4'-methoxyacetophenone

Short-term toxicity to fish

Link to relevant study record(s)

Reference

Endpoint:

short-term toxicity to fish

Type of information:

experimental study

Adequacy of study:

key study

Study period:

2017-06-26 to 2017-07-17

Reliability:

1 (reliable without restriction)

Rationale for reliability incl. deficiencies:

guideline study

Qualifier:

according to guideline

Guideline:

OECD Guideline 203 (Fish, Acute Toxicity Test)

Version / remarks:

1992

Deviations:

no

Qualifier:

according to guideline

Guideline:

EU Method C.1 (Acute Toxicity for Fish)

Version / remarks:

May 2008

Deviations:

no

GLP compliance:

yes (incl. QA statement)

Analytical monitoring:

yes

Details on sampling:

- Sampling method: Duplicate samples were taken from the test medium and the control at the start of the test, after 48 hours, and at the end of the test; taken from the approximate center of each test vessel without mixing the test medium. The concentration of the test item in the test medium and the control was determined in one of the duplicate samples from all sampling times.
- Sample storage conditions before analysis: Deep-frozen (at about -20 °C).

Vehicle:

no

Details on test solutions:

PREPARATION AND APPLICATION OF TEST SOLUTION

- Method: Immediately prior to the start of the test, the test medium of nominal 60 mg/L was prepared by completely weighing 300.04 mg of the test item into 5000 mL of test water. The test item was dissolved by ultrasonic treatment for 15 minutes and intensive stirring for 15 minutes.
- Controls: Negative control (without test item)

Test organisms (species):

Danio rerio (previous name: Brachydanio rerio)

Details on test organisms:

TEST ORGANISM
- Common name: Zebrafish
- Source: Breeding culture at IES Laboratories
- Age at study initiation: juvenile
- Length at study initiation: total body length (measured from the mouth to the end of the tail fin) of the fish was 2.4 ± 0.09 cm (Mean ± SD)
- Weight at study initiation: mean body wet weight was 0.11 ± 0.02 g (Mean ± SD)

ACCLIMATION
- Acclimation period: For at least one week.
- Acclimation conditions: Same as test.
- Type of food during acclimation: Fed with a commercial fish diet (Tetra Min® Hauptfutter, supplied by TETRA-Werke, 49324 Melle / Germany)
- Feeding frequency during acclimation: During holding and acclimatization until one day before the start of the test.
- Health during acclimation: During the last four weeks prior to the test, no fish died in the test fish batch and all fish were healthy.

FEEDING DURING TEST
The fish were not fed during the test.

Test type:

static

Water media type:

freshwater

Limit test:

yes

Total exposure duration:

96 h

Remarks on exposure duration:

Exposure duration according to the Commission Regulation (EC) No 440/2008, Part C.1 and the OECD Guideline for Testing of Chemicals No. 203 (1992).

Post exposure observation period:

Not applicable.

Hardness:

Test Water: 125 mg/L as CaCO3 (1.25 mmol/L)

Test temperature:

- Control: 22 °C
- Test item: 22 °C

pH:

- Control: 7.3 - 7.5
- Test item: 7.3 - 7.5

Dissolved oxygen:

- Control: 8.2 - 8.7 mg/L
- Test item: 8.3 - 8.8 mg/L

Salinity:

Not measured

Conductivity:

Not measured

Nominal and measured concentrations:

Nominal: 60 mg/L
Measured concentration: 59 mg/L (arithmetic mean over all measurements)

Details on test conditions:

The water temperature, pH values and oxygen concentrations were measured for each treatment at the start of the test and once daily during the test. At the same dates the appearance of the test medium was recorded. Additionally, the water temperature in the control was continuously recorded using a data logger.

TEST SYSTEM
- Test vessel: glass aquarium
- Material, size, headspace, fill volume: glass, 5 L nominal volume, 5 L fill volume
- No. of organisms per vessel: 7
- No. of vessels per concentration (replicates): 1
- No. of vessels per control (replicates): 1
- Biomass loading rate: 0.15 g fish wet weight per liter medium

TEST MEDIUM / WATER PARAMETERS
- Source/preparation of dilution water: Reconstituted test water; consisted of analytical grade salts dissolved in ultrapure water prepared in-house using an ELGA water purification system
- Ca:Mg and Na:K was 4:1 and 10:1,
- Culture medium different from test medium: Not specified
- Intervals of water quality measurement during test: daily

OTHER TEST CONDITIONS
- Adjustment of pH: No
- Photoperiod: 16-hour light to 8-hour dark photoperiod, with a 30-minute transition period
- Light intensity: 16 μE s-1 m-2.

EFFECT PARAMETERS MEASURED: Mortality

TEST CONCENTRATIONS
- Spacing factor for test concentrations: Not needed (limit test)
- Range finding study: No
- Test concentrations: 60 mg/L (nominal)

Reference substance (positive control):

no

Key result

Duration:

96 h

Dose descriptor:

LC50

Effect conc.:

> 60 mg/L

Nominal / measured:

nominal

Conc. based on:

test mat.

Basis for effect:

mortality (fish)

Details on results:

The test was considered valid as no fish in the control and the threshold concentration died during the 96-hour test period. The oxygen concentration in the test medium and the control did not drop below 60 % of the air saturation during the test.

Results with reference substance (positive control):

No positive control performed.

Reported statistics and error estimates:

The NOEC and LC0 were determined directly from the raw data. The LOEC, LC100 and LC50 at the observation times could not be quantified due to the absence of a toxic effect of the test item at the tested concentration.

Sublethal observations / clinical signs:

Table 1: Results of the HPLC/UV analytic of the test samples

Sampling Day/Sample Age	Nominal Concentration of Test Item cnom	Measured Concentration of Test Item x	Sample Preparation Factor F	Determined Concentration of Test Item c	% of Nominal Concentration
[d/h]	[mg/L]	[mg/L]		[mg/L]	[%]
0/0 fresh	Control 60	n.d. 48.8	1.2 1.2	< LOQ 58.6	n.a. 98
2/48 (aged)	Control 60	n.d. 48.9	1.2 1.2	< LOQ 58.3	n.a. 98
4/96 (aged)	Control 60	n.d. 48.6	1.2 1.2	< LOQ 58.7	n.a. 97

n.d. = no test item detected

n.a. = not applicable

LOQ: 10.5 mg/L

The tabulated values of the samples represent rounded results obtained by calculation using the exact raw data.

Table 2: Mortality and Visible Abnormalities Observed in the Test Fish

Nominal Test Item Concentration	Number of Abnormal and Dead Fish/ Number of Dead Fisch
	Observation Time
[mg/L]	2 hours	24 hours	48 hours	72 hours	96 hours
Control 60	0 / 0 0 / 0	0 / 0 0 / 0	0 / 0 0 / 0	0 / 0 0 / 0	0 / 0 0 / 0
LC50 95% C.I.	> 60 n.d.	> 60 n.d.	> 60 n.d.	> 60 n.d.	> 60 n.d.

95% C.I.: 95% confidence interval of the LC50

n.d.: Could not be determined

Validity criteria fulfilled:

yes

Conclusions:

The test item had no acute toxic effects on juvenile zebrafish (Danio rerio) at the threshold concentration of 60 mg/L (nominal).

Executive summary:

The acute toxicity of the test item to zebrafish (Danio rerio) was determined in a 96-hour test according to the Commission Regulation (EC) No 440/2008, Part C.1 and the OECD Guideline for Testing of Chemicals No. 203 (1992).

A limit test was performed to demonstrate that the test organism fish is not the most sensitive test organism to the test item compared to algae and daphnia. In this static test, zebrafish (Danio rerio) were exposed to an aqueous test medium containing the test item at the threshold concentration of nominal 60 mg/L. A control was tested in parallel. The selected test concentration of 60 mg/L corresponded approximately to the lower EC50 value determined in an algae study (OECD 201) and in an acute daphnia study (OECD 202). The lower EC50 value was determined in the algae test to be 53 mg/L, based on growth rate. In the test medium samples at the start of the test, after 48 hours and at the end of the test (96 hours) the measured test item concentrations ranged from 97 to 98 % of the nominal concentration of 60 mg/L, corresponding to a mean measured test item concentration of 59 mg/L. This shows the correct dosage of the test item and its stability during the test period of 96 hours. Therefore, all biological results are based on the nominal test item concentration of 60 mg/L. In the control and the test medium of 60 mg/L all fish survived until the end of the test and no visible abnormalities were observed at the test fish. Therefore, the 96-hour NOEC and LC0 of the test item to zebrafish were both determined to be at least 60 mg/L. The 96-hour LC50 and LC100 were clearly higher than 60 mg/L. These values could not be quantified due to the absence of toxicity of the test item at the tested concentration. In conclusion, the test item had no acute toxic effects on juvenile zebrafish (Danio rerio) at the threshold concentration of 60 mg/L. According to the threshold approach, it was demonstrated that the test organism fish is not the most sensitive species for the test item compared to daphnia and algae and that the 96-hour LC50 for fish is greater than the threshold concentration of 60 mg/L.

Description of key information

The acute toxicity of the test substance to Zebrafish (Danio rerio) was determined according to OECD 203. The 96 hour LC50 was determined to be >60 mg/L based on nominal concentration.

Key value for chemical safety assessment

Fresh water fish

Fresh water fish

Effect concentration:

60 mg/L

Additional information

The acute toxicity of the test item to zebrafish (Danio rerio) was determined in a 96-hour limit test under GLP-conditions, according to the Commission Regulation (EC) No 440/2008, Part C.1 and the OECD Guideline for Testing of Chemicals No. 203 (1992). In this static test, zebrafish (Danio rerio) were exposed to an aqueous test medium containing the test item at the threshold concentration of nominal 60 mg/L. The 96-hour NOEC of the test item to zebrafish was determined to be at least 60 mg/L. The 96-hour LC50 and LC100 were clearly higher than 60 mg/L. In conclusion, the test item had no acute toxic effects on juvenile zebrafish (Danio rerio) at the threshold concentration of 60 mg/L. According to the threshold approach, it was demonstrated that the test organism fish is not the most sensitive species for the test item compared to daphnia and algae and that the 96-hour LC50 for fish is greater than the threshold concentration of 60 mg/L.