Acetylglucosaminidase, β-

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:

20 July 2020 to March 11, 2021

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)

Version / remarks:

Please state version

Deviations:

no

GLP compliance:

yes

Analytical monitoring:

yes

Remarks:

Only the highest concentration could be measured, as the other concentrations fell below the assay detection limit. Therefore, nominal concentrations were used.

Details on sampling:

- Concentrations: 260 mg enzyme contrate dry matter/L (equivalent to 100 mg active enzyme protein/L)
- Sample storage conditions before analysis: During the duration of the test, the samples were treated the same as the test concentrations. Thereafter, the samples were stored frozen until analysis.

Vehicle:

no

Details on test solutions:

PREPARATION AND APPLICATION OF TEST SOLUTION
- Method: Dilution in water
- Controls: Elendt M4 medium
- Evidence of undissolved material (e.g. precipitate, surface film, etc.): No

Test organisms (species):

Daphnia magna

Details on test organisms:

Common name: Daphnia magna
Strain: Straus
Source: In-house laboratory cultures. A clone of this species was originally supplied by the University of Sheffield, UK.
Age at study initiation: Gravid adults were isolated 24 hours prior to initiation of the test. Young daphnids produced overnight were used for testing.
- Feeding during test: No

Test type:

static

Water media type:

freshwater

Limit test:

no

Total exposure duration:

48 h

Hardness:

250 mg/L CaCO3

Test temperature:

21-22°C

pH:

8.0-8.2 in control
7.3-8.2 in test concentrations

Dissolved oxygen:

8.6-8.8 mg O2/L in controls
6.1-8.8 mg O2/L

Nominal and measured concentrations:

16.3, 32.6, 65.2, 130.4 and 260.7 mg enzyme concentrate dry matter/L (nominal concentrations)
The sample 260.7 mg enzyme concentrate dry matter/L in Elendt M4 medium at 0 hours achieved 90.2% recovery, but fell below the limit of detection, and therefore, cannot be supported as a result. The respective sample in reverse osmosis water attained a recovery of 96.4%. The 48-hour old samples fell below the detection limit, which is close to the highest concentration.

Details on test conditions:

TEST SYSTEM
- Test vessels: 150 mL glass vessels each containing approximately 100 mL test solution.
- Experimental design: 20 animals per test concentration. 5 test concentrations plus 1 control. Duplicate samples were prepared for enzyme activity analysis and stored in the test concentrations.
- Type: covered to reduce evaporation
- No. of organisms per vessel: 5
- No. of vessels per concentration (replicates): 4
- No. of vessels per control (replicates): 4

TEST MEDIUM / WATER PARAMETERS
- Source/preparation of dilution water: Elendt M4 medium. Total hardness 250 mg/L as CaCO3.
- Culture medium different from test medium: no

OTHER TEST CONDITIONS
Method of initiation: Daphnia were placed in the vessels containing the test preparations.
Loading: 20 mL test solution per organism.
Photoperiod: 16 h light: 8 h dark with with 20 minute dawn and dusk transition periods.
Medium renewal: None.
Duration of exposure: 48 hours.
Criterion of effect: Daphnia were considered to be immobilized if they were unable to swim for approximately 15 seconds after gentle agitation.

EFFECT PARAMETERS MEASURED: The numbers of mobile, immobile and floating daphnia were counted approximately 24 and 48h after the start of the study.
TEST CONCENTRATIONS
16.3, 32.6, 65.2, 130.4 and 260.7 mg enzyme concentrate dry matter/L.

Reference substance (positive control):

no

Key result

Duration:

48 h

Dose descriptor:

EC50

Effect conc.:

> 260.7 mg/L

Nominal / measured:

nominal

Conc. based on:

other: enzyme concentrate dry matter

Basis for effect:

mobility

Duration:

48 h

Dose descriptor:

NOEC

Effect conc.:

260.7 mg/L

Nominal / measured:

nominal

Conc. based on:

other: enzyme concentrate dry matter

Basis for effect:

mobility

Details on results:

Details on results
- Mortality of control: 0
- Other adverse effects control: No
- Abnormal responses: No
- Effect concentrations exceeding solubility of substance in test medium: No

Validity criteria fulfilled:

yes

Conclusions:

Exposure of Daphnia magna to beta-N-acetylhexosaminidase based on nominal concentrations gave EC50 value of greater than 260.7 mg enzyme concentrate dry matter/L (equivalent to 100 mg active enzyme protein/L). The No Observed Effect Concentration (NOEC) was the highest concentration tested, 260.7 mg enzyme concentrate dry matter/L (equivalent to 100 mg active enzyme protein/L).

Executive summary:

A study was performed to assess the acute toxicity of beta-N-acetylhexosaminidase to Daphnia magna. The method followed was designed to be compatible with the OECD Guidelines for Testing of Chemicals (April 2004) No 202, "Daphnia sp., Acute Immobilisation Test" referenced as Method C.2 of Commission Regulation (EC) No. 440/2008.

20 daphnids (4 replicates of 5 animals) were exposed to aqueous dispersions of the test item at nominal concentrations of 16.3, 32.6, 65.2, 130.4 and 260.7 mg enzyme concentrate dry matter/L for 48 hours at a temperature of 21 °C to 22 °C under static test conditions. The number of immobilized Daphnia was recorded after 24 and 48 hours.

Analysis of the test preparations at 0 and 48 hours were performed by the Sponsor. Only the highest concentration could be measured, as the other concentrations fell below the assay detection limit. Therefore, nominal concentrations were used.

Exposure of Daphnia magna to beta-N-acetylhexosaminidase based on nominal concentrations gave EC50 value of greater than 260.7 mg enzyme concentrate dry matter/L (equivalent to 100 mg active enzyme protein/L). The No Observed Effect Concentration (NOEC) was the highest concentration tested, 260.7 mg enzyme concentrate dry matter/L (equivalent to 100 mg active enzyme protein/L).

Description of key information

Exposure of Daphnia magna to beta-N-acetylhexosaminidase based on nominal concentrations gave EC50 value of greater than 260.7 mg enzyme concentrate dry matter/L (equivalent to 100 mg aep/L). The No Observed Effect Concentration (NOEC) was the highest concentration tested, 260.7 mg enzyme concentrate dry matter/L (equivalent to 100 mg aep/L).

Key value for chemical safety assessment

Fresh water invertebrates

Fresh water invertebrates

Dose descriptor:

EC50

Effect concentration:

> 260 mg/L