Benzoyl chloride, 4-[[(2-methoxybenzoyl)amino]sulfonyl]-

Toxicity to aquatic algae and cyanobacteria

Link to relevant study record(s)

Reference

Endpoint:

effects on growth of green algae

Type of information:

experimental study

Adequacy of study:

key study

Study period:

17 Jun - 11 Jul 2006

Reliability:

1 (reliable without restriction)

Rationale for reliability incl. deficiencies:

guideline study

Qualifier:

according to guideline

Guideline:

EU Method C.3 (Algal Inhibition test)

Deviations:

no

GLP compliance:

yes (incl. QA statement)

Analytical monitoring:

yes

Remarks:

HPLC-UV/VIS

Details on sampling:

- Concentrations: 1.9, 4.3, 9.4, 20.7, 45.4 and 100 mg/L
- Sampling method: Aliquots of the samples from the biological test were directly analysed by HPLC
- Sample storage conditions before analysis: Routinely, the samples are analysed immediately. Only in exceptional cases, they are stored overnight deeply frozen and protected from light.

Vehicle:

no

Details on test solutions:

PREPARATION AND APPLICATION OF TEST SOLUTION
- Method: A stock solution was prepared to give the desired series of test concentrations. To
achieve this 124.9 mg of the test item were added to 1 L of dilution water and
treated for 1 h in an ultrasonic bath and afterwards stirred for 24 h on a magnetic
stirrer. Finally undissolved particles of the test item were removed by filtration using a
folded filter of pore size 7-12 µm.
- Controls: Yes
- Test concentration separation factor: 2.2
- Evidence of undissolved material: undissolved particles of the test item were removed by filtration

Test organisms (species):

Desmodesmus subspicatus (previous name: Scenedesmus subspicatus)

Details on test organisms:

TEST ORGANISM
- Common name: green algae
- Source and strain: Non-axenic strain of the test species obtained from the collection of Algal Cultures of the Institute of Plant Physiology at the University of Gottingen (Germany)
- Method of cultivation: Exponentially-growing stock cultures are maintained in the test facility under constant temperature conditions (23 +/- 2 °C) at a light intensity in the range 60 - 120
µE. x m-2 x s-1 (measured in the range 400 to 700 nm using a spherical quantum flux meter). The nutrient medium (according to BRINGMANN & KUHN (1977) is renewed once a week. Cell density measurements are made using a microcell counter

ACCLIMATION
- Acclimation period: Pre-cultures are set up three days before the start of a test.
- Culturing media and conditions: They are grown under identical exposure conditions as the stock cultures, except from the use of a different nutrient medium

Test type:

not specified

Water media type:

freshwater

Limit test:

no

Total exposure duration:

72 h

Test temperature:

21 °C to 25 °C

pH:

7.9 to 8.2

Nominal and measured concentrations:

Nominal: 1.9, 4.3, 9.4, 20.7, 45.4 and 100 mg/L
Measured: 1.75, 3.85, 8.95, 18.9, 43.5, 95.9 mg/L at 72 h

Details on test conditions:

TEST SYSTEM
- Test vessel: 300 mL Erlenmeyer flasks with cotton stoppers
- Initial cells density: 5000 cells/mL
- Control end cells density: 430000 cells/mL
- No. of vessels per concentration: 3
- No. of vessels per control: 6

GROWTH MEDIUM
- Standard medium used: yes
- Detailed composition if non-standard medium was used: Nutrient medium recommended in guidance 92/69/EEC C.3 was used.

OTHER TEST CONDITIONS
- Sterile test conditions: no
- Adjustment of pH: no
- Light intensity and quality: 4000 to 8000 Ix

EFFECT PARAMETERS MEASURED:
- Determination of cell concentrations: microscopic counting chamber

TEST CONCENTRATIONS
- Spacing factor for test concentrations: 2.2

Reference substance (positive control):

no

Key result

Duration:

72 h

Dose descriptor:

NOEC

Effect conc.:

9.4 mg/L

Nominal / measured:

nominal

Conc. based on:

test mat.

Basis for effect:

growth rate

Duration:

72 h

Dose descriptor:

EC10

Effect conc.:

99 mg/L

95% CI:

> 51 - < 2 116

Nominal / measured:

nominal

Conc. based on:

test mat.

Basis for effect:

growth rate

Duration:

72 h

Dose descriptor:

EC20

Effect conc.:

401 mg/L

95% CI:

> 145 - < 4 675 017

Nominal / measured:

nominal

Conc. based on:

test mat.

Basis for effect:

growth rate

Duration:

72 h

Dose descriptor:

NOEC

Effect conc.:

9.4 mg/L

Nominal / measured:

nominal

Conc. based on:

test mat.

Basis for effect:

biomass

Duration:

72 h

Dose descriptor:

EC10

Effect conc.:

18 mg/L

95% CI:

> 1.8 - < 32

Nominal / measured:

nominal

Conc. based on:

test mat.

Basis for effect:

biomass

Duration:

72 h

Dose descriptor:

EC50

Effect conc.:

253 mg/L

95% CI:

> 121 - < 5 788

Nominal / measured:

nominal

Conc. based on:

test mat.

Basis for effect:

biomass

Details on results:

- Exponential growth in the control: yes
- Observation of abnormalities: no

Reported statistics and error estimates:

- EC 50, EC 20 and EC 10 values for both, biomass [b] and growth rate [r], are calculated by probit analysis
- NOEC of both, biomass [b] and growth rate [r], are determined by a multisample comparison (according to DUNNETT 1955, 1964)

 

Mean growth rate and percentage change in growth rate

Nominal concentration [mg/L]	Growth rate [1/d]	Inhibition (+) / Increase (-) [%]
Control	1.51	0
1.9	1.53	-1.2
4.3	1.53	-1.3
9.4	1.52	-0-4
20.7	1.43	5.6
45.5	1.39	7.9
100	1.38	8.9

 

Mean biomass and percentage change in biomass

Nominal concentration [mg/L]	Area under growth curve	Inhibition (+) / Increase (-) [%]
Control	314167	0
1.9	339444	-8.0
4.3	340556	-8.4
9.4	322778	-2.7
20.7	266667	15.1
45.5	231667	26.3
100	217778	30.7

 

Validity criteria for the measurement of the algae toxicity

Target condition according to guideline:	Actual condition according to the study:	Validity criteria met:
Exponentially growing test organisms are exposed to the test substance in batch cultures over a period of normally 72 hours.	The algal inocula for a test are taken from an exponentially-growing pre-culture	Yes
The biomass in the control cultures should have increased exponentially by a factor of at least 16 within the 72-hour test period. This corresponds to a specific growth rate of 0.92 day-1.	The growth rate in control after 72 h was at least 1.48	Yes
The cultures should be maintained at a temperature in the range of 21 to 25°C, controlled at ± 2°C.	Temperature was mainted between 21 to 25°C.	Yes
For the final definitive test at least five concentrations, arranged in a geometric series with a factor not exceeding 3.2, should be selected. (For test substances showing a flat concentration response curve a higher factor may be justified. The concentration series should preferably cover the range causing 5-75 % inhibition of algal growth rate.)	6 concentrations with a spacing factor of 2.2 was used	Yes
The concentration of the test substance shall be maintained to within 80 % of the initial concentrations throughout a time corresponding to the duration of the test	Recovery rates ranged from 92 - 101.5 % of nominal values at 0 hours, and from 89.5 - 96% of nominal values at 72 hours, respectively.	Yes

Validity criteria fulfilled:

yes

Remarks:

See more details in the table in the section "Any other information on results incl. tables"

Conclusions:

Algae growth rate inhibition was observed; 72 h NOEC = 9.4 mg/L

Executive summary:

In a 72 h toxicity study, the cultures of Desmodesmus subspicatus were exposed to the test substance at nominal concentrations of 1.9, 4.3, 9.4, 20.7, 45.4 and 100 mg/L under static in accordance with the EEC Methods for Determination of Ecotoxicity Annex to Directive 92/69/EEC (OJ. No. L383A, 29.12.92) Part C, Method 3 'Algal inhibition test'. The NOEC and EC10 values based on growth rate were 9.4 and 99 mg/L, respectively. The NOEC was used as key value because on the unreliable wide confidence interval of the estimated EC10 value. The percentage growth rate inhibition in the treated algal culture as compared to the control ranged from -1.2 to 8.9 %. The results are expressed in terms of nominal concentrations. The test substance is an acid chloride which easily hydrolyses to the corresponding acid in aqueous media. Therefore the acid was analysed by HPLC. Recovery rates ranged from 92 - 101.5 % of nominal values at 0 hours, and from 89.5 - 96% of nominal values at 72 hours, respectively.

Description of key information

In a 72 h toxicity study, the cultures of Desmodesmus subspicatus were exposed to the test substance at nominal concentrations of 1.9, 4.3, 9.4, 20.7, 45.4 and 100 mg/L under static in accordance with the EEC Methods for Determination of Ecotoxicity Annex to Directive 92/69/EEC (OJ. No. L383A, 29.12.92) Part C, Method 3 'Algal inhibition test'. The NOEC and EC10 values based on growth rate were 9.4 and 99 mg/L, respectively. The NOEC was used as key value because on the unreliable wide confidence interval of the estimated EC10 value. The percentage growth rate inhibition in the treated algal culture as compared to the control ranged from -1.2 to 8.9 %. The results are expressed in terms of nominal concentrations. The test substance is an acid chloride which easily hydrolyses to the corresponding acid in aqueous media. Therefore the acid was analysed by HPLC. Recovery rates ranged from 92 - 101.5 % of nominal values at 0 hours, and from 89.5 - 96% of nominal values at 72 hours, respectively.

Key value for chemical safety assessment

EC10 or NOEC for freshwater algae:

9.4 mg/L