2,3-epoxypropyltrimethylammonium chloride

Long-term toxicity to aquatic invertebrates

Administrative data

Endpoint:

long-term toxicity to aquatic invertebrates

Type of information:

experimental study

Adequacy of study:

key study

Study period:

2004-08-11 to 1004-09-01

Reliability:

1 (reliable without restriction)

Rationale for reliability incl. deficiencies:

other: The study was conducted according to the appropriate OECD test guideline, and in compliance with GLP. Analytical monitoring was also conducted.

Data source

Reference

Reference Type:

study report

Title:

Unnamed

Year:

2004

Report date:

2004

Materials and methods

Test guideline

Qualifier:

according to guideline

Guideline:

OECD Guideline 211 (Daphnia magna Reproduction Test)

Deviations:

no

GLP compliance:

yes (incl. QA statement)

Test material

Test material information

Constituent 1

Reference substance name:

2,3-epoxypropyltrimethylammonium chloride

EC Number:

221-221-0

EC Name:

2,3-epoxypropyltrimethylammonium chloride

Cas Number:

3033-77-0

Molecular formula:

C6H14NO.Cl

IUPAC Name:

N,N,N-trimethyl(oxiran-2-yl)methanaminium chloride

Details on test material:

- Name of test material (as cited in study report): 2,3-Epoxypropyltrimethylammonium chloride / 72.5 % and GMAC (Glycidyl trimethyl ammonium chloride).

- Physical state: yellowish, clear liquid

- Storage condition of test material: refrigerator, 7 +/- 2 degC, protected from moisture and light.

The results are expressed on the basis of the active ingredient.

Sampling and analysis

Analytical monitoring:

yes

Details on sampling:

- Concentrations: The concentrations of 2,3-Epoxypropyltrimethylammonium chloride (EPTAC) were determined in all test item concentrations and controls on days 0, 7, 14, 19 (fresh media) and days 2, 9, 16, 21 (old media). 

Test solutions

Vehicle:

no

Details on test solutions:

PREPARATION AND APPLICATION OF TEST SOLUTION

- Method: a stock solution of 100 mg/L, i.e. 72.5 mg a.i./L was agitated to aid dispersion. No other information is reported. It is assumed by the reviewer that a similar method of dispersion was used to achieve test concentrations from serial dilutions of the stock solution or by adding the adequate amounts of the stock solution to achieve nominal concentrations in the test vessels.

Test organisms

Test organisms (species):

Daphnia magna

Details on test organisms:

TEST ORGANISM

- Clone: 5

- Source: the test organisms were bred at the testing facility, origin Insitut fur Wasser-, Boden- und Lufthygiene (WaBoLu).

- Age of parental stock (mean and range, SD): 2-24 h old, the test organisms were not the first brood progeny.

- Feeding during test

- Food type and amount: The food quantity was a D. subspicatus (0.395 - 1.45 mL) and C. vulgaris (0.375- 0.745 mL) suspension of algae per test vessel (corresponding to 0.1 - 0.2 mg C per Daphnia per day). lt varied according to the growth of the daphnids, but was the same for all test groups on any particular day.

- Frequency: At lest 5 times per week.


ACCLIMATION

- Acclimation conditions (same as test or not): the test organisms were bread in an incubator with similar conditions to the test (temperature, illumination and photoperiod), in 2-3 L glass vessels with approx. 1.8 L culture medium.

- Type and amount of food: a mix of D. subspicatus and C. vulgaris with algae cell density of >10^6 cells/ml ad libitum.

- Feeding frequency: at least 5x per week

- Health during acclimation: the stock is defined as healthy by the authors of the report.

Study design

Test type:

semi-static

Water media type:

freshwater

Limit test:

no

Total exposure duration:

21 d

Test conditions

Hardness:

> 140 mg/L as CaCO3

Test temperature:

range: 20.3 - 21.9 °C

pH:

pH range: 7.7-8.6 control, 7.8-8.5 0.48 mg a.i./L, 8.4-9.0 4.35 mg a.i./L.

Dissolved oxygen:

The final oxygen concentrations of the measured replicates in the test media of nominal 0.49, 4.35 mg a.i./L and the control were found to be greater than or equal to 7.66 mg/L. 

Nominal and measured concentrations:

Nominal concentrations: 0 .05, 0.16, 0.49, 1.45, 4.35 mg a.i./L.

Details on test conditions:

TEST SYSTEM

- Test vessel:

- Material, size, headspace, fill volume: glass beakers (5 (ID) x 8 (h) cm), 100 ml filled with 50 ml test solution.

- Renewal rate of test solution (frequency): 3 times per week

- No. of organisms per vessel: 1

- No. of vessels per concentration (replicates): 10

- No. of vessels per control (replicates): 10


TEST MEDIUM / WATER PARAMETERS

- Source/preparation of dilution water: Elendt M4, according to Elendt (1990), modified to a total hardness of 160-180 mg/L as CaCO3

- Total organic carbon: 1.44 mg/L

- Particulate matter: <0.1 mg/L

- Metals: within acceptable ranges

- Pesticides: <0.025 ug/L

- Chlorine: <0.003 mg/L

- Conductivity: <0.1 uOhm

- Culture medium different from test medium: culture medium - Elendt M4, according to Elendt (1990), modified to a total hardness of 160-180 mg CaCO3/L

- Intervals of water quality measurement: Dissolved oxygen concentration, pH, total hardness and water temperature were measured once a week, in fresh and old media, in one replicate of the control and the highest concentration of the test item.


OTHER TEST CONDITIONS

- Photoperiod: 16h/8h light/dark cycle

- Light intensity: max 20 uE x m^2/s


EFFECT PARAMETERS MEASURED:

o mortality of parent daphnia - measured once per day
o newborn offspring first appearance - daily;
o number of live and dead offspring - daily;
o aborted eggs - daily;
o abnormalities of parent daphnia - each day of observation;
o body length and weight (parent daphnia) - end of test.


RANGE-FINDING STUDY

- Test concentrations: two separate range finding studies: 0.1-100 mg/L and 2.5-10 mg/L

- Results used to determine the conditions for the definitive study: 100 and 30% immobilisation at 100 and 10 mg/L in the first test, while 10% immobilisation was achieved at 10 mg/L in the second test.

Reference substance (positive control):

yes

Remarks:

potassium dichromate

Results and discussion

Effect concentrations

Effect concentrations 1

Duration:

21 d

Dose descriptor:

NOEC

Effect conc.:

0.16 mg/L

Nominal / measured:

nominal

Conc. based on:

act. ingr.

Basis for effect:

reproduction

Effect concentrations 2

Duration:

21 d

Dose descriptor:

LOEC

Effect conc.:

0.49 mg/L

Nominal / measured:

nominal

Conc. based on:

act. ingr.

Basis for effect:

reproduction

Effect concentrations 3

Duration:

21 d

Dose descriptor:

EC50

Effect conc.:

0.55 mg/L

Nominal / measured:

nominal

Conc. based on:

act. ingr.

Basis for effect:

reproduction

Remarks on result:

other: 96% CL 0.47-0.66 mg/L

Details on results:

- Mortality of parent animals: At 1.45 and 4.35 mg a.i./L the immobilisation of parental amounted to 100% at 0.49 mg a.i./L to 50 % after 21 days.

- No. of offspring produced: 0.49 mg a.i./L treatment showed a significantly reduced number of offspring compared to the control.

- Body length and weight of parent animals: body length at the 0.49 mg a.i./L treatment level was significantly reduced compared to the control.

- Time to first brood release or time to hatch: no statistical difference between control and treatments reported.

- Brood size: the average number of juveniles per parent in the control group was 108 after 21 days. At 0.49 mg/L the number of juveniles per parent was statistically significantly reduced. 

Results with reference substance (positive control):

- Results with reference substance valid? yes, according to DIN 38412 L 30 the validity range is 1.0-2.5 mg/L during a 24 h exposure.

- Relevant effect levels: EC50 after 24h was 1.94 mg/L

- Other:

Reported statistics and error estimates:

The EC50 value and CI were calculated by probit analysis. The NOEC was calculated by Kruskal-Wallis ANOVA on Ranks, DUNN's method.

Any other information on results incl. tables

Table 1: Effect of test material on growth and survival of D. magna (average per 10 replicates unless otherwise stated) during the 21 day test.

Treatment (mg a.i./L) nominal concentration	Mortality of parents (dead or immobile)		Total no. of offspring	No. of juveniles per parent alive (± SD)	Day of first appearance of juveniles	Mean length (mm)	Mean weight (mg)
	No Dead (per 10 replicates)	% (per 10 replicates)
Control	1	10	970	108 ± 5.0	7.1	5.39	1.27
4.35	10	100	-	0 ± 0	-	-	-
1.45	10	100	-	0 ± 0	7.9	-	-
0.49	5	50	365	73 ± 15.5	8.0	4.6	0.22
0.16	1	10	961	107 ± 14.1	7.7	5.33	0.68
0.05	0	0	1025	103 ± 3.8	7.1	5.35	1.05

- denotes when an endpoint is not applicable, e.g. the parent daphnia have died before the first appearance of juveniles.

 

Table 2: Parental daphnia immobilisation results.

Endpoint	Result (mg a.i./L, nominal)
NOEC	0.16
LOEC	0.49
EC50	0.34 (0.27-0.42 95% CL)

 

Table 3: Nominal and measured concentrations during 21 days.

	Measured concentration (mg a.i./L) results
Nominalconcentr.	0 (control)	4.35	1.45	0.49	0.16	0.05
Range (min.-max.)	< LOQ	4.72	1.5-1.54	0.48-0.57	0.161-0.186	0.051-0.058

 

The highest treatment level is only presented as one measurement because the day 0 and 2 values are reported as an average and thereafter the organisms perished.

Applicant's summary and conclusion

Validity criteria fulfilled:

yes

Conclusions:

A 21 day NOEC value of 0.16 mg a.i./l has been determined for the effects of the test substance on reproduction of D. magna during a 21 day chronic test.