Benzenesulfonic acid, C10-13-alkyl derivs., sodium salts

Short-term toxicity to aquatic invertebrates

Link to relevant study record(s)

Reference

Reference 1

Endpoint:

short-term toxicity to aquatic invertebrates

Type of information:

experimental study

Adequacy of study:

supporting study

Study period:

From May 20, 1988 to May 28, 1988

Reliability:

2 (reliable with restrictions)

Rationale for reliability incl. deficiencies:

other: Acceptable, well-documented study report which meets basic scientific principles.

Reason / purpose for cross-reference:

reference to same study

Reason / purpose for cross-reference:

reference to other study

Qualifier:

no guideline followed

Principles of method if other than guideline:

Mature Hyalella azeteca were exposed to the test substance at concentrations of 0, 0.63, 1.3, 2.5, 5.0 and 10 mg active ingredient/L for 192 h. The Hyalella were observed for mortality after 48, 96, 144 and 192 h. Results were analysed for the calculation LC50.

GLP compliance:

yes

Remarks:

According to EPA GLP

Analytical monitoring:

no

Vehicle:

no

Details on test solutions:

PREPARATION AND APPLICATION OF TEST SOLUTION: 20 mg/L stock solution was prepared by diluting 0.0212 g of test material in 1000 mL of distilled water. The test solutions were prepared by adding the appropriate volume of stock solution to dilution water to provide the required total solution volume. Each test solution was mixed on a magnetic stirrer for approximately 30 sec and then divided into the appropriate replicate exposure vessels.

Test organisms (species):

other aquatic crustacea: Hyalella azteca

Details on test organisms:

TEST ORGANISM
- Common name: Amphipod
- Strain: Not reported
- Source: Springborn Life Sciences (SLS) culture facility
- Maintenance of stock culture: The culture water was well water with dissolved oxygen (in mg/L), pH and temperature (°C) of 8.4-8.7, 7.4 and 20, respectively. The culture areas received regulated photoperiod of 16 h light and 8 h darkness. The stock cultures were fed a solution of trout chow and a chocolate food supplement once each day. A temperature controlled water bath was used to maintain the culture solution temperatures at 20 ± 2 °C.
- Age at study initiation: Mature animals were used in the study
- Weight, length at study initiation: Not reported. All animals were of same size
- Method of breeding: Not specified
- Feeding during test: Animals were not fed during the 96 h exposure period. However, animals were fed solution of trout chow and a chocolate food supplement when test was extended beyond 96 h

ACCLIMATION: Acclimation period/conditions not reported in the study

Test type:

static

Water media type:

freshwater

Limit test:

no

Total exposure duration:

192 h

Remarks on exposure duration:

As the difference between the 48 and 96 h LC50 value exceeded a factor of two, the exposure was extended after 96 h to 144 h and then again to 192 h when an asymptotic LC50 had been reached

Post exposure observation period:

None

Hardness:

24-30 mg/L CaCO3 (measured by the EDTA titrimetric method)

Test temperature:

21-22 °C (measured with Brooklyn alcohol thermometer)

pH:

7.1, ranged 6.7- 8.5 (measured by La Motte pH meter and combination electrode)

Dissolved oxygen:

Ranged 3.0-8.8 mg/L (measured with a YSI Model #57 dissolved oxygen meter and probe)

Nominal and measured concentrations:

Nominal test concentrations were: 0, 0.63, 1.3, 2.5, 5.0 and 10 mg active ingredient/L

Details on test conditions:

TEST SYSTEM
- Test vessel: Glass beaker
- Type: Closed. (All beakers were covered with clear plastic wrap)
- Material, size, headspace, fill volume: 100 mL glass beaker containing 80 mL of exposure solution. The exposure solution was 4.6 cm deep, with a surface area of 1.6 cm2.
- Aeration: Yes. Aeration was initiated after Day 6 of the study to maintain appropriate dissolved oxygen levels
- No. of organisms per vessel: 1
- No. of vessels per concentration (replicates): 10
- No. of vessels per control (replicates): 10
- Biomass loading rate: 1 animal/80 mL test solution

TEST MEDIUM / WATER PARAMETERS
- Source/preparation of dilution water: Well water obtained from 100 m deep bedrock well
- Total organic carbon: Not reported
- Particulate matter: Not reported
- Metals: Not reported
- Pesticides: Not reported
- Chlorine: Not reported
- Total alkalinity: 22-24 mg/L as CaCO3 (potentiometric titration to an endpoint of pH 4.5)
- Ca/mg ratio: Not reported
- Specific conductivity: 90 - 110 µmhos/cm (measured by yellow springs Instrument Company Model #33 salinity-conductivity-temperature meter and probe)
- Culture medium different from test medium: No
- Intervals of water quality measurement: Observation of physical and biological characteristics of each replicate test solution were recorded at least every 48 h during the exposure period. The pH, dissolved oxygen concentration and temperature were measured at 0, 24, 48, 72, 96, 120, 144, 160 and 192 h in the vessel of the two highest treatment levels and the control.

OTHER TEST CONDITIONS
- Adjustment of pH: Not reported
- Photoperiod: 16 h of light and 8 h of darkness
- Light intensity: 200 footcandles (Sylvania GRO-LUX® and Cool White® fluorescent lights). Light intensity was measured with General Electric type 214 light meter

EFFECT PARAMETERS MEASURED: Hyalella were observed for mortality after 48, 96, 144 and 192 h.

NEGATIVE CONTROL PERFORMED: Yes (received only dilution water)

TEST CONCENTRATIONS
- Spacing factor for test concentrations: 2
- Range finding study: Prior to the initiation of each definitive exposure, preliminary range-finding test were conducted and data obtained from preliminary exposure was used to select the definitive test concentration
- Test concentrations: Not reported
- Results used to determine the conditions for the definitive study: Yes

Reference substance (positive control):

no

Duration:

48 h

Dose descriptor:

LC50

Effect conc.:

7.6 mg/L

Nominal / measured:

nominal

Conc. based on:

act. ingr.

Basis for effect:

mortality

Remarks on result:

other: 4.9-21

Duration:

96 h

Dose descriptor:

LC50

Effect conc.:

3.5 mg/L

Nominal / measured:

nominal

Conc. based on:

act. ingr.

Basis for effect:

mortality

Remarks on result:

other: 2.4 - 5.3

Duration:

144 h

Dose descriptor:

LC50

Effect conc.:

1.1 mg/L

Nominal / measured:

nominal

Conc. based on:

act. ingr.

Basis for effect:

mortality

Remarks on result:

other: 0.23 - 1.9

Duration:

192 h

Dose descriptor:

LC50

Effect conc.:

0.96 mg/L

Nominal / measured:

nominal

Conc. based on:

act. ingr.

Basis for effect:

mortality

Remarks on result:

other: 0.29 - 1.6

Details on results:

- Behavioural abnormalities: Several of the surviving amphipods exhibited lethargic behavior. See 'Table 1' in the section 'Any other information on results'.
- Observations on body length and weight: Not reported
- Other biological observations: None
- Mortality: See Table 1 below.
- Other adverse effects control: Not reported
- Abnormal responses: Not reported
- Any observations (e.g. precipitation) that might cause a difference between measured and nominal values: No visible signs of undissolved test material (e.g. precipitate, film on the solution¿s surface) were observed in any exposure solution. Water quality parameters measured during each test were generally unaffected by the concentration of test material tested and remained within acceptable ranges for the survival of the exposed organisms.
- Effect concentrations exceeding solubility of substance in test medium: No

Reported statistics and error estimates:

If at least one test concentration caused mortality of greater than or equal to 50% of the test population, then a computer program modified from the program of C. Stephan was used to calculate the LC50 values and 95% confidence intervals. Three statistical methods were available in the computer program: moving average angle analysis, probit analysis, and nonlinear interpolation with 95% confidence intervals calculated by binomial probability. See study report for more details on statistical methods.

All LC50s for Hyalella were estimated by probit analysis (48 hr, 96 hr, 144 hr, 196 hr).

Table 1: Cumulative percent mortalities and observations made during the 96 h static exposure of amphipods to linear alkylbenzene sulfonate (Hyalella azetec) (study # 34845)

Nominal concentration (mg active ingredient/L)	Percent Mortalities (%)
	48 h (mean)	96 h (mean)	144 h (mean)		192 h (mean)
Control	0	0	20^ef		30
0.63	0	10^b	60^be		60
1.3	0	0^d	30^ef		30
2.5	20	30^b	60^de		70
5	30^b	60^b	90^de		100
10	60^c	100	100		100

^aValue represents the mean percent of all replicate vessesls; (N=10)

^bTwo surviving amphipods exhibited lethargic behavior

^cAll surviving amphipods exhibited lethargic behavior

^dOne surviving amphipod exhibited lethargic behavior

^eUneaten food was observed in the bottom of the beakers and caught on the amphipods

^fSeveral of the surviving amphipods exhibited lethargic behavior

Results were based on 92.9% active ingredient Test material was C12.3 LAS.

Validity criteria fulfilled:

not applicable

Conclusions:

The 96 hr LC50 of C12.3 LAS (linear alkylbenzene sulfonate) to amphipods (Hyalella azetec) was 3.5 mg active ingredient/L, based on nominal concentration of test substance.

Executive summary:

The 96 hr LC50 of linear alkylbenzene sulfonate (C12.3 LAS) to Hyalella azetec was determined under static conditions. The nominal test concentrations were 0, 0.63, 1.3, 2.5, 5.0 and 10 mg active ingredient/L. The mortality of amphipods was determined at 48, 96, 144 and 192 h.

The 96 hr LC50 of linear alkylbenzene sulfonate to amphipods (Hyalella azetec) was 3.5 mg/L (2.4 -5.3). LC50s were estimated for the following time points: 48 hr       7.6 mg/L       (4.9 -21.0) 96 hr       3.5 mg/L       (2.4 -5.3) 144 hr       1.1 mg/L       (0.23 -1.9) 196 hr       0.96 mg/L       (0.29 -1.6)

Reference 2

Endpoint:

short-term toxicity to aquatic invertebrates

Type of information:

experimental study

Adequacy of study:

supporting study

Study period:

From Apr. 21, 1988 to Apr. 25, 1988

Reliability:

2 (reliable with restrictions)

Rationale for reliability incl. deficiencies:

other: Acceptable, well-documented study report which meets basic scientific principles.

Reason / purpose for cross-reference:

reference to same study

Reason / purpose for cross-reference:

reference to other study

Qualifier:

no guideline followed

Principles of method if other than guideline:

Immature larvae of midge (Chironomus riparius) were exposed to the test substance at test concentrations of 0, 1.3, 2.5, 5, 10 and 20 mg active ingredient/L for 96 h. The midge were observed for mortality after 48 h and 96 h of test initiation. Results were analysed for the calculation of 48 h and 96 h LC50.

GLP compliance:

yes

Remarks:

According to EPA GLP

Analytical monitoring:

no

Vehicle:

no

Details on test solutions:

PREPARATION AND APPLICATION OF TEST SOLUTION: 100 mg/L stock solution was prepared by diluting 0.1087 g of the test material in 1000 mL of distilled water. The test solutions were prepared by adding the appropriate volume of stock solution to the dilution water to provide the required total solution volume. Each test solution was mixed on a magnetic stirrer for approximately 30 seconds and then divided into the appropriate replicate exposure vessels.

Test organisms (species):

other aquatic arthropod: Chironomus riparius

Details on test organisms:

TEST ORGANISM
- Common name: Midge
- Strain: Not reported
- Source: Springborn Life Sciences (SLS) culture facility
- Maintenance of stock culture: The culture water was well water with dissolved oxygen (mg/L), pH and temperature (°C) of 7.7, 7.3 and 19 respectively. The culture areas received regulated photoperiod of 16 h light and 8 h darkness. The stock cultures were fed a solution of trout chow and a chocolate food supplement once each day. A temperature controlled water bath was used to maintain the culture solution temperatures at 20 ± 2 °C.
- Age at study initiation: Immature larvae of same size were used in the study
- Weight, length at study initiation: Not reported. Animals of same size are used in the study.
- Method of breeding: Not specified
- Feeding during test: Animals were not fed during the exposure period

ACCLIMATION: Acclimation period/conditoins not reported in the study

Test type:

static

Water media type:

freshwater

Limit test:

no

Total exposure duration:

96 h

Post exposure observation period:

None

Hardness:

24-30 mg/L CaCO3 (measured by the EDTA titrimetric method)

Test temperature:

21-22 °C (measured with Brooklyn alcohol thermometer)

pH:

7.1, ranged 6.9-7.7 (measured by La Motte pH meter and combination electrode)

Dissolved oxygen:

6.9-8.8 mg/L (measured with a YSI Model #57 dissolved oxygen meter and probe)

Nominal and measured concentrations:

Nominal test concentrations were: 0, 1.3, 2.5, 5, 10 and 20 mg active ingredient/L

Details on test conditions:

TEST SYSTEM
- Test vessel: Glass beaker
- Type: Open
- Material, size, headspace, fill volume: 250 mL glass beaker containing 150 mL of exposure solution. The test solutions were 4.7 cm deep, with a surface area of 33 cm2.
- Aeration: No
- No. of organisms per vessel: 10
- No. of vessels per concentration (replicates): Two
- No. of vessels per control (replicates): One
- Biomass loading rate: There were 10 animals/vessel. Loading rate was therefore 1 animal/15 mL test solution

TEST MEDIUM / WATER PARAMETERS
- Source/preparation of dilution water: Well water obtained from 100 m deep bedrock well
- Total organic carbon: Not reported
- Particulate matter: Not reported
- Metals: Not reported
- Pesticides: Not reported
- Chlorine: Not reported
- Total alkalinity: 22-24 mg/L as CaCO3 (potentiometric titration to an endpoint of pH 4.5)
- Ca/mg ratio: Not reported
- Specific conductivity: 90 - 110 µmhos/cm (measured by yellow springs Instrument Company Model #33 salinity-conductivity-temperature meter and probe)
- Culture medium different from test medium: No
- Intervals of water quality measurement: Observation of physical and biological characteristics of each replicate test solution were recorded at least every 48 h during the exposure period. The pH, dissolved oxygen concentration and temperature were measured at 0, 24, 48, 72 and 96 h in the vessel of the highest treatment level and the control.

OTHER TEST CONDITIONS
- Adjustment of pH: Not reported
- Photoperiod: 16 h of light and 8 h of darkness
- Light intensity: 200 footcandles (Sylvania GRO-LUX® and Cool White® fluorescent lights). Light intensity was measured with General Electric type 214 light meter

EFFECT PARAMETERS MEASURED: Test organisms were observed for mortality after 48 and 96 h.

NEGATIVE CONTROL PERFORMED: Yes (received only dilution water)

TEST CONCENTRATIONS
- Spacing factor for test concentrations: 2
- Range finding study: Prior to the initiation of each definitive exposure, preliminary range-finding test were conducted and data obtained from preliminary exposure was used to select the definitive test concentration
- Test concentrations: Not reported
- Results used to determine the conditions for the definitive study: Yes

Reference substance (positive control):

no

Duration:

48 h

Dose descriptor:

LC50

Effect conc.:

8.6 mg/L

Nominal / measured:

nominal

Conc. based on:

act. ingr.

Basis for effect:

mortality

Remarks on result:

other: 5.0-20.0

Duration:

96 h

Dose descriptor:

LC50

Effect conc.:

6.5 mg/L

Nominal / measured:

nominal

Conc. based on:

act. ingr.

Basis for effect:

mortality

Remarks on result:

other: 5.3 - 8.2

Details on results:

- Behavioural abnormalities: Several of the surviving midge exhibited lethargic behavior. See 'Table 1' in the section 'Any other information on results'.
- Observations on body length and weight: Not reported
- Other biological observations: None
- Mortality: Following 96 h of exposure, mortality of 100% and 95% was observed in the two highest treatment levels. Mortality of ¿5% was observed in the remaining lower treatment levels (i.e. 5, 2.5 and 1.3 mg active/L). Control mortality was 10%.
- Other adverse effects control: Other adverse effects on control are not reported in the study
- Abnormal responses: Not reported
- Any observations (e.g. precipitation) that might cause a difference between measured and nominal values: No visible signs of undissolved test material (e.g. precipitate, film on the solution¿s surface) were observed in any exposure solution. Water quality parameters measured during each test were generally unaffected by the concentration of test material tested and remained within acceptable ranges for the survival of the exposed organisms.
- Effect concentrations exceeding solubility of substance in test medium: No

Reported statistics and error estimates:

If at least one test concentration caused mortality of greater than or equal to 50% of the test population, then a computer program modified from the program of C. Stephan was used to calculate the LC50 values and 95% confidence intervals. Three statistical methods were available in the computer program: moving average angle analysis, probit analysis, and nonlinear interpolation with 95% confidence intervals calculated by binomial probability.

The 96 hr LC50 for midge was estimated by moving average angle analysis. The 48 hr LC50 for midge was estimated by non-linear interpolation (95% confidence interval calculated by binominal probability).

Table 1: Cumulative percent mortalities and observations made during the 96 h static exposure of midge (Chironomus riparius) to linear alkylbenzene sulfonate (study # 34845)

Nominal concentration (mg active ingredient/L)	Percent Mortalities (%)
	48 h			96 h
	A	B	Mean	A	B	Mean
Control	0	10	5	0	20	10
1.3	0	0	0	0	0	0
2.5	0	0	0	0	10	5
5	0	0	0*	0	10	5
10	80	60	70*	100	90	95**
20	100	100	100	100	100	100

* Several of the surviving midge exhibited lethargic behavior

**Surviving midge displayed lethargic behavior

Results were based on 92.9% active ingredient (C12.3LAS).

Validity criteria fulfilled:

not applicable

Conclusions:

The 96 hr LC50 of C12.3 LAS (linear alkylbenzene sulfonate) to midge (Chironomus riparius) was 6.5 mg active ingredient/L, based on nominal concentration of test substance.

Executive summary:

The 96 hr LC50 of linear alkylbenzene sulfonate (C12.3 LAS) to midge (Chironomus riparius) was determined under static conditions.  The nominal test concentrations were 0, 1.3, 2.5, 5, 10 and 20 mg active ingredient/L. The mortality of midge was determined at 48 h and 96 h.

Mortality of ¿5% was observed at the three lowest test concentrations (1.3, 2.5, 5.0 mg/L). Mortality of 95-100% was observed in the two highest exposure levels (10, 20 mg/L) after 96 h. Control mortality was 10%.

The 48 h and 96 h LC50 values were 8.6 and 6.5 mg active ingredient/L, respectively.

Reference 3

Endpoint:

short-term toxicity to aquatic invertebrates

Type of information:

experimental study

Adequacy of study:

supporting study

Study period:

From Apr. 21, 1988 to Apr. 25, 1988

Reliability:

2 (reliable with restrictions)

Rationale for reliability incl. deficiencies:

other: Acceptable, well-documented study report which meets basic scientific principles.

Reason / purpose for cross-reference:

reference to same study

Reason / purpose for cross-reference:

reference to other study

Qualifier:

no guideline followed

Principles of method if other than guideline:

Mature oligochaetes (Limnodrilus hoffmeisteri) were exposed to test substance concentrations of 0, 0.31, 0.63, 1.3, 2.5 and 5.0 mg active ingredient/L for 96 h. Oligochaetes were observed for mortality after 48 h and 96 h. Results were analysed for the calculation of 48 h and 96 h LC50.

GLP compliance:

yes

Remarks:

According to EPA GLP

Analytical monitoring:

no

Vehicle:

no

Details on test solutions:

PREPARATION AND APPLICATION OF TEST SOLUTION: 100 mg/L stock solution was prepared by diluting 0.1087 g of test material in 1000 mL of distilled water. The test solutions were prepared by adding the appropriate volume of stock solution to dilution water to provide the required total solution volume. Each test solution was mixed on a magnetic stirrer for approximately 30 seconds and then divided into the appropriate replicate exposure vessels

Test organisms (species):

other aquatic worm: Limnodrilus hoffmeisteri

Details on test organisms:

TEST ORGANISM
- Common name: Oligochaetes
- Strain: Not reported
- Source: Springborn Life Sciences (SLS) culture facility
- Maintenance of stock culture: The culture water was well water with dissolved oxygen (mg/L), pH and temperature (°C) of 7.6-8.3, 7.3 and 18-19 respectively. The culture areas received regulated photoperiod of 16 h light and 8 h darkness. The stock cultures were fed a solution of trout chow and a chocolate food supplement and tetra-Min (a flaked fish food) once each day. A temperature controlled water bath was used to maintain the culture solution temperatures at 20 ± 2 °C.
- Age at study initiation: Mature animals were used in the study.
- Weight, length at study initiation: Not reported. However, the animals were of same size.
- Method of breeding: Not specified
- Feeding during test: Animals were not fed during the exposure period

ACCLIMATION: Acclimation period/conditions not reported in the study

Test type:

static

Water media type:

freshwater

Limit test:

no

Total exposure duration:

96 h

Post exposure observation period:

None

Hardness:

24-30 mg/L CaCO3 (measured by the EDTA titrimetric method)

Test temperature:

21 °C (measured with Brooklyn alcohol thermometer)

pH:

7.1, ranged 6.6-7.3 (measured by La Motte pH meter and combination electrode)

Dissolved oxygen:

Ranged 1.0-8.8 mg/L (measured with a YSI Model #57 dissolved oxygen meter and probe)

Nominal and measured concentrations:

Nominal test concentrations were: 0, 0.31, 0.63, 1.3, 2.5 and 5.0 mg active ingredient/L

Details on test conditions:

TEST SYSTEM
- Test vessel: Glass beaker
- Type: Open
- Material, size, headspace, fill volume: 250 mL glass beaker containing 150 mL of exposure solution. The exposure solution depth was 4.7 cm with a surface area of 33 cm2.
- Aeration: No
- No. of organisms per vessel: 10
- No. of vessels per concentration (replicates): Two
- No. of vessels per control (replicates): One
- Biomass loading/ rate: There were 10 animals/vessel. Loading rate was therefore 1 animal/15 mL test solution.

TEST MEDIUM / WATER PARAMETERS
- Source/preparation of dilution water: Well water obtained from 100 m deep bedrock well
- Total organic carbon: Not reported
- Particulate matter: Not reported
- Metals: Not reported
- Pesticides: Not reported
- Chlorine: Not reported
- Total alkalinity: 22-24 mg/L as CaCO3 (potentiometric titration to an endpoint of pH 4.5)
- Ca/mg ratio: Not reported
- Specific conductivity: 90 - 110 µmhos/cm (measured by yellow springs Instrument Company Model #33 salinity-conductivity-temperature meter and probe)
- Culture medium different from test medium: No
- Intervals of water quality measurement: Observation of physical and biological characteristics of each replicate test solution were recorded at least every 48 h during the exposure period. The pH, dissolved oxygen concentration and temperature were measured at 0, 24, 48, 72 and 96 h in the vessel of the highest treatment level and the control.

OTHER TEST CONDITIONS
- Adjustment of pH: Not reported
- Photoperiod: 6 h of light and 8 h of darkness
- Light intensity: 200 footcandles (Sylvania GRO-LUX® and Cool White® fluorescent lights). Light intensity was measured with General Electric type 214 light meter

EFFECT PARAMETERS MEASURED: Oligochaetes were observed for mortality after 48 and 96 h of test initiation.

NEGATIVE CONTROL PERFORMED: Yes (received only dilution water)

TEST CONCENTRATIONS
- Spacing factor for test concentrations: 2
- Range finding study: Prior to the initiation of each definitive exposure, preliminary range-finding test were conducted and data obtained from preliminary exposure was used to select the definitive test concentration
- Test concentrations: Not reported
- Results used to determine the conditions for the definitive study: Yes

Reference substance (positive control):

no

Duration:

48 h

Dose descriptor:

LC50

Effect conc.:

2.4 mg/L

Nominal / measured:

nominal

Conc. based on:

act. ingr.

Basis for effect:

mortality

Remarks on result:

other: 1.9-3.1

Duration:

96 h

Dose descriptor:

LC50

Effect conc.:

1.8 mg/L

Nominal / measured:

nominal

Conc. based on:

act. ingr.

Basis for effect:

mortality

Remarks on result:

other: 1.4-2.2

Details on results:

- Behavioural abnormalities: Several of the surviving oligochaetes exhibited lethargic behavior. See 'Table 1' in the section 'Any other information on results'.
- Observations on body length and weight: Not reported
- Other biological observations: Clumpping of oligochaetes together in a ball was observed. See 'Table 1' in the section 'Any other information on results'.
- Mortality: see Table 1 below
- Other adverse effects control: not reported
- Abnormal responses: Not reported
- Any observations (e.g. precipitation) that might cause a difference between measured and nominal values: No visible signs of undissolved test material (e.g. precipitate, film on the solution¿s surface) were observed in any exposure solution. Water quality parameters measured during each test were generally unaffected by the concentration of test material tested and remained within acceptable ranges for the survival of the exposed organisms.
- Effect concentrations exceeding solubility of substance in test medium: No

Reported statistics and error estimates:

If at least one test concentration caused mortality of greater than or equal to 50% of the test population, then a computer program modified from the program of C. Stephan was used to calculate the LC50 values and 95% confidence intervals. Three statistical methods were available in the computer program: moving average angle analysis, probit analysis, and nonlinear interpolation with 95% confidence intervals calculated by binomial probability. See study report for more details on statistical methods.

The 96 hr LC50 for the Oligochaetes was estimated by moving average angle analysis. The 48 hr LC50 for the Oligochaetes was estimated by probit analysis.

Table 1: Cumulative percent mortalities and observations made during the 96 h static exposure of oligocheates (Limnodrilus hoffmeisteri) to linear alkylbenzene sulfonate (study # 34845)

Nominal concentration (mg active ingredient/L)	Percent Mortalities (%)
	48 h			96 h
	A	B	Mean	A	B	Mean
Control	0	0	0^a	0	0	0
0.31	0	0	0^a	0	0	0
0.63	0	0	0^a	0	0	0e
1.3	20	20	20^b	50	60	55^b
2.5	40	70	55^c	40	80	60^b
5.0	90	80	85^d	90	90	90^b

^aOligochaetes were clumped together in a ball

^bAll surviving oligochaetes exhibited lethargic behavior

^cSeveral surviving oligochaetes exhibited lethargic behavior

^dOne surviving oligochaete exhibited lethargic behavior

^eOne oligochaete body was split from its head and the head was still alive

Results were based on 92.9%active ingredient The test substance was C12.3 LAS.

Validity criteria fulfilled:

not applicable

Conclusions:

The 96 hr LC50 of C12.3 LAS (linear alkylbenzene sulfonate) to oligochaetes (Limnodrilus hoffmeisteri) was 1.8 mg active ingredient/L, based on nominal concentration of test substance.

Executive summary:

The 96 hr LC50 of linear alkylbenzene sulfonate (C12.3 LAS) to oligochaetes (Limnodrilus hoffmeisteri) was determined under static conditions. The nominal test concentrations were 0, 0.31, 0.63, 1.3, 2.5 and 5.0 mg active ingredient/L. The mortality of oligochaetes was determined at 48h and 96 h.

The 48 and 96 hr LC50 values were 2.4 and 1.8 mg active ingredient/L, respectively.

Reference 4

Endpoint:

short-term toxicity to aquatic invertebrates

Type of information:

experimental study

Adequacy of study:

key study

Study period:

1989-10-31 to 1989-11-2

Reliability:

2 (reliable with restrictions)

Rationale for reliability incl. deficiencies:

other: Followed guideline with minor deviation, well documented

Reason / purpose for cross-reference:

reference to same study

Qualifier:

according to guideline

Guideline:

OECD Guideline 202 (Daphnia sp. Acute Immobilisation Test)

Deviations:

yes

Remarks:

Measurement of test concentration is not available.

Principles of method if other than guideline:

Not applicable

GLP compliance:

yes (incl. QA statement)

Analytical monitoring:

yes

Details on sampling:

- Concentrations: Samples for analysis of each concentration tested were taken at 0, 24 and 48 h
- Sampling method: The samples were taken in glass bottles and preserved by adding 1ml of 37% formalin in 99 ml of test sample solution
- Sample storage conditions before analysis: Not available

Vehicle:

no

Details on test solutions:

PREPARATION AND APPLICATION OF TEST SOLUTION (especially for difficult test substances)
- Method: 200 mg of Linear alkyl benzene sulfonate (LAS ex Petresa) (E-3474.01) (ETS311) was accurately weighed out and dissolved in 2000 ml of dilution water, resulting in a solution containing 100mg/L. From this solution 32, 56, 100, 180, 320 and 560 ml were taken and diluted with dilution water to one liter. These solutions, containing 3.2, 5.6,10, 18, 32 and 56 mg/L and the solution containing 100 mg/L were used as test solutions; portions of 100ml of each test solution were placed in each of four 150 ml beakers.
- Eluate: Not applicable
- Differential loading: Not applicable
- Controls: Control vessel contained 100% dilution water.
- Chemical name of vehicle (organic solvent, emulsifier or dispersant): Not applicable
- Concentration of vehicle in test medium (stock solution and final test solution): Not applicable
- Evidence of undissolved material (e.g. precipitate, surface film, etc): None

Test organisms (species):

Daphnia magna

Details on test organisms:

TEST ORGANISM
- Common name: Daphnia magna
- Strain: Daphnia magna Straus 1820
- Source: TNO Nutrition and Food Research Institute Laboratory culture
- Age at study initiation (mean and range, SD): <24 h old
- Weight at study initiation (mean and range, SD): Not available
- Length at study initiation (length definition, mean, range and SD): Not available
- Valve height at study initiation, for shell deposition study (mean and range, SD): Not applicable
- Peripheral shell growth removed prior to test initiation: Not applicable
- Method of breeding: Organisms were cultured in laboratory kept under conditions specified in NPR 6503
- Feeding during test: Not fed
- Food type: Not applicable
- Amount: Not applicable
- Frequency: Not applicable


ACCLIMATION
- Acclimation period: Not available
- Acclimation conditions (same as test or not): Not available
- Type and amount of food: Not available
- Feeding frequency: Not available
- Health during acclimation (any mortality observed): Not available


QUARANTINE (wild caught)
- Duration: Not applicable
- Health/mortality: Not applicable

Test type:

static

Water media type:

freshwater

Limit test:

no

Total exposure duration:

48 h

Post exposure observation period:

Not applicable

Hardness:

The hardness of the test solution was 215 mg/L as CaCO3

Test temperature:

Temperature was maintained at 20 ± 1 °C

pH:

pH was measured at 0 and 48 h. Overall pH ranged 7.9-8.1. See the table 1 given below
Control, 3.2, 5.6, 10, 18, 32, 56 mg/L pH ranged as 8-8.1
100 mg/L: range 7.9-8

Dissolved oxygen:

DO was measured at 0 and 48 h. Overall DO ranged 8.8-9.6 mg/L. See the Table 2 given below.
Control: range 8.8-9.5 mg/L
3.2 mg/L: range 8.8-9.3 mg/L
5.6 mg/L: range 8.7-9.1 mg/L
10 mg/L: range 8.8-9.4 mg/L
18 mg/L: range 8.8-9.3 mg/L
32 mg/L and 56 mg/L: range 8.8-9.2 mg/L
100 mg/L: range 8.8-9.6 mg/L

Salinity:

Not applicable

Nominal and measured concentrations:

Nominal test concentrations based on the test material as received were 0, 3.2, 5.6, 10, 18, 32, 56 and 100 mg/L. The test material contained 12.09% active. Samples were taken for analytical measurement of test concentrations, but those results are not available in this study report.

Details on test conditions:

TEST SYSTEM
- Test vessel:
- Type (delete if not applicable): open / closed: Not available
- Material, size, headspace, fill volume: 150 ml all-glass beakers, containing 100 ml test solution.
- Aeration: Not aerated
- Type of flow-through (e.g. peristaltic or proportional diluter): Not applicable
- Renewal rate of test solution (frequency/flow rate): Not renewed
- No. of organisms per vessel: 5
- No. of vessels per concentration (replicates): 4
- No. of vessels per control (replicates): 4
- No. of vessels per vehicle control (replicates): Not applicable
- Biomass loading rate: 100 ml of test solution/5 organisms=20 ml test solution per organism


TEST MEDIUM / WATER PARAMETERS
- Source/preparation of dilution water: The dilution water was prepared by the addition of several salts to groundwater from a locality near Linschoten (The Netherlands). The composition of the dilution water (DSWL) was Na+: 1.20 mmol/L, K+: 0.23 mmol/L, Cl-: 2.59 mmol/L, SO4:2-: 0.67 mmol/L, HCO3-: 1.39mmol/L and contained several other trace elements (<<1mg/L). Water had hardness of 215 mg/L as CaCO3, pH of 8-8.2. This DSWL had proven to be suitable for the culture of the test organism used in this test.
- Total organic carbon: Not available
- Particulate matter: Not available
- Metals: Not available
- Pesticides: Not available
- Chlorine: Not available
- Alkalinity: Not available
- Ca/mg ratio: 1.37 mmol/L Ca 2+/ 0.78mmol/L Mg+2
- Conductivity: Not available
- Culture medium different from test medium: Not specified in the study report.
- Intervals of water quality measurement: pH and DO were measured at the beginning and at the end of the test. Temperature was maintained constant throughout the test.


OTHER TEST CONDITIONS
- Adjustment of pH: Not applicable
- Photoperiod: 16 h light and 8 h dark
- Light intensity: Not available


EFFECT PARAMETERS MEASURED (with observation intervals if applicable) : The immobile animals were counted after 24 h and 48 h. At the same time, the condition (Swimming behaviour, color or any other observable morphological or behavioural criterium) of the mobile animals was visually compared with that of the control animals (blanks)


TEST CONCENTRATIONS
- Spacing factor for test concentrations: 1.8
- Justification for using less concentrations than requested by guideline: Not applicable
- Range finding study: No
- Test concentrations: N/A
- Results used to determine the conditions for the definitive study: N/A

Reference substance (positive control):

no

Duration:

48 h

Dose descriptor:

EC50

Effect conc.:

2.9 mg/L

Nominal / measured:

nominal

Conc. based on:

act. ingr.

Basis for effect:

mobility

Remarks on result:

other: 95% CL: 2.5-3.5 mg/L

Duration:

24 h

Dose descriptor:

EC50

Effect conc.:

6.4 mg/L

Nominal / measured:

nominal

Conc. based on:

act. ingr.

Basis for effect:

mobility

Remarks on result:

other: 95% CL: 5.4-7.5 mg/L

Details on results:

- Behavioural abnormalities: At some test concentrations, the daphnids moved irregularly and more slowly than controls (see Table 3 below for details).
- Observations on body length and weight: None
- Other biological observations: At some test concentrations, some daphnids were paler than the controls (see Table 3 below for details).
- Mortality of control: None
- Other adverse effects control: None
- Abnormal responses: None
- Any observations (e.g. precipitation) that might cause a difference between measured and nominal values: None
- Effect concentrations exceeding solubility of substance in test medium: None

Results with reference substance (positive control):

Not applicable

Reported statistics and error estimates:

The EC50 values and their confidence intervals were calculated by means of a parametric model developed by a Kooijiman.

Table 3: Number of mobile animals and their condition at several concentrations of E-3474.01 (ETS 311) and after several exposure times(a, b, cand d are quadruplicate test vessels).

Time (h)	Concentration of test substance (mg.l"^1)
	0				3.2				5.6				10
	a	b	c	d	a	b	c	d	a	b	c	d	a	b	c	d
0	5	5	5	5	5	5	5	5	5	5	5	5	5	5	5	5
24	5	5	5	5	5	5	5	5	5	5	5	5	5	5	5	5
48	5^1)	5^1)	5^1)	5^1)	5^2)	5^2)	5^2)	5^2)	5^2)	5^2)	5^2)	5^3)4)	4^3)4)	5^3)4)	5^3)4)	5^3)4)
Time (h)	Concentration of test substance (mg.l'^1)
	18				32				56				100
	a	b	c	d	a	b	c	d	a	b	c	d	a	b	c	d
0	5	5	5	5	5	5	5	5	5	5	5	5	5	5	5	5
24	5	5	5	5	5	5	5	5	1	3	3	3	0	0	0	0
48	4^3)4)	4^3)4)	3^3)4)	3^3)4)	1^3)4)5)	2^3)4)5)	3^3)4)5)	2^3)4)5)	0	0	0	0	0	0	0	0

1)     Condition of all test animals, visually assessed, normal (= good),

2)     Condition of all test animals, visually assessed, equal to that of the control animals.

3)     The quoted number animals were mobile, according to the definition given in OECD Guideline no. 202, but their condition (see section 2.4) was not equal to that of the control animals; they moved irregularly and more slowly than the controls.

4)   The quoted number of mobile animals was paler than the control animals.

5) The animals were almost immobile

Validity criteria fulfilled:

yes

Remarks:

All criteria of OECD 202 were met

Conclusions:

The 48-hour EC50 of Linear alkyl benzene sulfonate (LAS ex Petresa) to Daphnia magna was 2.9 mg/L based on nominal concentration of the active ingredient.

Executive summary:

In a 48 h acute toxicity study, Daphnia magna were exposed to Linear alkyl benzene sulfonate (LAS ex Petresa) at nominal concentrations of 0 (Control), 3.2, 5.6, 10, 18, 32, 56 and 100 mg/L under static condition, based on the sample as received. The sample was 12.1% active ingredient, and results were adjusted to the active ingredient basis.

The 48 h EC50 value based on mobility was 2.9 mg a.i./L with 95% CL of 2.5 to 3.5 mg/L. This toxicity study is classified as acceptable and satisfies the guideline requirement for the current OECD 202 Daphnia acute toxicity study

Reference 5

Endpoint:

short-term toxicity to aquatic invertebrates

Type of information:

experimental study

Adequacy of study:

supporting study

Reliability:

2 (reliable with restrictions)

Rationale for reliability incl. deficiencies:

other: Well documented peer-reviewed publication.

Qualifier:

according to guideline

Guideline:

other: EPA-660/3-75-009 1975. Method for acute toxicity tests with fish, macroinvertebrates and amphibians.with fish, macroinvertebrates and amphibians.

Principles of method if other than guideline:

Acute toxicity tests were conducted on high molecular weight LAS, individual pure homologues, non-linear LAS components (dialkyl tetralin or indane sulfonates (DTIS), and model biodegradation intermediates (sulfophenyl undecane, SØU) in order to determine whether biodegradation decreases toxicity. In 250 mL beakers with 200 mL of well water of approximately 250 mg/L hardness, ten Daphnia, less than 18 hours old, were placed in each of the three beakers. No food was added for the duration of the test.

GLP compliance:

no

Analytical monitoring:

yes

Vehicle:

no

Details on test solutions:

In 250 mL beakers with 200 mL of well water of approximately 250 mg/L hardness, ten Daphnia, less than 18 hours old, were placed in each of the three replicate beakers per concentration. Five concentrations were tested.

Test organisms (species):

Daphnia magna

Details on test organisms:

Crustacea

Test type:

static

Water media type:

freshwater

Limit test:

no

Total exposure duration:

48 h

Post exposure observation period:

open system

Hardness:

250 mg/L

Test temperature:

not reported

pH:

not reported

Dissolved oxygen:

not reported

Reference substance (positive control):

not required

		Daphnia magna
	Average chain length	24 hour	48 hour
High molecular weight LAS	13.3	2.6 ± 0.1	2.3 ± 0.1
Individual homologues LAS
C10	10	53.1 ± 0.4	12.3 ± 2.6
C11	11	15.8 ± 3.0	5.7 ± 0.6
C12	12	10.7 ± 1.6	3.5 ± 1.0
C13	13	2.7 ± 0.4	2.0 ± 0.3
C14	14	1.2 ± 0.2	0.7 ± 0.2
Nonlinear LAS components (DTIS)
C10	10	106.0 ± 27.0	98.0 ± 21.3
C12	12	55.1 ± 9.1	34.1 ± 5.1
C14	14	12.4 ± 1.4	10.0 ± 1.0
Model biodegradation intermediates
C4 (SØ Butyrate)	4	~12,000	~6,000
C5 (SØ Valerate)	5	~12,000	~5,000
C11 (SØU)	11	355 ± 150*	208 ± 85*

*Subsequent repurification of this sample yielded a product with the same isomeric composition but with LC₅₀ values over 1000 mg/L for daphnids (Swisher et al., 1976).

Validity criteria fulfilled:

yes

Conclusions:

Partial biodegradation of LAS is shown to significantly reduce the specific toxicity of the remaining LAS to Daphnia magna.

Executive summary:

The toxicity of high molecular weight LAS, individual LAS homologues, and nonlinear LAS components (DTIS) was measured in a series of aquatic toxicity tests with Daphnia magna. Results show that biodegradation of LAS influences the toxicity, with the remaining LAS becoming less toxic. The 48 -hour LC50 of high molecular weight LAS to Daphnia magna is 2.4 mg/L.

Description of key information

Five studies characterize the potential for acute toxicity to aquatic invertebrates. In the key study (The Procter & Gamble Company 1990), Daphnia magna were exposed for 48 hours to LAS at nominal concentrations of 0 (Control), 3.2, 5.6, 10, 18, 32, 56 and 100 mg/L under static conditions. The sample was 12.1% active ingredient, and results were adjusted to the active ingredient basis. The resultant 48 h EC₅₀value based on mobility was 2.9 mg a.i./L. In the second study (Kimerle and Swisher 1977), the toxicity of high molecular weight LAS, individual LAS homologues, and nonlinear LAS components (dialkyl tetralin or indane sulfonates, DTIS) was measured in a series of aquatic toxicity tests with Daphnia magna. Results show that biodegradation of LAS influences the toxicity, with the remaining LAS becoming less toxic. The 48 hour LC₅₀of high molecular weight LAS to Daphnia magna is 2.4 mg/L. In the third study, the 96 hr LC50 of linear alkylbenzene sulfonate (C12.3 LAS) to amphipods (Hyalella azetec) was determined under static conditions. The nominal test concentrations were adjusted to active ingredient/L. The mortality of amphipods was determined at 48, 96, 144 and 192 h, and an LC50 calculated. The 96 hr LC50 of C12.3 LAS (linear alkylbenzene sulfonate) to Hyalella azetec was 3.5 mg active ingredient/L, based on nominal concentration of test substance. In the fourth study, the 96 hr LC50 of linear alkylbenzene sulfonate (C12.3 LAS) to immature larvae of midge (Chironomus riparius) was determined under static conditions.  The nominal test concentrations were 0, 1.3, 2.5, 5, 10 and 20 mg active ingredient/L. The 48 h and 96 h LC50 values were 8.6 and 6.5 mg active ingredient/L, respectively. In the fifth study, mature oligochaetes (Limnodrilus hoffmeisteri) were exposed to test substance concentrations of 0, 0.31, 0.63, 1.3, 2.5 and 5.0 mg active ingredient/L for 96 h under static conditions. Oligochaetes were observed for mortality after 48 h and 96 h. Results were analysed for the calculation of 48 h and 96 h LC50. The 48 and 96 hr LC50 values were 2.4 and 1.8 mg active ingredient/L, respectively.