Methanaminium, N,N,N-trimethyl-, 2,2-dimethylpropanoate (1:1)

Long-term toxicity to aquatic invertebrates

Link to relevant study record(s)

Reference

Endpoint:

long-term toxicity to aquatic invertebrates

Type of information:

experimental study

Adequacy of study:

key study

Study period:

June 30, 2017 to December 13, 2017

Reliability:

1 (reliable without restriction)

Qualifier:

according to guideline

Guideline:

OECD Guideline 211 (Daphnia magna Reproduction Test)

Deviations:

no

GLP compliance:

yes (incl. QA statement)

Specific details on test material used for the study:

SOURCE OF TEST MATERIAL
- Source (i.e. manufacturer or supplier) and lot/batch number of test material:
Lot No. 13AP016
- Purity, including information on contaminants, isomers, etc.:
Purity >99%

STABILITY AND STORAGE CONDITIONS OF TEST MATERIAL
- Stability and homogeneity of the test material in the vehicle/solvent under test conditions (e.g. in the exposure medium) and during storage:
Standard stock solution I (1020 mg/L) was prepared by adding 0.0510 g test substance into
50.0 mL with deionized water.
Standard stock solution II (1000 μg/L) was prepared by diluting 0.098 mL standard stock
solution (1020 mg/L) into 100 mL with deionized water.

Under the HPLC conditions, the retention time of test substance was about 0.97 min.

- Solubility and stability of the test material in the solvent/vehicle and the exposure medium:
The concentration of the test substance (Methanaminium, N,N,N-trimethyl-, salt with 2,2-dimethylpropanoic acid ( 1: 1 )) in test medium was determined by HPLC-MS/MS to represent the stability of the test substance in test period. A linear regression equation was obtained with the mean peak squares (Cps.s) vs. the concentration of the Methanaminium, N,N,N-trimethyl-, salt with 2,2-dimethylpropanoic acid (1 :1) (c, mg/L): A=J39.55c+1161.1, with good linearity of r2 =0.9967. The results show that linearity for the concentration range of 10.0 μg/L to 400 μg/L was good. The minimum detection amount of HPLC-MS/MS was 3.00 μg/L, the minimum detection concentration of this method was 10.0 μg/L. The analytical results indicated that the concentration of the test substance was consistent in the test medium throughout the test period (deviation within 20% of the initial concentration). Thus a semi-static method of 72h renewal test procedure was reasonable.

The stock solution of 10.0 mg/L was prepared by dissolving 0.0105-0.0109 g test substance into 1000 mL Elendt M7. The stock solution was tested with ultrasound for 15 minutes and no visible insolubles were detected.

TREATMENT OF TEST MATERIAL PRIOR TO TESTING
- Final concentration of a dissolved solid, stock liquid or gel:
0.018, 0.027, 0.041, 0.061, 0.091 mg/L

Analytical monitoring:

yes

Remarks:

During the test period, the pH values of the control and test media were between 7.23 and 7.33, the Dissolved Oxygen (DO) values varied between 8.35 and 8.96 mg/L, the hardness values varied from 212 and 228 mg (CaC03)/L, and the temperature of the contro

Details on sampling:

- Sampling method:
10.0 mL water samples were taken from each treatment at Od (fresh)~ 3d(aged), 9d(fresh) ~ 12d(aged) and 18(fresh) ~ 21d(aged). The 10 .0 mL samples were filtered through 0.22 μm millipore membrane. The filtrate were analysed by HPLC-MS/MS.

Vehicle:

no

Details on test solutions:

PREPARATION AND APPLICATION OF TEST SOLUTION (especially for difficult test substances)
- Method: Adding the test substance into deionized water.
- Controls: Elendt M7 (dilution medium without test substance)
Refer to the attached tables.

Test organisms (species):

Daphnia magna

Details on test organisms:

TEST ORGANISM
- Common name: Water flea
- Age at study initiation (mean and range, SD): 6 to 24 hours old
- Source: Nanjing University (2010)
- Feeding during test
- Food type: Chlorella vulgaris
- Amount: 0.1-0.2 mg C/Daphnia
- Frequency: Once per day

Test type:

semi-static

Water media type:

freshwater

Limit test:

no

Total exposure duration:

21 d

Hardness:

212 to 228 mg (CaC03)/L
Refer to the attached tables.

Test temperature:

19.8 °C to 20.4 °C
Refer to the attached tables.

pH:

pH 7.23 to 7.33
See attached tables

Dissolved oxygen:

8.35 to 8.96 mg/L
Refer to the attached tables.

Nominal and measured concentrations:

Nominal concentrations of 0.018, 0.027, 0.041,0.061 and 0.091 mg/L
All test concentrations were analyzed at Od (fresh) ~ 3d(aged), 9d(fresh) ~12d(aged) and 18(fresh) ~ 2ld(aged).
For measured concentration, refer to the attached tables.

Details on test conditions:

TEST SYSTEM
- Test vessel: glass beaker
- Type: closed (lid with holes)
- Material, size, headspace, fill volume: glass, size 100 mL, fill volume 50 mL
- Aeration: none
- Renewal rate of test solution (frequency): 72h
- No. of organisms per vessel: 1
- No. of vessels per concentration (replicates): 10
- No. of vessels per control (replicates): 1
- No. of vessels per vehicle control (replicates): 10

TEST MEDIUM / WATER PARAMETERS
- Culture medium different from test medium: no
Refer to the attached tables

OTHER TEST CONDITIONS
- Photoperiod: 16-hour light and 8-hour dark cycle daily
- Light intensity: 1,000 to 1,500 lux

EFFECT PARAMETERS MEASURED (with observation intervals if applicable) :
Oxygen concentration, temperature, hardness and pH values were measured once a week in the control(s) and the highest test substance concentration.

TEST CONCENTRATIONS
- Range finding study
- Test concentrations: 0.01, 0.1 mg/L
- Results used to determine the conditions for the definitive study:
The analytical results from the acetonitrile diluted group show that the analysed concentrations were consistent in the test medium throughout the test period (deviation within 20% of the initial concentration). Thus a semi-static method of 72h renewal test procedure was reasonable.
Refer to the attached tables.

Reference substance (positive control):

no

Key result

Duration:

21 d

Dose descriptor:

LOEC

Effect conc.:

ca. 0.091 mg/L

Nominal / measured:

nominal

Conc. based on:

test mat.

Basis for effect:

reproduction

Key result

Duration:

21 d

Dose descriptor:

NOEC

Effect conc.:

ca. 0.061 mg/L

Nominal / measured:

nominal

Conc. based on:

test mat.

Basis for effect:

reproduction

Key result

Duration:

21 d

Dose descriptor:

LOEC

Effect conc.:

ca. 0.091 mg/L

Nominal / measured:

nominal

Conc. based on:

test mat.

Basis for effect:

mortality

Key result

Duration:

21 d

Dose descriptor:

NOEC

Effect conc.:

ca. 1.061 mg/L

Nominal / measured:

nominal

Conc. based on:

test mat.

Basis for effect:

mortality

Details on results:

- Mortality of control: no mortality in control group
- Any observations (e.g. precipitation) that might cause a difference between measured and nominal values: No visible insolubles were detected.

Reported statistics and error estimates:

(1 ) The reproduction inhibition rate (Ir) for the individual animal was calculated as follows:
%Ir=((N c-NT)/N c)*100

where:
Ir = reproduction inhibition rate;
N c = average number of live off springs of a particular daphnia in the control;
NT = average number of live off springs of a particular daphnia in the treatment;

(2) For the estimation of the lowest observed effect concentration (LOEC), and hence the no observed effect concentration (NOEC), for effects of the chemical on growth rate, the mean growth rate for each concentration were compared with the control mean using a multiple comparison method of Dunnett' s tests (Dunnett' s tests, ANOVA, SPSS 17.0).


The coefficient of variation around the mean number of living offspring produced per parent animal in the control(s) was 11.7%. Therefore, the study met the acceptability criteria (Dissolved Oxygen (DO) values were above 3mg/L, mortality of parent animals in the control was <= 10%, the number of live offspring produced per control parent animal surviving at the end of the test was >= 60) and was considered valid.

Refer to the attached tables.

Validity criteria fulfilled:

yes

Conclusions:

The results showed that under valid semi-static test conditions (72h-renewal), for Daphnia magna reproduction and adult survival, the lowest observed effect concentration (LOEC) was 0.091 mg/L(the average measured concentration 0.080 mg/L) and the no observed effect concentration (NOEC) was 0.061 mg/L (the average measured concentration 0.050 mg/L). For survival of adults the LOEC was 0.091 mg/L (the average measured concentration 0.080 mg/L), the NOEC was 0.061 mg/L (the average measured concentration 0.050 mg/L).

Key value for chemical safety assessment

Fresh water invertebrates

Fresh water invertebrates

Dose descriptor:

NOEC

Effect concentration:

ca. 0.061 mg/L