Quaternary ammonium compounds, C14-18 (even numbered) and C18 unsaturated-alkyl-hydroxyethyl-dimethyl, chlorides

Short-term toxicity to aquatic invertebrates

Link to relevant study record(s)

Reference

Endpoint:

short-term toxicity to aquatic invertebrates

Type of information:

experimental study

Adequacy of study:

key study

Study period:

29 March to 08 June 2011

Reliability:

1 (reliable without restriction)

Rationale for reliability incl. deficiencies:

guideline study

Remarks:

This study was performed according to OECD Guideline 202 with GLP statement. All validity criteria were fulfilled.

Qualifier:

according to guideline

Guideline:

OECD Guideline 202 (Daphnia sp. Acute Immobilisation Test)

Deviations:

no

Qualifier:

according to guideline

Guideline:

EU Method C.2 (Acute Toxicity for Daphnia)

Principles of method if other than guideline:

Not applicable

GLP compliance:

yes (incl. QA statement)

Remarks:

Swiss GLP Compliance Programme (inspected on 22-26 March and 12-16 April 2010 / signed on 11 January 2011)

Specific details on test material used for the study:

SOURCE OF TEST MATERIAL- Source and lot/batch No.of test material: Chimex / 0141653- Expiration date of the lot/batch: 30 June 2011- Purity test date: 28 September 2010STORAGE CONDITIONS OF TEST MATERIAL- Storage condition of test material: At room temperature at about 20 °C, in the dark

Analytical monitoring:

yes

Details on sampling:

Analysis of the Test Item ConcentrationsFor the determination of the actual test item concentrations, triplicate samples were taken from each treatment at the start and at the end of the test (after 48 hours).For the 48-hour stability samples, additional flasks with adequate volumes of the freshly prepared test media were incubated under the same conditions as in the actual test (but without daphnids) over 48 hours. Sampling from the test beakers containing the daphnids was not possible since the test medium volume per beaker (50 mL) was too small for the analytical requirements (500 mL per sample).All samples were stored deep-frozen (at about -20 °C) immediately after sampling. In pre-experiments (non-GLP), the test item proved to be sufficiently stable in the test water under these storage conditions.The concentrations of the test item Chimexane CL were analyzed in one of the triplicate test media samples from the two highest nominal concentrations (0.15 and 0.32 mg/L) and from the control from both sampling times (0 and 48 hours).The samples of the lower test concentrations were not analyzed since these concentrations were below the 48-hour NOEC determined in this test and, thus, were not relevant for the interpretation of the biological results.

Vehicle:

yes

Details on test solutions:

PREPARATION AND APPLICATION OF TEST SOLUTION - Method: A stock solution with a nominal concentration of 32 mg/L was prepared by dissolving 32.1 mg of the test item in 1000 mL of test water using intense stirring for 5 minutes at room temperature. Adequate volumes of this stock solution were diluted with test water to prepare the test media of all test item concentrations. The test media were prepared just before introduction of the daphnids (i.e., start of the test).- Controls: Test water without test item) was tested in parallel

Test organisms (species):

Daphnia magna

Details on test organisms:

TEST ORGANISM - Strain/clone: Daphnia magna Straus; a clone of this species (clone 5) - Source: University of Sheffield / UK in 1992; Since that time, the clone has been bred at Harlan Laboratories in reconstituted water of the quality identical to the water quality used in the tests (in respect to pH, main ions, and total hardness) and under temperature and light conditions identical to those of the tests. - During breeding, daphnids are generally fed three times a week with an algal suspension of the green algae Desmodesmus subspicatus CHODAT, Strain No. 86.81 SAG, supplied by the Collection of Algal Cultures (SAG, Institute for Plant Physiology, University of Göttingen, 37073 Göttingen / Germany) and cultivated at Harlan Laboratories under standardized conditions or a mixture of this algal suspension and a commercial fish diet (Tetra Min® Hauptfutter, supplied by TETRA-Werke, 49304 Melle / Germany).- At the start of the test, the organisms used in the test were 6-24 hours old and were not first brood progeny. - Feeding during test: Daphnids were not fed during the test.

Test type:

static

Water media type:

freshwater

Limit test:

no

Total exposure duration:

48 h

Post exposure observation period:

None

Hardness:

2.5 mmol/L (= 250 mg/L as CaCO3)

Test temperature:

20-21 °C

pH:

At the beginning and end of the test period, the pH value of the test media was between 7.9 and 8.0.

Dissolved oxygen:

At the beginning and end of the test period, the dissolved oxygen concentrations in the test media and control were at least 8.0 mg/L.

Salinity:

None

Conductivity:

No data

Nominal and measured concentrations:

Nominal concentrations: 0.015, 0.032, 0.070, 0.15 and 0.32 mg/L (corresponding to 0.0045, 0.0096, 0.021, 0.045 and 0.096 mg hydroxyethyl oleyl dimonium chloride/L corrected for water content)

Details on test conditions:

TEST SYSTEM - Test vessel: Test was performed in 100-mL glass beakers filled with 50 mL of test medium - Type: closed; test vessels were covered with glass plates to reduce the loss of water by evaporation and to avoid the entry of dust into the solutions. - Aeration: The test water was aerated prior to the start of the study until oxygen saturation was reached. During the test period, the test water was not aerated. - Renewal of test solution: A static test without test medium renewal was performed. - No. of organisms per vessel: 5 - No. of vessels per concentration (replicates): 4 - No. of vessels per control (replicates): 4 - The volume of test solution provided for each daphnid was 10 mL. Thus, the requirement of the test guidelines for the minimum volume of 2 mL test medium per daphnid was fulfilled. The daphnids were randomly distributed to the test vessels at initiation of the test. TEST MEDIUM / WATER PARAMETERS - Source/preparation of dilution water: Reconstituted test water according to ISO 6341 was used in the study. - The ratio of Ca:Mg and Na:K was 4:1 and 10:1, respectively, based on molarity. OTHER TEST CONDITIONS - Test was performed in a temperature-controlled room with continuous monitoring of the room temperature. The water temperature was maintained at 20-21 °C. - Photoperiod: 16-hour light to 8-hour dark cycle with a 30-minute transition period - Light intensity: Light intensity during the light period was approximately between 485 and 640 Lux. EFFECT PARAMETERS MEASURED- Determination of Immobility: The immobility of the daphnids was determined by visual inspection after 24 and 48 hours of exposure. Those daphnids not able to swim within 15 seconds after gentle agitation of the test vessel were considered to be immobilized.- Water Quality Criteria: At the start and end of the test, the pH values, dissolved oxygen concentrations and water temperature were determined at each treatment. The appearance of the test media was visually recorded at the start of the test and after 24 and 48 hours. RANGE-FINDING STUDY - The selection of the test concentrations was based on the results of a range-finding test (non-GLP).

Reference substance (positive control):

yes

Remarks:

potassium dichromate

Key result

Duration:

24 h

Dose descriptor:

EC0

Effect conc.:

0.13 mg/L

Nominal / measured:

meas. (geom. mean)

Conc. based on:

test mat.

Basis for effect:

mobility

Remarks on result:

other: corresponding to 0.039 mg hydroxyethyl oleyl dimonium chloride/L

Key result

Duration:

24 h

Dose descriptor:

EC50

Effect conc.:

0.23 mg/L

Nominal / measured:

meas. (geom. mean)

Conc. based on:

test mat.

Basis for effect:

mobility

Remarks on result:

other: corresponding to 0.069 mg hydroxyethyl oleyl dimonium chloride/L

Key result

Duration:

24 h

Dose descriptor:

EC100

Effect conc.:

> 0.26 mg/L

Nominal / measured:

meas. (geom. mean)

Conc. based on:

test mat.

Basis for effect:

mobility

Remarks on result:

other: corresponding to > 0.078 mg hydroxyethyl oleyl dimonium chloride/L

Key result

Duration:

48 h

Dose descriptor:

EC0

Effect conc.:

0.13 mg/L

Nominal / measured:

meas. (geom. mean)

Conc. based on:

test mat.

Basis for effect:

mobility

Remarks on result:

other: corresponding to 0.039 mg hydroxyethyl oleyl dimonium chloride/L

Key result

Duration:

48 h

Dose descriptor:

EC50

Effect conc.:

0.18 mg/L

Nominal / measured:

meas. (geom. mean)

Conc. based on:

test mat.

Basis for effect:

mobility

Remarks on result:

other: (corresponding to an EC50 for hydroxyethyl oleyl dimonium of 0.054 mg/L with 95% confidence limits of 0.039 and 0.078 mg/L

Key result

Duration:

48 h

Dose descriptor:

EC100

Effect conc.:

0.26 mg/L

Nominal / measured:

meas. (geom. mean)

Conc. based on:

test mat.

Basis for effect:

mobility

Remarks on result:

other: corresponding to 0.078 mg hydroxyethyl oleyl dimonium chloride/L

Details on results:

- During the first 24 hours of the test, no immobilized test organisms were determined in the control and up to and including the mean measured test item concentration of 0.13 mg/L. At the highest mean measured test concentration of 0.26 mg/L, 95% of test organisms were found to be immobile after 24 hours.- The 24-hour EC50 of the test item was calculated to be 0.23 mg/L (corresponding to 0.069 mg hydroxyethyl oleyl dimonium chloride/L). The 95% confidence limits could not be determined. The 24-hour EC0 was at the mean measured concentration of 0.13 mg/L (corresponding to 0.039 mg hydroxyethyl oleyl dimonium chloride/L). The 24-hour EC100 was above the highest mean measured test concentration of 0.26 mg/L (corresponding to 0.078 mg hydroxyethyl oleyl dimonium chloride/L).- After 48 hours of exposure, no immobilized test organisms were determined in the control and up to and including the mean measured test item concentration of 0.13 mg/L. At the highest mean measured test concentration of 0.26 mg/L, all test organisms were found to be immobile.- The 48-hour EC50 of the test item was calculated to be 0.18 mg/L with 95% confidence limits of 0.13 and 0.26 mg/L (corresponding to an EC50 for hydroxyethyl oleyl dimonium of 0.054 mg/L with 95% confidence limits of 0.039 and 0.078 mg/L). The 48-hour EC0 and NOEC of Chimexane CL were both at the mean measured test item concentration of 0.13 mg/L (corresponding to 0.039 mg hydroxyethyl oleyl dimonium chloride/L). The 48-hour EC100 was at the mean measured test item concentration of 0.26 mg/L (corresponding to 0.078 mg hydroxyethyl oleyl dimonium chloride/L). Thus, the concentration-effect relationship in this test was rather steep.- No remarkable observations were made concerning the appearance of the test media. All test media were clear solutions throughout the entire test duration.

Results with reference substance (positive control):

For evaluation of the quality of the daphnia clone and the experimental conditions, potassium dichromate is tested as a positive control twice a year. The result of the latest positive control test in April 2011 (48-hour EC50: 0.62 mg/L, Harlan Laboratories Study D25745) showed that the sensitivity of the test organisms was within the internal historical range (48-hour EC50 from 2000 to 2011: 0.43-1.1 mg/L).

Reported statistics and error estimates:

The 24-hour EC50 could not be calculated by Probit Analysis or Moving Average Interpolation due to the steep concentration-effect relationship. Instead, the 24-hour EC50 was calculated by linear interpolation of the immobility between the two consecutive test concentrations with 0 and 95% immobility on a semi-logarithmic scale (the 95% confidence limits could not be determined).The 48-hour EC50 was determined as the geometric mean value of the two consecutive test concentrations with 0 and 100% immobility (EC and EC0100). The 95% confidence limits correspond to the EC and EC0100 values.

Table 6.1.3/1: Effect of Chimexane CL on the Mobility of Daphnia magna

 

Nominal test item concentration (mg/L)	Mean measured concentration (mg/L)	No. of daphnids tested	Immobilized daphnids after 24 hours		Immobilized daphnids after 48 hours
			No.	%	No.	%
Control	-	20	0	0	0	0
0.015	n.a.	20	0	0	0	0
0.032	n.a.	20	0	0	0	0
0.070	n.a.	20	0	0	0	0
0.15	0.13	20	0	0	0	0
0.32	0.26	20	19	95	20	100

n.a.: not analyzed

 

Analysis of the Test Item Concentrations

The measured concentrations of Chimexane CL in the test media of the two highest nominal test concentrations of 0.15 and 0.32 mg/L (corresponding to 0.045 and 0.096 mg hydroxyethyl oleyl dimonium chloride/L) were 91 and 82% of the nominal values, respectively, at the start of the test. At the end of the test, 89 and 79% of the nominal values were found, respectively.

All reported biological results are related to the mean measured test item concentrations of 0.13 and 0.26 mg/L (corresponding to 0.039 and 0.078 mg hydroxyethyl oleyl dimonium chloride/L), which were calculated as the geometric means of the concentrations measured at the start and at the end of the test.

 

Water Quality Criteria

At the beginning and end of the test period, the dissolved oxygen concentrations in the test media and control were at least 8.0 mg/L, the pH value of the test media was between 7.9 and 8.0, and the water temperature during the test was 20-21 °C.

The test is considered to be valid, as in the control no daphnids showed immobilization or other signs of disease or stress (e.g., discoloration or unusual behavior such as trapping at the surface water). Furthermore, the dissolved oxygen concentration at the end of the test was ≥3 mg/L in the control and test vessels.

Validity criteria fulfilled:

yes

Conclusions:

Under the test conditions, the 48-hour EC50 of the test substance was calculated to be 0.18 mg/L with 95% confidence limits of 0.13 and 0.26 mg/L (corresponding to an EC50 for hydroxyethyl oleyl dimonium of 0.054 mg/L with 95% confidence limits of 0.039 and 0.078 mg/L).

Executive summary:

A Study was performed according to OECD Guideline 202 with GLP statement, to assess the 48 h-acute toxicity of the of the test substance to Daphnia magna, under static conditions.

 

The nominal test item concentrations tested were 0.015, 0.032, 0.070, 0.15 and 0.32 mg/L (corresponding to 0.0045, 0.0096, 0.021, 0.045 and 0.096 mg Hydroxyethyl oleyl dimonium chloride/L corrected for water content). Additionally, a control group was tested in parallel.

 

The measured concentrations of Chimexane CL in the test media of the two highest nominal test concentrations of 0.15 and 0.32 mg/L (corresponding to 0.045 and 0.096 mg hydroxyethyl oleyl dimonium chloride per liter) were 91 and 82% of the nominal values, respectively, at the start of the test. At the end of the test, 89 and 79% of the nominal values, respectively, were found. All reported biological results are related to the mean measured test item concentrations of 0.13 and 0.26 mg/L (corresponding to 0.039 and 0.078 mg hydroxyethyl oleyl dimonium chloride per liter, respectively), which were calculated as the geometric means of the concentrations measured at the start and at the end of the test.

 

During the first 24 hours of the test, no immobilized test organisms were determined in the control and up to and including the mean measured test item concentration of 0.13 mg/L. At the highest mean measured test concentration of 0.26 mg/L, 95% of test organisms were found to be immobile after 24 hours. After 48 hours of exposure, no immobilized test organisms were determined in the control and up to and including the mean measured test item concentration of 0.13 mg/L. At the highest mean measured test concentration of 0.26 mg/L, all test organisms were found to be immobile. No remarkable observations were made concerning the appearance of the test media. All test media were clear solutions throughout the entire test duration.

 

Under the test conditions, the 48-hour EC50 of the test substance was calculated to be 0.18 mg/L with 95% confidence limits of 0.13 and 0.26 mg/L (corresponding to an EC50 for hydroxyethyl oleyl dimonium of 0.054 mg/L with 95% confidence limits of 0.039 and 0.078 mg/L).

Description of key information

Under the test conditions, the 48-hour EC50 of the test substance was calculated to be 0.18 mg/L with 95% confidence limits of 0.13 and 0.26 mg/L corresponding to an EC50 for hydroxyethyl oleyl dimonium of 0.054 mg/L with 95% confidence limits of 0.039 and 0.078 mg/L.

Key value for chemical safety assessment

Fresh water invertebrates

Fresh water invertebrates

Effect concentration:

0.054 mg/L

Additional information

A reliable study was available and was performed according to OECD Guideline 202 with GLP statement, to assess the 48 h-acute toxicity of the of the test substance to Daphnia magna, under static conditions. The nominal test item concentrations tested were 0.015, 0.032, 0.070, 0.15 and 0.32 mg/L (corresponding to 0.0045, 0.0096, 0.021, 0.045 and 0.096 mg Hydroxyethyl oleyl dimonium chloride/L corrected for water content). Additionally, a control group was tested in parallel.

This study was considered as acceptable for that endpoint.