2-hydroxyethanesulphonic acid, compound with 4,4'-[hexane-1,6-diylbis(oxy)]bis[benzenecarboxamidine] (2:1)

Short-term toxicity to aquatic invertebrates

Administrative data

Endpoint:

short-term toxicity to aquatic invertebrates

Type of information:

experimental study

Adequacy of study:

key study

Study period:

May-July 2018

Reliability:

1 (reliable without restriction)

Rationale for reliability incl. deficiencies:

guideline study

Data source

Reference

Reference Type:

study report

Title:

Unnamed

Year:

2018

Report date:

2018

Materials and methods

Test guideline

Qualifier:

according to guideline

Guideline:

OECD Guideline 202 (Daphnia sp. Acute Immobilisation Test)

GLP compliance:

yes

Test material

Test material information

Constituent 1

Reference substance name:

2-hydroxyethanesulphonic acid, compound with 4,4'-[hexane-1,6-diylbis(oxy)]bis[benzenecarboxamidine] (2:1)

EC Number:

211-533-5

EC Name:

2-hydroxyethanesulphonic acid, compound with 4,4'-[hexane-1,6-diylbis(oxy)]bis[benzenecarboxamidine] (2:1)

Cas Number:

659-40-5

Molecular formula:

C20H26N4O2.2C2H6O4S

IUPAC Name:

4-[6-(4-carbamimidoylphenoxy)hexoxy]benzenecarboximidamide;2-hydroxyethanesulfonic acid

Test material form:

solid: particulate/powder

Details on test material:

Appearance: White to slightly yellow powder

Specific details on test material used for the study:

Batch/Lot Number: 42964
Expiry date: 31 January 2019

Sampling and analysis

Analytical monitoring:

yes

Test solutions

Vehicle:

yes

Remarks:

aqueous test medium

Details on test solutions:

A stock solution with a concentration of 100 mg/L (nominal) was prepared with Test Item and test medium (ISO medium) by direct addition with mixing and using approximately 5-minute ultra-sonication. The test solutions were prepared by appropriate dilution of this stock solution just before introduction of the Daphnia (start of experiment, Day 0), distributed into test vessels prior to introduction of Daphnia

Test organisms

Test organisms (species):

Daphnia magna

Details on test organisms:

Species and strain: Daphnia magna
Source: Szent István University, Department of Aquaculture, 2100 Gödöllő, Páter Károly u. 1. – Hungary
Justification of strain: Daphnia magna is the standard species of the acute immobilisation test.
Number of animals: There were 20 animals in test and control groups respectively, divided into 4 replicates
(5 animals / replicate)
Age of the animals: They were less than 24 h old at the beginning of the test
Acclimatization: There was no acclimatization because the water used was similar to the culture water

Study design

Test type:

static

Water media type:

other: Reconstituted water (ISO medium, according to OECD 202)

Total exposure duration:

48 h

Test conditions

Hardness:

The reconstituted water (ISO medium) had a total hardness of 247 mg/L (as CaCO3).

Test temperature:

The water temperature was measured at the start and at the end of each renewal period in each test vessel. The test temperature was in the range of 20.4 – 20.7°C measured in the test vessels.
The additionally measured temperature in the climate chamber was between 20.1 – 21.0°C.

pH:

The pH of the test solution was not adjusted and not varied by more than 1.5 units in any one test. The pH was measured at the start and at the end of each renewal period in each test vessel and was in the range of 7.65 – 7.92.

Dissolved oxygen:

The dissolved oxygen concentration was measured in each test vessel at the start and at the end of each renewal period and was in the range of 8.1 – 8.3 mg/L.

Nominal and measured concentrations:

The following nominal concentrations were tested: 0.010, 0.017, 0.031, 0.056 and 0.1 mg/L (nominal).
The Test Item was under the limit of quantification (LOQ = 0.01 mg/L) at the concentration of 0.010 mg/L at the start of the test and also at the 48 hours measurement at 0.010 and 0.017 mg/L nominal concentrations. The exposure concentrations at the 2 lowest levels were extrapolated from the measured geometric mean concentrations of the 3 highest concentration levels using the separation factor of 1.97 between them.
The corresponding calculated Test Item concentrations were: 0.0053 and 0.0103 mg/L and the measured geometric means were 0.0203, 0.0412 and 0.0785 mg/L. The biological results are based on the measured and calculated concentrations

Details on test conditions:

The test duration was 48 hours. Twenty animals, divided into four groups (glass beaker) of five animals each (~16 mL test solution/animal) were used. Taking into account that the stability of the Test Item is not available, the test was performed under static conditions. The animals were not fed during the test.
The choice of the test concentration was done on the basis of the results of a preliminary range-finding test.

Reference substance (positive control):

yes

Results and discussion

Effect concentrations

Effect concentrations 1

Key result

Duration:

48 h

Dose descriptor:

EC50

Effect conc.:

0.076 mg/L

Nominal / measured:

meas. (geom. mean)

Conc. based on:

test mat.

Basis for effect:

mobility

Effect concentrations 2

Duration:

48 h

Dose descriptor:

EC100

Effect conc.:

> 0.079 mg/L

Nominal / measured:

meas. (geom. mean)

Conc. based on:

test mat.

Basis for effect:

mobility

Effect concentrations 3

Duration:

48 h

Dose descriptor:

NOEC

Effect conc.:

0.041 mg/L

Nominal / measured:

meas. (geom. mean)

Conc. based on:

test mat.

Basis for effect:

mobility

Effect concentrations 4

Duration:

48 h

Dose descriptor:

LOEC

Effect conc.:

0.079 mg/L

Nominal / measured:

meas. (geom. mean)

Conc. based on:

test mat.

Basis for effect:

mobility

Details on results:

The following nominal concentrations were tested: 0.010, 0.017, 0.031, 0.056 and 0.1 mg/L (nominal).
The Test Item was under the limit of quantification (LOQ = 0.01 mg/L) at the concentration of 0.010 mg/L at the start of the test and also at the 48 hours measurement at 0.010 and 0.017 mg/L nominal concentrations. The exposure concentrations at the 2 lowest levels were extrapolated from the measured geometric mean concentrations of the 3 highest concentration levels using the separation factor of 1.97 between them.
The corresponding calculated Test Item concentrations were: 0.0053 and 0.0103 mg/L and the measured geometric means were 0.0203, 0.0412 and 0.0785 mg/L. The biological results are based on the measured and calculated concentrations

Reported statistics and error estimates:

DEVIATION FROM THE STUDY PLAN
The following deviation from the Study Plan was recorded during the study:
1) Only the LOQ is reported in the Analytical Report instead of the LOQ and LOD together due to technical and calculation reasons.
This deviation was considered not to have adversely affecting the results or integrity of the study.

There was no immobilized animal in the Control group and the dissolved oxygen concentration at the end of the test in Control and test vessels was more than 3 mg/L. All validity criteria were within acceptable limits and therefore the study can be considered as valid.

Any other information on results incl. tables

For the determination of the LOEC and NOEC, the immobilization at the test concentrations was tested on significant differences to the control values by Dunnett’s Test using TOXSTAT software.

Applicant's summary and conclusion

Validity criteria fulfilled:

yes

Conclusions:

EC50 (48h) daphnia magna = 0.0759 mg/L

Executive summary:

In conclusion, under the conditions of this study, the calculated endpoints for the effect of the Test Item were the following:

Parameter (48 hours)    EC50 (measured)      EC100           NOEC (measured)      LOEC (measured)

                                        0.0759 mg/L         >0.0785 mg/L     0.0412 mg/L              0.0785 mg/L

95 % conf. limits   0.0647 – 0.0981 mg/L              ---                            ---                              ---