Benzenesulfonic acid, 2,2'-(1,2-ethenediyl)bis[5-nitro-, disodium salt, reaction products with 4-[(4-aminophenyl)azo]benzenesulfonic acid monosodium salt

Toxicity to aquatic algae and cyanobacteria

Link to relevant study record(s)

Reference

Reference 1

Endpoint:

toxicity to aquatic algae and cyanobacteria

Type of information:

other: read-across from supporting substance (structural analogue or surrogate)

Adequacy of study:

key study

Reliability:

2 (reliable with restrictions)

Rationale for reliability incl. deficiencies:

guideline study with acceptable restrictions

Remarks:

Source study has reliability 2. Details on the read across are attached in section 13.

Qualifier:

according to guideline

Guideline:

OECD Guideline 201 (Alga, Growth Inhibition Test)

GLP compliance:

not specified

Test organisms (species):

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

Details on test organisms:

TEST ORGANISM
- Source (laboratory, culture collection): culture collection algae and protoyoo (Cumbria, Reino Unido)

ACCLIMATION
- Temperature: 22 ± 2 °C
- Light intensity: 2500 Lux.

Test type:

flow-through

Water media type:

not specified

Limit test:

no

Total exposure duration:

72 h

Test temperature:

22 ± 2 °C

pH:

pH at start: 7.20
pH at the end: 7.03
pH minimum concentration at start: 7.19
pH minimum concentration at the end: 7.17
pH maximum concentration at start: 7.49
pH maximum concentration at the end: 7.22

Nominal and measured concentrations:

Nominal concentrations are: 10 ,17, 29, 37, 48, 62, 100 mg/l
Actual concentrations are: 10.1, 17.2, 29.3, 37.3, 48.4, 62.6, 100.9 mg /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.

Basis for effect:

growth rate

Cells concentrations

replications	control		concentration (mg/l)
no. cells/ml	0h	72h	10	17	29	37	48	62	100
1	4.10 10^4	6.70 10^5	6.73 10^5	6.05 10^5	5.68 10^5	5.77 10^5	5.64 10^5	5.38 10^5	4.98 10^5
2	3.90 10^4	6.90 10^5	5.73 10^5	6.11 10^5	5.72 10^5	5.73 10^5	5.60 10^5	5.36 10^5	4.96 10^5
Promedio	4.00 10^4	6.80 10^5	6.23 10^5	6.08 10^5	5.70 10^5	5.75 10^5	5.62 10^5	5.37 10^5	4.97 10^5
sd	1414	14142	70711	4243	2828	2828	2828	1414	1414
CV	4	2	11	1	0.5	0.5	1	0.3	0.3

% growth inhibition: defined as biomass at the end of the exposition period (72h) minus the initial biomass (0h).

conc. (mg/l)	% growth inhibition
control	0
10	8.9
17	11.3
29	17.2
37	16.4
48	18.4
62	22.3
100	28.6

Validity criteria fulfilled:

yes

Conclusions:

The substance was tested according to OECD 201.
ErC50 (72h) > 100 mg/l.

Executive summary:

Method

Inhibition on the algae growth was assessed exposing the unicellular alga Pseudokirchneriella subcapitata (Selenastrum capricornutum) to test substance according to OECD guideline 201 (1984). Nominal concentrations tested were 10, 17, 29, 37, 48, 62 and 100 mg/l.

Results

At the end of the 72 -hour exposure period, effect on growth rate was ca. 29 % at the highest tested dose of 100 mg/l. Therefore, ErC50 (72h) > 100 mg/l.

Reference 2

Endpoint:

toxicity to aquatic algae and cyanobacteria

Type of information:

other: read-across from supporting substance (structural analogue or surrogate)

Adequacy of study:

key study

Study period:

1991

Reliability:

2 (reliable with restrictions)

Rationale for reliability incl. deficiencies:

guideline study

Remarks:

Source study has reliablity 1. Details on the read across are attached in section 13.

Qualifier:

according to guideline

Guideline:

OECD Guideline 201 (Alga, Growth Inhibition Test)

Deviations:

no

GLP compliance:

yes

Analytical monitoring:

yes

Details on sampling:

- Concentrations: 62.3, 250, 1000 mg/l
- Sampling method: The stock solution of the biological test was diluted 1000fold with test medium. Due to precipitation effects, samples with a nominal concentration of 250 mg/l and 1000 mg/l must have been filtered first. Samples with a nominal concentration of 62.3 mg/l and 250 mg/l were diluted 10fold with test medium. Samples with a nominal concentration of 1000 mg/l were diluted 100fold with test medium.
- Sample storage conditions before analysis: the samples were stored deep-frozen (-20 °C) until the analysis was performed.

Vehicle:

no

Test organisms (species):

Desmodesmus subspicatus (previous name: Scenedesmus subspicatus)

Details on test organisms:

TEST ORGANISM
- Scenedesmus subspicatus CHODAT
- Common name: unicellular green algal species.
- Strain: No. 8681 SAG
- Source (laboratory, culture collection): Sammlung von Algenkulturen, Pflanzenphysiologisches Institut der Universität, D-3400 Göttingen, F.R:G.
- Age of inoculum (at test initiation): algae were grown in the laboratories of RCC
- Method of cultivation: under standardized conditions according to the OECD guideline 201.

ACCLIMATION
- Acclimation period: 72h
- Culturing media and conditions (same as test or not): 10000 cells per ml test solution were taken from a sterile pre-culture, at the same conditions as in the test.

Test type:

static

Water media type:

not specified

Limit test:

no

Total exposure duration:

96 h

Post exposure observation period:

Samples of 1 ml test solutions were sterily taken out of all flasks after 24, 48, 72 and 96 hours of exposure.

Hardness:

n.a.

Test temperature:

In the test media during the test duration temperature was 21.0 - 23.4 °C.

pH:

At the start of the test pH-values in the test media were in range of pH 7.8 to 7.9, at the end of the test pH-values were meausured between pH 8.2 and 10.3.

Dissolved oxygen:

n.a.

Salinity:

n.a.

Conductivity:

n.a.

Nominal and measured concentrations:

Nominal concentrations tested were 62.3, 125, 250, 500 and 1000 mg test substance/l and a control without any additions. The concentrations in the analysed test media of the concentrations 62.3 and 1000 mg/l were in the range of 92.8 to 103.9% of the nominal values. The mesured concentration in the stock solution amounted to 105.5% of the nominal value.

Details on test conditions:

TEST SYSTEM
- Test vessel: 250 ml Erlenmeyer flasks stoppered with permaeble plugs.
- Test solutions were incubated on a laboratory shaker in an air-conditioned room
- Initial cells density: 10000 cells/ml
- No. of vessels per concentration (replicates): 3

GROWTH MEDIUM
- Standard medium used: yes

TEST MEDIUM / WATER PARAMETERS
- Source/preparation of dilution water: according to OECD guideline 201

OTHER TEST CONDITIONS
- Adjustment of pH: 7.8 at the begining of the test
- Photoperiod: continuous illumination
- Light intensity and quality: 8000 lux (± 20 %)

EFFECT PARAMETERS MEASURED:
- Determination of cell concentrations: electronical particle counter (AL CELLCOUNTER Model 871)

TEST CONCENTRATIONS
- Spacing factor for test concentrations: 2

Reference substance (positive control):

yes

Remarks:

Potassium dichromate

Duration:

72 h

Dose descriptor:

EC50

Effect conc.:

> 344.8 mg/L

Nominal / measured:

nominal

Conc. based on:

test mat.

Basis for effect:

biomass

Remarks on result:

other: 95% conf. limits

Duration:

72 h

Dose descriptor:

EC10

Effect conc.:

27.4 mg/L

Nominal / measured:

nominal

Conc. based on:

test mat.

Basis for effect:

biomass

Remarks on result:

other: 95% conf. limits

Duration:

72 h

Dose descriptor:

EC100

Effect conc.:

> 1 000 mg/L

Nominal / measured:

nominal

Conc. based on:

test mat.

Basis for effect:

biomass

Duration:

96 h

Dose descriptor:

EC50

Effect conc.:

335.5 mg/L

Nominal / measured:

nominal

Conc. based on:

test mat.

Basis for effect:

biomass

Remarks on result:

other: 95% conf. limits

Duration:

96 h

Dose descriptor:

EC10

Effect conc.:

19 mg/L

Nominal / measured:

nominal

Conc. based on:

test mat.

Basis for effect:

biomass

Remarks on result:

other: 95% conf. limits

Duration:

96 h

Dose descriptor:

EC100

Effect conc.:

> 1 000 mg/L

Nominal / measured:

nominal

Conc. based on:

test mat.

Basis for effect:

biomass

Duration:

72 h

Dose descriptor:

EC50

Effect conc.:

> 1 000 mg/L

Nominal / measured:

nominal

Conc. based on:

test mat.

Basis for effect:

growth rate

Duration:

72 h

Dose descriptor:

EC0

Effect conc.:

ca. 62.3 mg/L

Nominal / measured:

nominal

Conc. based on:

test mat.

Basis for effect:

growth rate

Duration:

96 h

Dose descriptor:

EC50

Effect conc.:

> 1 000 mg/L

Nominal / measured:

nominal

Conc. based on:

test mat.

Basis for effect:

growth rate

Duration:

96 h

Dose descriptor:

EC0

Effect conc.:

ca. 62.3 mg/L

Nominal / measured:

nominal

Conc. based on:

test mat.

Basis for effect:

growth rate

Details on results:

The concentrations in the analysed test media of the concentrations 62.3 and 1000 mg/l were in the range of 92.8 to 103.9% of the nominal values. In these test media, incubated under test conditions (but withouth algae), the test substance was sufficiently stable. In the analytical samples of the nominal concentration of 250 mg/l a precipitation was observed and must have been removed by filtration prior to analysis. Therefore low recovery values of the test substance were found in this concentration.
The reported results are related to the nominal concentrations of the substance.

Influence of the test substance on algae growth (cells × 10³/ml) during the test duration of 96 hours

 

Concentration	24h rep./mean	48h rep. /mean	72h rep. /mean	96h rep./mean
Control	42.6	260.8	1178.7	3013.0
62.3 mg/l	36.0	198.9	1032.6	2322.4
125.0 mg/l	33.3	155.6	778.1	1598.0
250.0 mg/l	28.7	145.3	660.3	1501.3
500.0 mg/l	26.6	120.1	552.6	1564.7
1000 mg/l	23.7	91.4	322.6	1041.3

 

Rep.: replicate

 

Influence of test substance on algae growth (growth rate μ and percentage inhibition of μ)

 

Nominal test subst. concentration	μ (0 -72h)	Iμ (0 -72h)	μ (0 -96h)	Iμ (0 -96h)
Control	1.590	-	1.428	-
62.3 mg/l	1.547	2.7	1.363	4.6
125.0 mg/l	1.453	8.6	1.270	11.0
250.0 mg/l	1.397	12.2	1.253	12.3
500.0 mg/l	1.337	15.9	1.265	11.4
1000.0 mg/l	1.160	27.0	1.163	18.6

 

Results obtained for the concentrations of the test substance in test medium:

Nominal concentration (mg/l)	Sampling day	Conc. Test substance (mg/l)*	% of nominal
62.3 62.3   250.0 250.0   1000 1000   Control sample Control sample	0 4   0 4   0 4   0 4	64.7 57.8   57.3 56.8   1028.3 950.4   ---** ---**	103.9 92.8   22.9 22.7   102.8 95.0   --- ---

* Mean value of a repeated test

** Counts were below lowest calibration point

Validity criteria fulfilled:

yes

Conclusions:

The substance was tested according to guideline OECD 201.
At the highest concentration tested, algae biomass was inhibited by about 70% growth rate only by about 19% at the end of the test. EC50 (96h) > 1000 mg/l.

Executive summary:

Method

Toxicity to aquatic algae was determined according to OECD guideline 201. A 96 -hour static test was run using green algae Scenedesmus subspicatus Chodat; effect on growth rate was assessed.

Nominal concentrations tested were 62.3, 125, 250, 500 and 1000 mg test substance/l and a control without any additions.

Results

Test sample has a low inhibition effect on the growth of algae. At the highest concentration tested, algae biomass was inhibited by about 70 %, growth rate only by about 19 % at the end of the test. In conclusion, ErC50 (96h) > 1000 mg/l.

Description of key information

OECD 201, 1991, algae, EC50(96h) > 1000 mg/l.

OECD 201, 2003, algae, EC50 (72 -96h) > 100 mg/l

Additional information

No experimental data on target substance was available, thus a read across approach followed as detailed in section 13.

The assessment relied on studies on Similar Sustance 01 and Similar Substance 02, conducted according to OECD guideline 201.

Similar Substance 01 (91 % of active substance) was tested at nominal concentrations of 0 (control), 62.3, 125, 250, 500 and 1000 mg/l. Exposure duration was up to 96 h under static exposure condition. Inhibition of growth rate was ca. 19 % after 96 h of exposure, thus ErC50 > 1000 mg/l.

Similar Substance 02 (10 % of active substance) was tested at nominal concentration of 0 (control), 10, 17, 29, 37, 48, 62, 100 mg/l for 72 hours under static exposure conditions. At the end of exposure, inhibition of growth rate was ca. 29 % at 100 mg/l, thus implying ErC50 (72h) > 100 mg/l.