Dimethyldioctylammonium chloride

Long-term toxicity to aquatic invertebrates

Link to relevant study record(s)

Reference

Endpoint:

long-term toxicity to aquatic invertebrates

Type of information:

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

Adequacy of study:

key study

Reliability:

2 (reliable with restrictions)

Rationale for reliability incl. deficiencies:

other: Read-across to K1 study therefore K2 is the maximum Klimisch value.

Justification for type of information:

Read-across approach - see read-across justification in section 13.

Reason / purpose for cross-reference:

read-across source

GLP compliance:

yes

Analytical monitoring:

yes

Details on sampling:

Water samples were collected from one test chamber of each treatment and control group two days prior to the start of the test to confirm the operation of the diluter. Water samples also were collected from alternating replicate test chambers at the beginning of the test, at weekly intervals during the test and at the end of the test to determine concentrations of the test substance. All samples were collected at mid-depth from each test chamber, placed in a culture tube containing an equal volume of a methanol:0.2 % formic acid solution, and processed immediately for analysis.

Vehicle:

yes

Details on test solutions:

Stock solutions were prepared weekly during the test. All glassware used in preparation of the stock solutions was acid washed in 10 % HCl prior to use. The stock and test concentrations were prepared on the basis of active ingredient (AI) of the test substance. A primary stock solution was prepared in DMF at a nominal concentration of 800 μg AI/mL. Proportional dilutions of the primary stock were made in DMF to prepare additional stock solutions at nominal concentrations of 120, 200, 310 and 500 μg AI/mL. The stock solutions were mixed by inversion, and were clear and colorless in appearance. The stock solutions were delivered to the diluter mixing chambers (at a rate of 15.5 μL/minute) where they were mixed with dilution water (at a rate of 155 mL/minute) to achieve the desired test concentrations of 12, 20, 31, 50 and 80 μg AI/L. The solvent control was prepared by injecting DMF into the mixing chamber assigned to the solvent control. The concentration of DMF in the solvent control and all treatment groups was 0.1 mL/L. The test solutions appeared clear and colorless in the diluter mixing chambers and in the test chambers at test initiation and termination.

Test organisms (species):

Daphnia magna

Details on test organisms:

The cladoceran, Daphnia magna, was selected as the test species for this study. Daphnids are representative of an important group of aquatic invertebrates and were selected for use in the test based upon past history of use and ease of culturing in the laboratory. Daphnid neonates used in the test were less than 24 hours old.
Adult daphnids were cultured in water from the same source and at approximately the same temperature as used during the test. During the 14-day period preceding the test, water temperatures ranged from 19.9 to 21.2 °C, measured with a hand-held liquid-in-glass thermometer. The pH of the water ranged from 8.4 to 8.6, measured with a Fisher Scientific Accumet Model 915 pH meter. Dissolved oxygen concentrations ranged from 7.6 to 8.9 mg/L (≥84% of saturation), measured with a Yellow Springs Instruments Model 51B dissolved oxygen meter.
The adult daphnids in the cultures were at least 15 days old prior to collection of the juveniles for testing. The adults showed no signs of disease or stress during the culture period, and no ephippia were produced in the cultures. Neonate daphnids were obtained for testing from five individual adult daphnids that had produced at least one prior brood. At test initiation, the juvenile daphnids were collected from the cultures and indiscriminately transferred one and two at a time to transfer chambers (e.g., 10 mL glass beakers) until each chamber contained five neonates. Each transfer chamber was indiscriminately assigned to a test compartment, and the neonates were transferred to the test compartments to initiate the test. All transfers were made below the water surface using wide-bore pipettes.
During culture and testing, daphnids were fed a mixture of yeast, Cerophyll, and trout chow, as well as a suspension of the freshwater green alga, Selenastrum capricornutum. Daphnids were fed three times per day through Day 7 of the test and then were fed four times per day until the last day of the test. There were no known levels of contaminants reasonably expected to be present in the diet that were considered to interfere with the purpose or conduct of the test.

Test type:

flow-through

Water media type:

freshwater

Limit test:

no

Total exposure duration:

21 d

Hardness:

124 to 132 mg/L as CaCO3

Test temperature:

20.0 to 20.2°C

pH:

8.4 to 8.5

Dissolved oxygen:

>= 82 % of saturation (7.4 mg/L)

Nominal and measured concentrations:

- Control: < LOQ
- Solvent control: < LOQ
- Nominal concentration: 12, 20, 31, 50 and 80 µg AI/L
- Measured concentration: 9.2, 17, 27, 46 and 77 µg AI/L

Details on test conditions:

- Continuous-flow through system: Two replicates of each concentration (12, 20, 31, 50 and 80 µg AI/L) and controls were tested with ten test animals. The test was carried out under a 16-hour light – 8-hour darkness regimen.
- Test conditions: pH 8.4 - 8.5, dissolved oxygen >= 82 % saturation, temperature 20.0 - 20.2 °C.
- Feed: The Daphnids were fed a mixture of yeast, Cerophyll and trout chow, as well as a suspension of freshwater green algae Selenastrum capricornutum.
- Observations: First-generation Daphnid were observed daily during the test and the number of dead and immobile Daphnids were recorded along with any clinical signs of toxicity. The presence of eggs in the brood pouch, aborted eggs, males or ephippia were also recorded daily. With the onset of reproduction, neonates produced by the first-generation Daphnids were counted and then discarded every Monday, Wednesday and Friday during the test. The body length and dry weight of each surviving first-generation Daphnid were measured at the end of the test.

Duration:

21 d

Dose descriptor:

EC50

Effect conc.:

0.077 mg/L

Nominal / measured:

meas. (arithm. mean)

Conc. based on:

act. ingr.

Basis for effect:

immobilisation

Duration:

21 d

Dose descriptor:

EC50

Effect conc.:

> 0.077 mg/L

Nominal / measured:

meas. (arithm. mean)

Conc. based on:

act. ingr.

Basis for effect:

reproduction

Duration:

21 d

Dose descriptor:

LOEC

Effect conc.:

0.046 mg/L

Nominal / measured:

meas. (arithm. mean)

Conc. based on:

act. ingr.

Basis for effect:

growth

Key result

Duration:

21 d

Dose descriptor:

NOEC

Effect conc.:

0.027 mg/L

Nominal / measured:

meas. (arithm. mean)

Conc. based on:

act. ingr.

Basis for effect:

growth

Duration:

21 d

Dose descriptor:

other: Maximum Acceptable Toxicant Concentration

Effect conc.:

35 mg/L

Nominal / measured:

meas. (arithm. mean)

Conc. based on:

act. ingr.

Basis for effect:

growth

Details on results:

- Effect data (Mortality): Survival in the 9.2, 17, 27, 46 and 77 µg AI/L treatment groups was 95, 100, 95, 90 and 50 % respectively at test termination.
- Survival: NOEC 46 µg/L; LOEC 77 µg/L; 21-day EC50 77 µg AI/L
- Other effects: Surviving Daphnids in the control groups and in the 9.2, 17, 27 and 46 µg AI/L treatment groups appeared normal during the test, with the exception of one Daphnid in the 17 µg AI/L treatment group that was lethargic at test termination. In the highest concentration, all surviving Daphnids at test termination were smaller in size compared to the controls.
There was no apparent delay in the onset of neonate production at any test concentration. With the exception of two immobile neonates in the 77 µg/L group on Day 16 of the test, no immobile neonates were noted in the control or treatment groups during the test. On Day 21, four aborted or shed eggs were present in one of the 77 µg/L test concentrations. No males or ephippia were produced during the test.
Daphnids in the 9.2, 17, 27, 46 and 77 µg/L treatment groups produced an average of 6.05, 8.25, 8.15, 8.37 and 6.82 live young per reproductive day, respectively. There was a statistically significant difference in reproduction in the lowest concentration tested, however, the slight decrease in this group was not considered to be treatment-related, since there were no treatment-related effects on reproduction at the higher concentrations.
Reproduction: NOEC 77 µg/L; 21-day EC50 > 77 µg/L
Daphnids in the 9.2, 17, 27, 46 and 77 µg AI/L treatment groups had mean lengths of 4.39, 4.47, 4.50, 4.32 and 3.74 mm, respectively and mean dry weights of 0.64, 0.69, 0.70, 0.53 and 0.37 mg respectively. Growth measurements from the surviving daphnids in the 77 µg AI/L treatment group were excluded from statistical analysis due to the statistically significant effects on survival in that group. There was a statistically significant decrease in mean length and dry weight in the 9.2 and 46 µg AI/L groups in comparison with the controls. However, this was not considered to be treatment-related since there were no treatment-related effects on growth at the next two higher test concentrations.
Growth: NOEC 27 µg/L; LOEC 46 µg/L
- Results of controls: None

Validity criteria fulfilled:

yes

Conclusions:

Daphnia magna exposed to test item at mean measured concentrations of 9.2 to 77 μg AI/L for 21 days showed no significant reductions in survival, reproduction or growth at concentrations of 27 μg AI/L and below. Treatment-related reductions in growth (length and dry weight) were apparent at concentrations of 46 μg AI/L and above, while a treatment-related reduction in survival was apparent only at 77 μg AI/L, the highest concentration tested. Consequently, the NOEC for this study, based on the most sensitive endpoint tested (growth) was 27 μg AI/L and the LOEC was 46 μg AI/L. The MATC was calculated to be 35 μg AI/L. The 21-day EC50 value for adult mortality/immobility was estimated to be 77 μg AI/L. The 21-day EC50 value for reproduction was >77 μg AI/L.

Executive summary:

A study was carried out according to OECD Guideline 211 (Daphnia magna Reproduction Test) and EPA OPPTS 850.1300 (Daphnid Chronic Toxicity Test). Daphnia magna were exposed to a geometric series of five test concentrations (12, 20, 31, 50 and 80 µg AI/L), a negative control and a solvent control for 21 day under continuous flow-through conditions. Observations were made daily to assess for effects on survival, growth and reproduction.

Daphnia magna exposed concentrations of 9.2 to 77 µg AI/L for 21 days showed no significant reductions in survival, reproduction or growth at concentrations of 27 µg AI/L and below. Treatment-related reductions in growth (length and dry weight) were apparent at concentrations of 46 µg AI/L and above, while a treatment-related reduction in survival was apparent only at 77 µg AI/L.

Survival: NOEC 46 µg AI/L; LOEC 77 µg AI/L

Reproduction: NOEC 77 µg AI/L ; 21-day EC50 > 77 µg AI/L

Growth: NOEC 27 µg AI/L; LOEC 46 µg AI/L

Description of key information

Only one long term invertebrate test is available, based on the read-across substance N,N-Didecyl-N,N-dimethylammonium carbonate (DDACarbonate). The study is a reliable, conducted according to OECD Guideline 211 and EPA OPPTS 850.1300, complying with GLP. Daphnia magna were exposed to a geometric series of five test concentrations (12, 20, 31, 50 and 80 µg AI/L) for 21 days under continuous flow-through conditions.  Treatment-related reductions in growth (length and dry weight) were apparent at concentrations of 46 µg AI/L and above, while a treatment-related reduction in survival was apparent only at 77 µg AI/L. No effects were observed on reproduction at any of the test concentrations. The NOEC based on the most sensitive endpoint, growth was determined at 27 µg AI/L and the LOEC 46 µg AI/L.

Key value for chemical safety assessment

Fresh water invertebrates

Fresh water invertebrates

Effect concentration:

0.027 mg/L