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Ecotoxicological information

Short-term toxicity to fish

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Administrative data

Endpoint:
short-term toxicity to fish
Type of information:
experimental study
Adequacy of study:
key study
Reliability:
1 (reliable without restriction)
Rationale for reliability incl. deficiencies:
other: study design is comparable to OECD guideline 203 and the study is very well documented, analytical monitoring of test substance concentration conducted, validity criteria fulfilled

Data source

Reference
Reference Type:
study report
Title:
Unnamed
Year:
1984

Materials and methods

Test guideline
Qualifier:
equivalent or similar to guideline
Guideline:
OECD Guideline 203 (Fish, Acute Toxicity Test)
GLP compliance:
not specified

Test material

Constituent 1
Chemical structure
Reference substance name:
Benzophenone
EC Number:
204-337-6
EC Name:
Benzophenone
Cas Number:
119-61-9
Molecular formula:
C13H10O
IUPAC Name:
benzophenone
Details on test material:
- Name of test material (as cited in study report): benzophenone
- Substance type: aromatic ketone
- Physical state: solid (25.3°C; melting point: ca. 48°C)
- Analytical purity: 99% (obtained from Aldrich Chemical Co.)
- Impurities (identity and concentrations): not reported
- Stability under test conditions: not reported
- Storage condition of test material: not reported

Sampling and analysis

Analytical monitoring:
yes
Details on sampling:
- Concentrations:
Test 1: 0-21.9 mg/L nominal (0- 23.8 mg/L effective) were tested
Test 2: 0-ca. 22.3 mg/L nominal (0- 23.8 mg/L effective) were tested
- Sampling method: sampling was performed at approximately mid-depth at 0 and 96 h and one of each duplicate exposure chambers at 24, 48, and 72 h. The test substance concentration was measured using gas liquid chromatography (GLC). GLC analyses was performed by either direct aqueous injection or solvent extraction and subsequent analysis.
- Sample storage conditions before analysis: all samples were analyzed immediately or adequately preserved for later analysis.

Test solutions

Vehicle:
no
Details on test solutions:
PREPARATION AND APPLICATION OF TEST SOLUTION (especially for difficult test substances)
- Method: Flow-through exposure was conducted using the cycling proportional diluter (acc. to Mount and Brungs 1967) with duplicate exposures for every tested concentration. In the case of acetophenone a diluted stock solution was prepared; the diluted stock was a premixed aqueous solution of desired concentration (4100 mg/L).
- Differential loading: volume of exposure tank: 6.3 L; additions: 5.49 V/d
- Controls: 2 controls (test substance omitted) performed.

Test organisms

Test organisms (species):
Pimephales promelas
Details on test organisms:
TEST ORGANISM
- Common name: fathead minnow (Pimephales promelas)
- Source: US EPA Environmental Research Laboratory-Duluth, MN
- Age at study initiation (mean and range, SD):
Test 1: 31 days
Test 2: 30 days
- Length at study initiation (length definition, mean, range and SD):
Test1: mean length: 21.8 mm (SD: 1.640 mm)
Test2: mean length: 19.7 mm (SD: 1.725 mm)
- Weight at study initiation (mean and range, SD): mean weight:
Test1: 0.165 g (SD: 0.0395 g)
Test2: 0.091 g (SD: 0.0126 g)
- Method of breeding: Fathead minnows used in the tests were cultured from brood stock provided by US EPA Environmental Research Laboratory-Duluth, MN. Adults were held at 25°C in flowing water with a 16 h light-controlled photoperiod and fed frozen adult brine shrimp (Artemia sp.). They were provided with asbestos pipe (cut in half longitudinally) as spawning substrate. Eggs were spawned and fertilized on the undersides of these substrates. The embryos were tended by the male parent until eye-up at which time the substrates with embryos attached were removed to another 25°C bath where hatching occurred. Fry of similar (within 24 hr) age were transferred to flow-through rearing tanks at 25°C with a 16 h light photoperiod. Fry were fed freshly hatched brine shrimp nauplii to excess three times daily until 24 h before a test (twice on most week-end days) and a dry flake ration (Tetramin) during their first week. Fish approx. 30 days old were then used in the toxicity tests.
- Feeding during test: no

ACCLIMATION
- Acclimation period: ca. 30 d
- Acclimation conditions (same as test or not): temperature: 25°C; photoperiod: 16 h light/8 h dark; flow-through conditions
- Type and amount of food: fry were fed freshly hatched brine shrimp nauplii to excess three times daily until 24 h before a test (twice on most week-end days) and a dry flake ration (Tetramin) during their first week.
- Feeding frequency: fry were fed freshly hatched brine shrimp nauplii to excess three times daily until 24 h before a test (twice on most week-end days) and a dry flake ration (Tetramin) during their first week. No feeding during the test performance.
- Health during acclimation (any mortality observed): Only groups of fish having healthy appearance and no history of unusual thermal exposure or abnormally high mortality rate were used for toxicant exposure.
- other: Prior to exposure fish were pooled if reared in more than one tank, and by inspection unusually large or small individuals were netted out with minimal disturbance of the remainder fish.

Study design

Test type:
flow-through
Water media type:
freshwater
Total exposure duration:
96 h

Test conditions

Hardness:
Test 1: 47.9 mg/L CaCO3
Test 2: 43.7 mg/L CaCO3
Test temperature:
Test 1: 25.3°C
Test 2: 25.4°C
pH:
Test 1: 7.72
Test 2: 7.46
Dissolved oxygen:
Test 1: 6.9 mg/L
Test 2: 7.46 mg/L
Nominal and measured concentrations:
Test 1:
Test concentrations (mg/L)

nominal 0 (control) 2.8 4.7 7.9 13.1 21.9
measured <0.24 2.84/2.85 4.44/4.12 7.24/7.40 11.8/12.0 23.4/23.8 (corrected average values; tests conducted in duplicate)

Test 2:
Test concentrations (mg/L)

nominal 0 (control) 2.89 4.82 8.03 13.4 22.3
measured <0.25 2.34/2.03 3.97/4.24 7.20/7.37 11.6/12.0 22.4/24.4 (corrected average values; tests conducted in duplicate)
Details on test conditions:
TEST SYSTEM
- Test vessel: tank/container (6.3 L)
- Type (delete if not applicable): open
- Material, size, headspace, fill volume: size: 6.3 L (no further data)
- Aeration: not reported
- Type of flow-through (e.g. peristaltic or proportional diluter): proportional diluter according to Mount and Brungs 1967
- Renewal rate of test solution (frequency/flow rate): additions: 5.49 volume/d (corresponding to 34.59 L/d)
- No. of organisms per vessel: 25
- No. of vessels per concentration (replicates): 2
- No. of vessels per control (replicates): 2
- Biomass loading rate: 0.155 g/L


TEST MEDIUM / WATER PARAMETERS
- Source/preparation of dilution water: Lake Superior Water
- Total organic carbon: not reported
- Particulate matter: <1 mg/L (total non-filterable residue)
- Metals: sodium: 1.1-1.2 mg/L, calcium: 13.0-14.7 mg/L, magnesium: 2.9-3.6 mg/L, potassium: 0.48-0.59 mg/l, aluminium: 1-26 µg/L, cadminum<0.1 µg/L, chromium: 2-20 µg/L, cobalt<0.5 µg/L, copper: 0.3-3.2 µg/L, iron: 2-83 µg/L; lead: 7-20 µg/L, manganese: 0.2-11.5 µg/L, nickel<0.5 µg/L, and zinc: 1.0-2.7µg/L
- Pesticides: -
- Chlorine: 1.2-1.3 mg/L
- Alkalinity: 35 mg/L CaCO3
- Ca/Mg ratio: 3
- Conductivity: 78-86 µmhos/cm (specific conductance)
- Culture medium different from test medium: no
- Intervals of water quality measurement:
a. temperature was measured daily
b. dissolved oxygen was measured at least three times (0, 24, 96 h) in high, medium, low and control exposure chambers
c. total hardness was determined at least once for each test
d. total alkalinity was determined at least once for each test by titrimetric method acc. to APHA 1980
e. pH was measured at least once during each test in high, medium, low and control exposure chambers
f. acidity was determined for all tests in high, medium, low and control exposure chambers

OTHER TEST CONDITIONS
- Adjustment of pH: no
- Photoperiod: 16 h light/8 h dark
- Light intensity: not reported


EFFECT PARAMETERS MEASURED (with observation intervals if applicable) :
-mortality: dead fish were removed after 3, 5, 12, 24, 48, 72, and 96 h
-behavior: routinely observed during exposure

TEST CONCENTRATIONS
- Spacing factor for test concentrations: 1.66
- Range finding study: not reported

Results and discussion

Effect concentrationsopen allclose all
Duration:
96 h
Dose descriptor:
LC50
Effect conc.:
15.3 mg/L
Nominal / measured:
meas. (not specified)
Conc. based on:
test mat.
Basis for effect:
mortality (fish)
Remarks on result:
other: Test 1 95% CL: 14.4-16.3 mg/L
Duration:
96 h
Dose descriptor:
EC50
Effect conc.:
15.3 mg/L
Nominal / measured:
meas. (not specified)
Conc. based on:
test mat.
Basis for effect:
behaviour
Remarks on result:
other: Test 1 95% CL: 14.4-16.3 mg/L
Duration:
96 h
Dose descriptor:
LC50
Effect conc.:
14.2 mg/L
Nominal / measured:
meas. (not specified)
Conc. based on:
test mat.
Basis for effect:
mortality (fish)
Remarks on result:
other: Test 2 95% CL: 13.2-15.3 mg/L
Duration:
96 h
Dose descriptor:
EC50
Effect conc.:
14.2 mg/L
Nominal / measured:
meas. (not specified)
Conc. based on:
test mat.
Basis for effect:
behaviour
Remarks on result:
other: Test 2 95% CL: 13.2-15.3 mg/L
Details on results:
- Behavioural abnormalities:
Test 1: Many of the fish in the highest concentration that did not lose equilibrium, swam in a slow corkscrew fashion, with head down after 13 hours. Later, they were spread out and motionless on the bottom of the tank and were hypoactive.

Test 2: Immediate stress to fish when added to the two highest concentrations. Fish in the second highest concentration moved in slow, head down spirals after 24 hours. Fish in the middle concentration were spread out on the bottom of the tank. After 72 hours, fish in the second highest concentration were motionless on the bottom of the tank.
Reported statistics and error estimates:
Estimation of the LC50 and EC50 was made using the 'trimmed Spearman-Karber' method (Hamilton et al. 1977).

Any other information on results incl. tables

Sublethal observations / clinical signs:

The following results were obtained in Test 1:

a. mortality (25 fish each were used)

Time (h)

control

A

B

C

D

E

1

-

-

-

-

-

-

 

-

-

-

3

6

2

-

-

-

-

-

-

-

-

2

-

17

25

3

-

-

-

-

-

-

-

-

4

-

24

25

6

-

-

-

-

-

-

-

-

5

1

24

25

13

-

-

-

-

-

-

-

-

5

1

24

25

22

-

-

-

-

-

-

-

-

6

2

24

25

24

-

-

-

-

-

-

-

-

6

2

24

25

48

-

-

-

-

-

-

-

-

6

2

24

25

72

-

-

-

-

-

-

-

-

6

2

24

25

96

-

-

-

-

-

-

-

-

6

2

24

25

Control: < 0.24 mg/L benzophenone

A: 2.84-2.85 mg/L benzophenone

B: 4.44-4.12 mg/L benzophenone

C: 7.24-7.40 mg/L benzophenone

D: 11.8-12.0 mg/L benzophenone

E: 23.4-23.8 mg/L benzophenone

Based on the results a 96 h LC50=15.3 mg/l was obtained (confidence limits: 14.4 – 16.3)

 

b.effect (25 fish each were used)

Time (h)

control

A

B

C

D

E

1

-

-

-

-

-

-

 

-

4

1

24

25

2

-

-

-

-

-

-

-

-

5

1

24

25

3

-

-

-

-

-

-

-

-

6

1

24

25

6

-

-

-

-

-

-

-

-

6

1

24

25

13

-

-

-

-

-

-

-

-

7

3

24

25

22

 

 

 

 

 

 

 

 

7

 

24

25

24

-

-

-

-

-

-

-

-

7

3

24

25

48

-

-

-

-

-

-

-

-

6

3

24

25

72

-

-

-

-

-

-

-

-

6

2

24

25

96

-

-

-

-

-

-

-

-

6

2

24

25

Concentrations tested see above

Based on the results a 96 h EC50=15.3 mg/l was obtained (confidence limits:14.4 – 16.3)

 

The following results were obtained in Test 2:

a. mortality (25 fish each were used)

Time (h)

control

A

B

C

D

E

1

-

-

-

-

-

-

1

0

0

1

8

1

2

-

-

-

-

-

-

1

0

0

2

24

22

3

-

-

-

-

-

-

1

0

0

2

25

25

6

-

-

-

-

-

-

1

0

3

3

25

25

13

-

-

-

-

-

-

1

0

4

4

25

25

24

-

-

-

-

-

-

1

0

4

4

25

25

48

-

-

-

-

-

-

1

0

5

5

25

25

72

-

-

-

-

-

-

1

0

5

5

25

25

96

-

-

-

-

-

-

1

0

6

6

25

25

Control: <0.25 mg/L benzophenone

A: 2.34-2.03 mg/L benzophenone

B: 3.97-4.24 mg/L benzophenone

C: 7.20-7.37mg/L benzophenone

D: 11.6-12.0 mg/L benzophenone

E: 22.4-24.4 mg/L benzophenone

Based on the results a 96 h LC50=14.2 mg/l was obtained (confidence limits: 13.2 – 15.3)

 

b.effect (25 fish each were used)

Time (h)

control

A

B

C

D

E

1

-

-

-

-

-

-

1

0

8

11

25

25

2

-

-

-

-

-

-

2

0

3

7

25

25

3

-

-

-

-

-

-

1

0

3

5

25

25

6

-

-

-

-

-

-

1

0

5

5

25

25

13

-

-

-

-

-

-

1

0

6

5

25

25

24

-

-

-

-

-

-

1

0

9

7

25

25

48

-

-

-

-

-

-

1

0

6

5

25

25

72

-

-

-

-

-

-

1

0

6

5

25

25

96

-

-

-

-

-

-

1

0

6

6

25

25

Concentrations tested see above

Based on the results a 96 h LC50=14.2 mg/l was obtained (confidence limits: 13.2 – 15.3)

Applicant's summary and conclusion

Validity criteria fulfilled:
yes
Conclusions:
In two flow-through tests (comparable to OECD guideline 203) acetophenone shows acute toxicity towards Pimephales promelas:
Test 1
-mortality: 96 h LC50=15.3 mg/L
-behavior: 96 h EC50=15.3 mg/L

Test 2
-mortality: 96 h LC50=14.2 mg/L
-behavior: 96 h EC50=14.2 mg/L

Executive summary:

The acute toxicity to fish was investigated under flow-through conditions and the study was conducted comparable to OECD guideline 203 'Fish, acute toxicity'. In two separate tests total 6 concentrations inclusive the controls were tested in duplicate each. The applied nominal concentrations ranged in test 1 from 0 to 21.9 mg/L (<0.24 to 23.8 mg/L effective), in test 2 from 0 to 21.9 mg/L (<0.24 to 23.8 mg/L effective), respectively. 50 Fish were used per concentration step. Lake Superior Water was used as dilution water. The actual concentration was measured using GLC. Benzophenone shows toxicity towards Pimephales promelas: 96 h LC50 and EC50=15.3 mg/L (test 1), 96 h LC50 and EC50=14.2 mg/L (test 2). The effect concentration of the two tests is close together. The mean effect concentration is: 96 h LC50 and EC50=14.75 mg/L

The study is very well documented and can be classified as reliable without restrictions satisfying the guideline requirements for an acute toxicity study with fish.

Results Synopsis

Test1:

Test organism size/age: Pimephales promelas(30 d old, length: 21.8 mm; weight: 0.165 g)

Test Type: Flow-through

96 h LC50: 15.3 mg/L (measured)

96 h EC50: 15.3 mg/L (measured)

Endpoint(s) Effected: mortality, behaviour

Test2:

Test organism size/age: Pimephales promelas(31 d old, length: 19.7 mm; weight: 0.091 g)

Test Type: Flow-through

96 h LC50: 15.3 mg/L (measured)

96 h EC50: 15.3 mg/L (measured)

Endpoint(s) Effected: mortality, behaviour

The mean effect concentration of Test 1 and Test 2: 96 h LC50 and EC50=14.75 mg/L

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