXSTAT release 3.5,1996,DD Gulley
AMBoelter,HL bergman) Additionally the EC10 was determined to meet the recommendations as put down in " A review of Statistical Data Analysis and experimental design in OECD aquatic Toxicology Test Guidelines By S Pack August 1993.calculation of the EC50 and the EC 10 values
was based on linear regression analysis of the % of growth inhibition and the % growth rate reduction versus the corresponding nominal
concentrations of the test substance.

Validity criteria fulfilled:

yes

Conclusions:

Under the conditions of the present study with Selenastrum capricornutum, BBSA reduced growth rate of this fresh water algae species significantly at nominal concentrations of 22 mg/l and higher.
The EC50 for cell growth inhibition (EBC50: 0-72h) was 49 mg/l with a 95% confidence interval ranging from 16 to 82 mg/l. The EC10 for cell growth inhibition (EBC10: 0-72h) was 9,1 mg/l with a 95% confidence interval ranging from 0 to 42 mg/l.
Since growth rate is derived from the slope under the growth curve in a logarithmic plot, the measure of the specific growth rate is preferable over biomass following from the mathematical nature of exponential growth.
The EC50 for growth rate reduction (ERC50: 0-72h) was 83 mg/l with a 95% confidence interval ranging from 69 to 97 mg/l. The EC10 for growth rate reduction (ERC10:0-72h) was 20 mg/l with a 95% confidence interval ranging from 6,6 to 33 mg/l.
The NOEC for cell growth inhibition was determined to be 22 mg/l and the NOEC for growth rate reduction was 10 mg/l.

Description of key information

The cell growth and growth rate inhibition In of Selenastrum capricornutumwere measured in a standard OECD 201 test study. The reported concentrations are nominal concentrations.

The EC50 for cell growth inhibition (EBC50-72h) was 49 mg/l. The EC10 for cell growth inhibition (EBC10-72h) was 9,1 mg/l.

The EC50 for growth rate reduction (ERC50-72h) was 83 mg/l. The EC10 for growth rate reduction (ERC10-72h) was 20 mg/l.

The NOEC for cell growth inhibition was determined to be 22 mg/l and the NOEC for growth rate reduction was 10 mg/l.

Key value for chemical safety assessment

EC50 for freshwater algae:

83 mg/L

EC10 or NOEC for freshwater algae:

20 mg/L