Benzethonium chloride

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:

September - October 2001

Reliability:

1 (reliable without restriction)

Rationale for reliability incl. deficiencies:

guideline study

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)

Deviations:

no

GLP compliance:

yes (incl. QA statement)

Specific details on test material used for the study:

TEST MATERIAL
- Identity: LZ1442.3, Benzethonium chloride
- Chemical name: Benzethonium chloride
- Appearance: White powder
- Storage conditions: Room temperature
- Stability under test conditions: stable for min. 3 years from manufacturing date
- Batch No.: OC0509
- Expiry date: 7 June2002
- Purity: Responsibility of sponsor
- Date received: 20 August 2001
- Safety precautions: Routine safety precautions (lab coat, gloves, safety glasses, face mask) for unknown materials were applied to assure personnel health and safety.

Analytical monitoring:

yes

Details on sampling:

At the start of the definitive test, two samples (20 ml) were taken from the freshly-prepared control and test media. After 48 hours, the contents of the test vessels from each group were pooled and further samples were taken for analysis. On each occasion, one of the samples was analysed immediately and the other was stored in a refrigerator in case further analysis was required.

Vehicle:

no

Details on test solutions:

The test substance (10 mg) was dissolved in dilution medium (1 I) in a volumetric flask. The contents of the flask were shaken vigorously and then diluted to provide the test media. The rangefinding test used test concentrations of 0.1, 1, 10 and 100 mg/l. The definitive test concentrations, which were selected based on the results of this rangefinding test, were 0.0427, 0.0939, 0.207, 0.455 and 1 mg/l as nominal concentrations.

Test organisms (species):

Daphnia magna

Details on test organisms:

Daphnia magna (Straus) used in this study were cultured in-house and were obtained from a strain originating from the Institute National de Recherche Chimique Applique (IRChA), France. Stock cultures of Daphnia magna were maintained in glass vessels containing approximately 800 mls of Elendt M4 culture medium in a temperature-controlled laboratory at nominally 20 ± 2°C. A photoperiod of 16 hours light - 8 hours dark was maintained. The culture medium was renewed at least twice each week.Cultures were fed daily with a suspension of the unicellular green alga, Chlorella vulgaris, to provide 0.1 to 0.2 mg carbon per daphnid, per day. Culture conditions ensure that the stock animals reproduce by parthenogenesis. The day before the start ofthe study, all juvenile Daphnia were removed from the laboratory cultures. The following morning, juveniles produced by the gravid (egg-bearing) adult Daphnia were removed from the culture vessels and held in a separate holding vessel; these animals, which were less than 24 hours old, were used in the test.

Test type:

static

Water media type:

freshwater

Limit test:

no

Total exposure duration:

48 h

Post exposure observation period:

None

Hardness:

The total hardness and alkalinity of the batch ofElendt M4 medium used were 244 and 53 mg!l as CaC03, respectively.

Test temperature:

19.5-19.8°C

pH:

7.7-8.0

Dissolved oxygen:

94-100% ASV

Salinity:

Freshwater used

Nominal and measured concentrations:

The rangefmding test used test concentrations of 0.1, 1, 10 and 100 mg/l. The definitive test concentrations, which were selected based on the results of this rangefinding test, were 0.0427, 0.0939, 0.207, 0.455 and 1 mg/l as nominal concentrations. The measured concentrations ranged between 79 and 91% of their nominal values at the start of the test and between 94 and 103% of their starting values after 48 hours. The overall mean measured levels of Benzethoniumchloride were 0.0387, 0.0808, 0.164, 0.378 and 0.866 mg/l.

Details on test conditions:

A rangefinding test was conducted followed by a definitive test with five test concentrations plus one control (dilution medium). Twenty Daphnia, four replicates of five animals per vessel, were exposed in each control and test group. The first instar Daphnia were placed in groups of five, at random into glass bottles filled to capacity with 60 ml of medium to give a loading of 12 ml medium per organism. Information on the physical chemical nature of the test substance suggested that the animals would float in the test medium, to prevent this the bottles were sealed with a teflon faced butyl seal.

Reference substance (positive control):

no

Key result

Duration:

24 h

Dose descriptor:

EC50

Effect conc.:

0.249 mg/L

Nominal / measured:

meas. (arithm. mean)

Conc. based on:

test mat.

Basis for effect:

mobility

Key result

Duration:

48 h

Dose descriptor:

EC0

Effect conc.:

0.22 mg/L

Nominal / measured:

meas. (arithm. mean)

Conc. based on:

test mat.

Basis for effect:

mobility

Key result

Duration:

48 h

Dose descriptor:

NOEC

Effect conc.:

0.08 mg/L

Nominal / measured:

meas. (arithm. mean)

Conc. based on:

test mat.

Basis for effect:

mobility

Details on results:

After 48 hours, the lowest measured concentration at which 100% immobility occurred was 0.378 mg/l and the highest measured concentration at which no immobilisation was noted was 0.0808 mg/l. Based on these findings the following values have been estimated:
EC50 (24h) = 0.249 mg/l (95% confidence limits 0.164 and 0.378 mg/l)
EC50 (48h) = 0.220 mg/l (95% confidence limits 0.164 and 0.378 mg/l)

Results with reference substance (positive control):

No reference substance tested

Validity criteria fulfilled:

yes

Conclusions:

The 48-hour EC50 value was found to be 0.220 mg/l (95% confidence limits of 0.164 and 0.378 mg/l). The NOEC was 0.0808 mg/l.

Executive summary:

The acute toxicity of Benzethoniumchloride to Daphnia magna was assessed under static exposure conditions. The study was conducted in accordance with OECD Guidelines No. 202. Groups of twenty Daphnia, less than 24 hours old, were exposed for 48 hours to the test item at nominal concentrations of 0.0427, 0.0939, 0.207, 0.455 and 1 mg/l. The test media were prepared in Elendt M4 medium from an aqueous stock solution in which test substance was added directly to the dilution medium. The overall mean measured levels of Benzethoniumchloride were 0.0387, 0.0808, 0.164, 0.378 and 0.866 mg/l. After 48 hours, the lowest measured concentration resulting in 100% immobility was 0.378 mg/l and the highest measured concentration at which no immobilisation occurred was 0.0808 mg/l.

Based on these findings the 48-hour ECso value was found to be 0.220 mg/l (95% confidence limits of0.164 and 0.378 mg/l) and the NOEC to be 0.0808 mg/l.

Description of key information

The acute toxicity of Benzethoniumchloride to Daphnia magna was assessed under static exposure conditions. The study was conducted in accordance with OECD Guidelines No. 202. Based on the findings the 48-hour EC5o value was found to be 0.220 mg/l (95% confidence limits of 0.164 and 0.378 mg/l) and the NOEC to be 0.0808 mg/l.

Key value for chemical safety assessment

Fresh water invertebrates

Fresh water invertebrates

Effect concentration:

0.22 mg/L

Additional information

Source: GLP-report