4-ethylphenol

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:

2004-09-28 to 2005-06-07

Reliability:

1 (reliable without restriction)

Rationale for reliability incl. deficiencies:

guideline study

Data source

Reference

Reference Type:

study report

Title:

Unnamed

Year:

2005

Report date:

2005

Materials and methods

Test guideline

Qualifier:

according to guideline

Guideline:

OECD Guideline 202 (Daphnia sp. Acute Immobilisation Test)

Deviations:

no

GLP compliance:

yes (incl. QA statement)

Test material

Test material information

Constituent 1

Reference substance name:

2-ethylphenol

EC Number:

201-958-4

EC Name:

2-ethylphenol

Cas Number:

90-00-6

Molecular formula:

C8H10O

IUPAC Name:

2-ethylphenol

Constituent 2

Reference substance name:

4-ethylphenol

EC Number:

204-598-6

EC Name:

4-ethylphenol

Cas Number:

123-07-9

Molecular formula:

C8H10O

IUPAC Name:

4-ethylphenol

Constituent 3

Reference substance name:

3-ethylphenol

EC Number:

210-627-3

EC Name:

3-ethylphenol

Cas Number:

620-17-7

Molecular formula:

C8H10O

IUPAC Name:

3-ethylphenol

Test material form:

not specified

Sampling and analysis

Analytical monitoring:

yes

Details on sampling:

At test initiation (0 hour) and test termination ( 48 hours), one sample from each test solution and control was analyzed for ethyl phenols concentration. At test initiation, test solution samples were taken from the intermediate vessels prior to division into replicate vessels. At test termination, the replicate solutions were composited prior to sampling. Each sample was collected by pipet from the approximate midpoint of the test vessel.
Three quality control (QC) samples were also prepared at each sampling interval at nominal ethyl phenols concentrations of 1.00 to 50.0 mg/L and remained with the exposure solution samples throughout the analytical process. Analysis of the QC samples was used to judge the precision and quality control maintained during the analytical process.

Test solutions

Vehicle:

yes

Details on test solutions:

The dilution water source used during the definitive test was from the same source as the water in the daphnid cultures and was characterized as having a total hardness and a total alkalinity as CaCO3 of 190 mg/Land 120 mg/L, respectively, a specific conductivity of 500 µmhos/cm, a pH of 8.0, and a dissolved oxygen concentration of 9.4 mg/L.
Prior to test initiation, a 300 mg/mL stock solution was prepared by placing 7.4 mL (7.5332 g based on a specific gravity of 1 .018 g/mL) of ethyl phenols in a 25-mL volumetric flask and diluting to volume with acetone (CAS No. 67-64-1 ). The resultant 300 mg/mL stock solution was observed to be clear and colorless with no visible sign of undissolved test substance. The 300 mg/mL stock was used to prepare secondary stock solutions (150, 75. 38 and 19 mg/mL). All resulting stock solutions were observed to be clear and colorless with no visible undissolved test substance, and were used to prepare exposure solutions .
A solvent control solution was prepared by adding 0.10 mL of acetone to 1.0 L of dilution water. Each test solution was divided into four replicate capped vessels, each containing 200 ml of
solution. Test vessels were fitted with Teflon-lined screw-top caps to minimize the potential for volatilization of the test substance.
In addition, four control vessels containing dilution water were established and maintained under the same conditions as the treatment level solutions.

Test organisms

Test organisms (species):

Daphnia magna

Details on test organisms:

The Daphnia magna used in this toxicity test were obtained from laboratory cultures maintained at Springborn Smithers. The adult daphnids used to produce offspring for this test ( 1) did not contain ephippia, (2) produced offspring prior to being 12 days old and (3) less than 20% mortality was observed in the culture population in the 48 hours prior to test initiation. The culture water was prepared by fortifying well water based on the formula for hard water (U.S. EPA, 1975) and filtering it through an Amberlite XAD-7 resin column to remove any potential organic contaminants.
For the two weeks prior to testing, this water had total hardness and total alkalinity ranges as calcium carbonate (CaCO3) of 180 to 190 mg/L and 120 to 130 mg/L, respectively and a specific conductivity range of 500 to 600 micromhos per centimeter (µmhos/cm). A pH range of 7.6 to 9.6, a temperature range of 18 to 20 °C, and dissolved oxygen concentration range of 6.8 to 12.0 mg/L were measured in the culture solutions during the two weeks prior to test initiation.
The daphnid culture area received a regulated photoperiod of 16 hours of light and 8 hours of darkness. Light at an intensity of 112 to 140 footcandles at the surface of the culture solutions was provided by fluorescent bulbs. Daphnids were fed unicellular green algae, Ankistrodesmus falcatus (4 x 10^7 cells/mL), in addition to a suspension of YCT (yeast, cereal leaves and flaked fish food).

Study design

Test type:

static

Water media type:

freshwater

Limit test:

no

Total exposure duration:

48 h

Test conditions

Hardness:

Total hardness as CaCO3: 190 mg/L

Test temperature:

19 to 20 °C

pH:

at test start: 8.4
at test end: 8.2 - 8.4

Dissolved oxygen:

at test start: 9.5 - 9.6 mg/L
at test end: 8.8 - 8.9 mg/L

Conductivity:

Specific conductivity: 500 µmhos/cm

Nominal and measured concentrations:

nominal: 1.9, 3.8, 7.5, 15 and 30 mg/L
measured: 1.1, 2.4, 6.2, 12 and 27 mg/L

Details on test conditions:

Due to the volatile nature of the test substance. the following procedures were used to minimize the loss of test substance: 1.) the test substance was transferred volumetrically rather than weighed, 2.) 250-mL screw-top exposure vessels were used rather than traditional 1.6-L battery jars which allow air exchange, and 3.) water quality measurements were not performed at 24 hours of exposure to limit the exposure of the test substance to air.
The test vessels were impartially placed in a temperature-controlled water bath designed to maintain exposure solution temperatures at 20 ± 1 °C. Test vessels were 250-mL glass flasks with screw-top Teflon-lined caps, each containing 200 mL of test solution. Test vessels were fitted with screw-top caps to minimize the potential for volatilization of the test substance and to limit exposure to the strong odor of the test solutions. The test area was illuminated with fluorescent bulbs at an intensity of 63 to 78 footcandles at the solutions' surface. The photoperiod during the test was the same as in the culture area. Sudden transitions from light to dark and vice versa were avoided. Light intensity was measured with an Extech 401025 light meter.
The test was initiated when five daphnids, < 24 hours old, were impartially selected and distributed to each vessel (replicates A, B, C and D). Daphnids were impartially added to an intermediate vessel, one daphnid at a time. until each vessel contained five daphnids. The test was initiated when the daphnids from each intermediate vessel were impartially added to each respective test vessel. Daphnids were not fed during the exposure.
The number of immobilized daphnids in each replicate test vessel was recorded at 24 and 48 hours of exposure. Immobilization was defined as those animals not able to swim within
15 seconds after gentle agitation of the test vessel. Biological observations and observations of the physical characteristics of each replicate test solution were also made and recorded at 0, 24 and 48 hours. The pH, dissolved oxygen concentration and temperature were measured at 0, and 48 hours in each treatment level and the controls.

Results and discussion

Effect concentrations

Effect concentrations 1

Duration:

48 h

Dose descriptor:

NOEC

Effect conc.:

2.4 mg/L

Nominal / measured:

meas. (geom. mean)

Conc. based on:

test mat.

Basis for effect:

mobility

Effect concentrations 2

Key result

Duration:

48 h

Dose descriptor:

EC50

Effect conc.:

9 mg/L

Nominal / measured:

meas. (geom. mean)

Conc. based on:

test mat.

Basis for effect:

mobility

Remarks on result:

other: 95% confidence (6.2 - 12 mg/L)

Reported statistics and error estimates:

If at least one test concentration caused immobilization of greater than or equal to 50% of the test population, then a computer program, TOXSTAT® (Gulley. et al., 1996), was used to calculate the EC50 values and 95% confidence intervals. The No-Observed-Effect Concentration (NOEC) during the 48-hour exposure period was also determined.

Any other information on results incl. tables

Table 1: Mean measured concentrations tested and corresponding cumulative percent and number of immobilized organisms made during the 48-hour static acute exposure of daphnids (Daphnia magna) to ethyl phenols (actual number of immobilized daphnids is presented in parentheses). 

	Cumulative Percent Immobilization (%)
	24-Hour					48-Hour
Mean Measured Concentration (mg/L)	A	B	C	D	Mean	A	B	C	D	Mean
Control	0	0	0	0	0	0	0	0	0	0
Solvent Control	0	0	0	0	0	0	0	0	0	0
1.1	0	0	0	0	0	0	0	0	0	0
2.4	0	0	0	0	0	0	0	0	0	0
6.2	0	0	0	0	0	0	0	0	0	0
12	40 (2)	60 (3)	60 (3)	100 (5)	65	100 (5)	80 (4)	100 (5)	100 (5)	95
27	100 (5)	100 (5)	100 (5)	100 (5)	100	100 (5)	100 (5)	100 (5)	100 (5)	100

Applicant's summary and conclusion

Validity criteria fulfilled:

yes

Conclusions:

The 48h EC50 value for ethyl phenols was determined to be 9.0 mg/L, the NOEC was determined to be 2.4 mg/L.

Executive summary:

The purpose of this study was to estimate the acute toxicity (EC50) of ethyl phenols to daphnids (Daphnia magna) under static test conditions.

Based on the results of this test, the 48-hour EC50 value for ethyl phenols and Daphnia magna was determined to be 9.0 mg/L, with 95% confidence intervals of 6.2 to 12 mg/L. The No-Observed-Effect Concentration (NOEC) was determined to be 2.4 mg/L.