Benzododecinium chloride

Short-term toxicity to fish

Link to relevant study record(s)

Reference

Reference 1

Endpoint:

short-term toxicity to fish

Type of information:

experimental study

Adequacy of study:

key study

Study period:

2001

Reliability:

1 (reliable without restriction)

Rationale for reliability incl. deficiencies:

guideline study

Qualifier:

according to guideline

Guideline:

OECD Guideline 203 (Fish, Acute Toxicity Test)

Deviations:

no

GLP compliance:

yes (incl. QA statement)

Analytical monitoring:

yes

Vehicle:

yes

Remarks:

Water

Test organisms (species):

Oryzias latipes

Details on test organisms:

Test organism: Length 2.1 cm, Weight 0.12 g

Test type:

semi-static

Water media type:

freshwater

Limit test:

no

Total exposure duration:

96 h

Test temperature:

24 +/- 1ºC

pH:

7.3 to 8

Dissolved oxygen:

5.4 to 8 mg/L

Nominal and measured concentrations:

0, 1.0, 2.2, 4.6, 10, 22, 46, 100 mg/L nominal concentration
0, 1.1, 1.9, 4.3, 11, 21, 47 and 110 mg/L measured concentration (geometric mean)

Details on test conditions:

- Renewal rate of test solution: total amount of test solution every 24 hours
- Test solution volume: 5.0 L per vessel
- No. of vessels per concentration (replicates): 1
- No. of organisms per vessel: 10 per concentration
- Lighting: 16 hours light / 8 hours dark

Reference substance (positive control):

not specified

Key result

Duration:

96 h

Dose descriptor:

EC50

Effect conc.:

ca. 9.35 mg/L

Nominal / measured:

nominal

Conc. based on:

test mat.

Basis for effect:

mortality (fish)

Remarks on result:

other: 95% confidence limit (mg/L) - 6.73 and 12.8

Remarks:

Statistical method: Probit

Validity criteria fulfilled:

not specified

Conclusions:

Under the study conditions, the 96 h LC50 value (95% Confidence interval) and NOEC value were determined to be 9.35 mg/L (nominal) (6.73 & 12.8) and 4.6 mg/L (nominal), respectively.

Executive summary:

A study was conducted to determine the acute toxicity of test substance, C12 ADBAC (purity: >99%), to Oryzias latipes, according to the OECD Guideline 203, in compliance with GLP. The study was performed under semi-static conditions, with a daily renewal of media. Groups of 10 juvenile fish were exposed for 96 h to five concentrations of test substance nominal concentrations of 0, 1.0, 2.2, 4.6, 10, 22, 46 and 100 mg/L, in diluent water. A control group of 10 fish was placed into diluent water alone. Test solution was renewed daily. The exposure levels of test substance in aqueous samples of test media were monitored using a Fluorometry. The mean measured concentrations of test substance were determined to be 0, 1.1, 1.9, 4.3, 11, 21, 47 and 110 mg/L (geometric mean). Since the intended exposure concentrations were substantially achieved and were maintained between renewals of the media, the test results were expressed in terms of their nominal values. Observations of the fish were made after approximately 24, 48, 72 and 96 h of exposure. The 96 h LC50 and LC100 values were determined to be  9.35 mg/L (95% confidence interval: 6.73 & 12.8) and 22 mg/L respectively. The no observed effect concentration (NOEC) was determined to be 4.6 mg/L. No mortality was observed in control group fish and dissolved oxygen concentration was 5.4 to 8. Under the study conditions, the 96 h LC50 value (95% Confidence interval) and NOEC value were determined to be 9.35 mg/L (6.73 & 12.8) (nominal) and 4.6 mg/L (nominal), respectively (NITE, 2001).

Reference 2

Endpoint:

short-term toxicity to fish

Type of information:

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

Adequacy of study:

supporting study

Study period:

From October 15, 1990 to October 19, 1990

Reliability:

2 (reliable with restrictions)

Rationale for reliability incl. deficiencies:

guideline study

Remarks:

KL2 due to RA

Justification for type of information:

Refer to section 13 of IUCLID for details on the read-across justification. The study with the read across substance is considered sufficient to fulfil the information requirements.

Reason / purpose for cross-reference:

read-across source

Qualifier:

according to guideline

Guideline:

EPA OPP 72-1 (Fish Acute Toxicity Test)

Deviations:

no

GLP compliance:

yes

Analytical monitoring:

yes

Details on sampling:

- Concentrations: 0, 0.180, 0.320, 0.490, 0.560 and 0.750 mg/L
- Sampling method: water quality characteristics of the test solutions were measured at test initiation, at the 24, 48 and 72 h renewal events, and at test termination.

Details on test solutions:

Preparation of test solutions:
The test solutions were prepared from a single stock solution of 14C-ADBAC and ADBAC. The stock was prepared by adding 11.70 mg of radiolabelled test substance and 919.5 mg of non-radiolabelled test substance in deionized water and brought to a volume of 250 mL to yield a nominal concentration 3.72 mg/mL test substance. The test solutions were prepared by adding measured aliquots of this stock solution to dilution water at test temperature (22±1°C), and adjusting the final volume to 22L in the test aquaria.

Test organisms (species):

Lepomis macrochirus

Details on test organisms:

Test organism:
- Common name: Bluegill sunfish
- Source: Kurtz’s Fish Hatchery Elverson, PA
- Feeding during test: No
- Length at study initiation: 33-41 mm (Average length 36 mm)
- Weight at study initiation: 0.57-0.98 g (Average weight 0.76 g)
Acclimation:
- Acclimation period: 2 wks
- Food: Commercial trout chow (Purina brand), frozen brine shrimp (Artemia) and Tetra-Min
- Feeding frequency: 5-6 times/wk
- Water (maintenance): dilution water
- Photoperiod: 16h/8h light/dark photoperiod
- Temperature: 22 ± 2.2°C

Test type:

static

Water media type:

freshwater

Limit test:

no

Total exposure duration:

96 h

Hardness:

48-84 mg/L as CaCO3

Test temperature:

21.5-22.0°C

pH:

6.8-7.2

Dissolved oxygen:

6.9-8.8 mg/L

Nominal and measured concentrations:

Nominal test concentration: 0, 0.180, 0.320, 0.490, 0.560 and 0.750 mg/L
Mean measured test concentrations: 0, 0.1973, 0.3171, 0.4555, 0.515 and 0.638 mg/L

Details on test conditions:

Test system:
- Test vessel: glass aquaria (capacity 25L)
- Material, size, headspace, fill volume: 22L
- Aeration: No aeration was provided during the test
- Renewal rate of test solution (frequency/flow rate): 24h renewals
- No. of organisms per vessel: 10
- No. of vessels per concentration: 2
- No. of vessels per control: 2
- Biomass loading rate: 0.35 g/L

Test medium / water parameters:
- Source/preparation of dilution water: the dilution water consisted of water from two deep wells. The well water was initially treated to remove iron and organic impurities (activated carbon filters). Some of this water was then sequentially passed through reverse osmosis (RO) purifiers to reduce ion concentrations. To obtain test water of proper hardness, the RO water and the treated well water (organic impurity removal and iron removal only) were mixed in a 500 gallon capacity fibreglass storage tank to obtain a hardness of approximately 50 mg/L as CaCO3.
- Total organic carbon: <0.8 mg/L
- Dissolved oxygen (DO) concentration: 6.9-8.8 mg/L, which represented 83-106% of air saturation at 22°C
- Conductivity: 192 µMhos/cm
- Metals: <2 µg/L Zinc, <1 µg/L cadmium, <12 µg/L copper
- Chlorine (residual): <0.05 mg/L
- Alkalinity: 68-128 mg/L as CaCO3
- Culture medium different from test medium: No
- Intervals of water quality measurement: Water quality characteristics of the test solutions were measured at test initiation, at the 24, 48 and 72h renewal events, and at test termination.

Other test conditions:
- Photoperiod: 16h/8h light/dark photoperiod
- Light intensity: 74.2-81.3 foot candles

Effect parameters measured (with observation intervals if applicable):
Observations of symptoms of toxicity and mortality were made at 24, 48, 72 and 96h. Mortality is defined as the lack of movement by the test organism after touching the caudal peduncle. Data from the 96 h static-renewal acute test with bluegill sunfish were used to estimate median lethal concentration (LC50).

Test concentrations:
- Range finding study: A 96 h static acute range-finding test was conducted to determine the concentration range of the test substance to use in the definitive test. The range-finding test was conducted with one replicate test chamber concentration with 10 fish/replicate.
- Test concentrations: 0, 0.100, 0.500, 0.625, 0.825 and 1.000 mg/L
- Results used to determine the conditions for the definitive study: Mortality was ≤10% in the control and the 0.1 mg/L nominal test concentration during the 96 h static exposure. The 96h LC50 value (nominal concentration) was 0.556 mg/L of the test substance. Based on these results, nominal test concentrations were selected for the definitive study.

Reference substance (positive control):

no

Key result

Duration:

96 h

Dose descriptor:

LC50

Effect conc.:

0.515 mg/L

Nominal / measured:

meas. (arithm. mean)

Conc. based on:

act. ingr.

Basis for effect:

mortality (fish)

Remarks on result:

other: confidence interval: 0.456-0.638 mg/L

Key result

Duration:

96 h

Dose descriptor:

NOEC

Effect conc.:

0.456 mg/L

Nominal / measured:

meas. (arithm. mean)

Conc. based on:

act. ingr.

Basis for effect:

mortality (fish)

Details on results:

Test substance
- No mortality was observed at the test concentrations of 0.456 mg/L or less during the entire test period. Immobilisation and erratic swimming were noted for some fish in the 0.515 mg/L group. 

Control
- All control fish appeared normal and survived for the duration of the test.

Reported statistics and error estimates:

Estimates of LC50 values were obtained using version 1.61 of the computer program, TOXDAT, cited in Peltier and Weber (Methods for measuring the acute toxicity of effluents to freshwater and marine organisms. 1985, US Environmental Protection Agency).

Sublethal observations / clinical signs:

Table 1. Cumulative mortality data

Concentration (mg/L)	Percent Mortality
	24 h	48 h	72 h	96 h
0.6384	100	100	100	100
0.515	30	50	50	50
0.4555	0	0	0	0
0.3171	0	0	0	0
0.1973	0	0	0	0
Control	0	0	0	0

 

Table 2. Results based on the mortality data

LC50 (mg/L)	24 h	48 h	72 h	96 h
	0.540	0.515	0.515	0.515
95% confidence limits	(0.456-0.638)	(0.456-0.638)	(0.456-0.638)	(0.456-0.638)

 

Validity criteria fulfilled:

yes

Conclusions:

Based on the results of the read across study, similar ((i.e., 96 h LC50 = 0.52 mg a.i./L) or higher effect levels can be expected for the test substance.

Executive summary:

A study was conducted to determine the short-term toxicity to fish of the read across substance, C12 -16 ADBAC (30% active and 95.5-96.5% radiolabelled purity) according to US EPA OPP 72 -1, in compliance with GLP. The experiment was carried out in Bluegill sunfish (Lepomis macrochirus) under static conditions. Fish were exposed for 96 h to nominal read across substance concentrations of 0, 0.180, 0.320, 0.490, 0.560 and 0.750 mg a.i./L, equivalent to mean measured concentrations of 0, 0.1973, 0.3171, 0.4555, 0.515 and 0.638 mg a.i./L. Analytical dose verification was performed. Twenty fish in two replicates were exposed to each test concentration. The solutions were renewed at 24, 48 and 72 h. Observations of symptoms of toxicity and mortality were conducted at 24, 48, 72 and 96 h. The LC50 was calculated based on measured concentrations following a binomial method. No mortality was observed at the test concentrations of 0.456 mg/L or less during the entire test period. Immobilisation and erratic swimming were noted for some fish in the 0.515 mg/L group. Under the study conditions, the 96 h LC50 and NOEC were calculated to be 0.515 and 0.456 mg a.i./L, respectively (Pate and McIntyre, 1991). Based on the results of the read across study, similar or higher effect levels can be expected for the test substance.

Reference 3

Endpoint:

short-term toxicity to fish

Type of information:

experimental study

Adequacy of study:

supporting study

Study period:

From October 15, 1990 to October 19, 1990

Reliability:

1 (reliable without restriction)

Rationale for reliability incl. deficiencies:

guideline study

Qualifier:

according to guideline

Guideline:

EPA OPP 72-1 (Fish Acute Toxicity Test)

Deviations:

no

GLP compliance:

yes

Specific details on test material used for the study:

- Physical state: Yellow liquid (Non-radiolabelled test substance); clear liquid (Radiolabelled test substance)
- Analytical purity: Non-radiolabelled test substance - 30% w/w active substance in aqueous solution; Radiolabelled test substance - 96.5% (by TLC) and 95.5% (by HPLC).
- Lot/batch No.: 05-6K (Non-radiolabelled test substance)
- Stability under test conditions: Non-radiolabelled test substance is hydrolytically and photolytically stable under the conditions of this study and has been shown to be stable in aqueous, alcohol and alcohol/aqueous solutions for extended periods, e.g. at least five years under standard laboratory conditions.
- Storage condition of test material: Radiolabelled and non-radiolabelled test substance was stored refrigerated or frozen in dark for the duration of the study.
- Density: Approximately 0.97 g/mL (Non-radiolabelled test substance)
- Boiling point: Approximately 212°C (Non-radiolabelled test substance)
- Specific activity (if radiolabelling): 25 mCi/mmol

Analytical monitoring:

yes

Details on sampling:

- Concentrations: 0, 0.180, 0.320, 0.490, 0.560 and 0.750 mg/L
- Sampling method: water quality characteristics of the test solutions were measured at test initiation, at the 24, 48 and 72 h renewal events, and at test termination.

Details on test solutions:

Preparation of test solutions:
The test solutions were prepared from a single stock solution of 14C-ADBAC and ADBAC. The stock was prepared by adding 11.70 mg of radiolabelled test substance and 919.5 mg of non-radiolabelled test substance in deionized water and brought to a volume of 250 mL to yield a nominal concentration 3.72 mg/mL test substance. The test solutions were prepared by adding measured aliquots of this stock solution to dilution water at test temperature (22±1°C), and adjusting the final volume to 22L in the test aquaria.

Test organisms (species):

Lepomis macrochirus

Details on test organisms:

Test organism:
- Common name: Bluegill sunfish
- Source: Kurtz’s Fish Hatchery Elverson, PA
- Feeding during test: No
- Length at study initiation: 33-41 mm (Average length 36 mm)
- Weight at study initiation: 0.57-0.98 g (Average weight 0.76 g)
Acclimation:
- Acclimation period: 2 wks
- Food: Commercial trout chow (Purina brand), frozen brine shrimp (Artemia) and Tetra-Min
- Feeding frequency: 5-6 times/wk
- Water (maintenance): dilution water
- Photoperiod: 16h/8h light/dark photoperiod
- Temperature: 22 ± 2.2°C

Test type:

static

Water media type:

freshwater

Limit test:

no

Total exposure duration:

96 h

Hardness:

48-84 mg/L as CaCO3

Test temperature:

21.5-22.0°C

pH:

6.8-7.2

Dissolved oxygen:

6.9-8.8 mg/L

Nominal and measured concentrations:

Nominal test concentration: 0, 0.180, 0.320, 0.490, 0.560 and 0.750 mg/L
Mean measured test concentrations: 0, 0.1973, 0.3171, 0.4555, 0.515 and 0.638 mg/L

Details on test conditions:

Test system:
- Test vessel: glass aquaria (capacity 25L)
- Material, size, headspace, fill volume: 22L
- Aeration: No aeration was provided during the test
- Renewal rate of test solution (frequency/flow rate): 24h renewals
- No. of organisms per vessel: 10
- No. of vessels per concentration: 2
- No. of vessels per control: 2
- Biomass loading rate: 0.35 g/L

Test medium / water parameters:
- Source/preparation of dilution water: the dilution water consisted of water from two deep wells. The well water was initially treated to remove iron and organic impurities (activated carbon filters). Some of this water was then sequentially passed through reverse osmosis (RO) purifiers to reduce ion concentrations. To obtain test water of proper hardness, the RO water and the treated well water (organic impurity removal and iron removal only) were mixed in a 500 gallon capacity fibreglass storage tank to obtain a hardness of approximately 50 mg/L as CaCO3.
- Total organic carbon: <0.8 mg/L
- Dissolved oxygen (DO) concentration: 6.9-8.8 mg/L, which represented 83-106% of air saturation at 22°C
- Conductivity: 192 µMhos/cm
- Metals: <2 µg/L Zinc, <1 µg/L cadmium, <12 µg/L copper
- Chlorine (residual): <0.05 mg/L
- Alkalinity: 68-128 mg/L as CaCO3
- Culture medium different from test medium: No
- Intervals of water quality measurement: Water quality characteristics of the test solutions were measured at test initiation, at the 24, 48 and 72h renewal events, and at test termination.

Other test conditions:
- Photoperiod: 16h/8h light/dark photoperiod
- Light intensity: 74.2-81.3 foot candles

Effect parameters measured (with observation intervals if applicable):
Observations of symptoms of toxicity and mortality were made at 24, 48, 72 and 96h. Mortality is defined as the lack of movement by the test organism after touching the caudal peduncle. Data from the 96 h static-renewal acute test with bluegill sunfish were used to estimate median lethal concentration (LC50).

Test concentrations:
- Range finding study: A 96 h static acute range-finding test was conducted to determine the concentration range of the test substance to use in the definitive test. The range-finding test was conducted with one replicate test chamber concentration with 10 fish/replicate.
- Test concentrations: 0, 0.100, 0.500, 0.625, 0.825 and 1.000 mg/L
- Results used to determine the conditions for the definitive study: Mortality was ≤10% in the control and the 0.1 mg/L nominal test concentration during the 96 h static exposure. The 96h LC50 value (nominal concentration) was 0.556 mg/L of the test substance. Based on these results, nominal test concentrations were selected for the definitive study.

Reference substance (positive control):

no

Key result

Duration:

96 h

Dose descriptor:

LC50

Effect conc.:

0.515 mg/L

Nominal / measured:

meas. (arithm. mean)

Conc. based on:

act. ingr.

Basis for effect:

mortality (fish)

Remarks on result:

other: confidence interval: 0.456-0.638 mg/L

Key result

Duration:

96 h

Dose descriptor:

NOEC

Effect conc.:

0.456 mg/L

Nominal / measured:

meas. (arithm. mean)

Conc. based on:

act. ingr.

Basis for effect:

mortality (fish)

Details on results:

Test substance
- No mortality was observed at the test concentrations of 0.456 mg/L or less during the entire test period. Immobilisation and erratic swimming were noted for some fish in the 0.515 mg/L group. 

Control
- All control fish appeared normal and survived for the duration of the test.

Reported statistics and error estimates:

Estimates of LC50 values were obtained using version 1.61 of the computer program, TOXDAT, cited in Peltier and Weber (Methods for measuring the acute toxicity of effluents to freshwater and marine organisms. 1985, US Environmental Protection Agency).

Sublethal observations / clinical signs:

Table 1. Cumulative mortality data

Concentration (mg/L)	Percent Mortality
	24 h	48 h	72 h	96 h
0.6384	100	100	100	100
0.515	30	50	50	50
0.4555	0	0	0	0
0.3171	0	0	0	0
0.1973	0	0	0	0
Control	0	0	0	0

 

Table 2. Results based on the mortality data

LC50 (mg/L)	24 h	48 h	72 h	96 h
	0.540	0.515	0.515	0.515
95% confidence limits	(0.456-0.638)	(0.456-0.638)	(0.456-0.638)	(0.456-0.638)

 

Validity criteria fulfilled:

yes

Conclusions:

Under the test conditions, the 96h LC50 and NOEC (Lepomis macrochirus) of the test substance were calculated to be 0.515 mg a.i./L and 0.456 mga.i./L respectively.

Executive summary:

A study was conducted to determine the short-term toxicity to fish of the test substance, C12 -16 ADBAC (30% active and 95.5-96.5% radiolabelled purity) according to US EPA OPP 72 -1, in compliance with GLP. The experiment was carried out in Bluegill sunfish (Lepomis macrochirus) under static conditions. Fish were exposed for 96 h to nominal test substance concentrations of 0, 0.180, 0.320, 0.490, 0.560 and 0.750 mg a.i./L, equivalent to mean measured concentrations of 0, 0.1973, 0.3171, 0.4555, 0.515 and 0.638 mg a.i./L. Analytical dose verification was performed. Twenty fish in two replicates were exposed to each test concentration. The solutions were renewed at 24, 48 and 72 h. Observations of symptoms of toxicity and mortality were conducted at 24, 48, 72 and 96 h. The LC50 was calculated based on measured concentrations following a binomial method. No mortality was observed at the test concentrations of 0.456 mg/L or less during the entire test period. Immobilisation and erratic swimming were noted for some fish in the 0.515 mg/L group. Under the test conditions, the 96 h LC50 and NOEC were calculated to be 0.515 and 0.456 mg a.i./L, respectively (Pate, 1991).

Description of key information

The 96 h LC50 value of the test substance, which was determined at 9.35 mg/L or 9350 µg/L (nominal), based on the effects on mortality, has been considered further for hazard and risk assessment.

Key value for chemical safety assessment

Fresh water fish

Fresh water fish

Effect concentration:

9 350 µg/L

Additional information

Study 1: A study was conducted to determine the acute toxicity of test substance, C12 ADBAC (purity: >99%), to Oryzias latipes, according to the OECD Guideline 203, in compliance with GLP. The study was performed under semi-static conditions, with a daily renewal of media. Groups of 10 juvenile fish were exposed for 96 h to five concentrations of test substance nominal concentrations of 0, 1.0, 2.2, 4.6, 10, 22, 46 and 100 mg/L, in diluent water. A control group of 10 fish was placed into diluent water alone. Test solution was renewed daily. The exposure levels of test substance in aqueous samples of test media were monitored using a Fluorometry. The mean measured concentrations of test substance were determined to be 0, 1.1, 1.9, 4.3, 11, 21, 47 and 110 mg/L (geometric mean). Since the intended exposure concentrations were substantially achieved and were maintained between renewals of the media, the test results were expressed in terms of their nominal values. Observations of the fish were made after approximately 24, 48, 72 and 96 h of exposure. The 96 h LC50 and LC100 values were determined to be  9.35 mg/L (95% confidence interval: 6.73 & 12.8) and 22 mg/L respectively. The no observed effect concentration (NOEC) was determined to be 4.6 mg/L. No mortality was observed in control group fish and dissolved oxygen concentration was 5.4 to 8. Under the study conditions, the 96 h LC50 value (95% Confidence interval) and NOEC value were determined to be 9.35 mg/L (6.73 & 12.8) (nominal) and 4.6 mg/L (nominal), respectively (NITE, 2001).

Study 2: A study was conducted to determine the short-term toxicity to fish of the read across substance, C12 -16 ADBAC (30% active and 95.5-96.5% radiolabelled purity) according to US EPA OPP 72 -1, in compliance with GLP. The experiment was carried out in Bluegill sunfish (Lepomis macrochirus) under static conditions. Fish were exposed for 96 h to nominal read across substance concentrations of 0, 0.180, 0.320, 0.490, 0.560 and 0.750 mg a.i./L, equivalent to mean measured concentrations of 0, 0.1973, 0.3171, 0.4555, 0.515 and 0.638 mg a.i./L. Analytical dose verification was performed. Twenty fish in two replicates were exposed to each test concentration. The solutions were renewed at 24, 48 and 72 h. Observations of symptoms of toxicity and mortality were conducted at 24, 48, 72 and 96 h. The LC50 was calculated based on measured concentrations following a binomial method. No mortality was observed at the test concentrations of 0.456 mg/L or less during the entire test period. Immobilisation and erratic swimming were noted for some fish in the 0.515 mg/L group. Under the study conditions, the 96 h LC50 and NOEC were calculated to be 0.515 and 0.456 mg a.i./L, respectively (Pate and McIntyre, 1991). Based on the results of the read across study, similar or higher effect levels can be expected for the test substance.

The available information with test or read across substance suggests that fish is not the most sensitive species. Although a direct comparison of the target substance is not possible with the read across substance due to its UVCB nature, generally their results support the fact that the toxicity of the quaternary ammonium compounds increase with carbon chain length until C14 carbon atoms, afterwhich the trend was reported to be opposite (Nica et al., 2017). Therefore, the LC50 value determined in the study with the test substance has been preferably taken forward for hazard and risk assessment.

Reference: Nicaa VD, Galletb J, Villaa S, and Mezzanotte V, 2017. Toxicity of Quaternary Ammonium Compounds (QACs) as single compounds and mixtures to aquatic non-target microorganisms: Experimental data and predictive models. Ecotoxicol. Environ. Safety 142: 567 -577.