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Short-term toxicity to fish

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short-term toxicity to fish
Type of information:
experimental study
Adequacy of study:
key study
1 (reliable without restriction)
Rationale for reliability incl. deficiencies:
other: GLP, guideline study with measured concentrations of the test substance.
Reason / purpose for cross-reference:
reference to other study
according to guideline
OECD Guideline 203 (Fish, Acute Toxicity Test)
Principles of method if other than guideline:
ISO International Standard 7346-1: "Water quality -
Determination of the acute lethal toxicity of substances to
a freshwater fish [Brachydanio rerio Hamilton-Buchanan
(Teleostei, Cyprinidae)]" - Part 2: Semi-static method,
Second edition, 1996-06-15.

European Economic Community (EEC), EEC directive 92/69, Part
C: Methods for the determination of ecotoxicity,
Publication No. L383, December 1992, C.1. "Acute toxicity
for fish".

OECD guidelines for Testing of Chemicals, guideline No. 203:
"Fish Acute Toxicity Test", Adopted 17 July, 1992

Guidance document on aquatic toxicity testing of difficult
substances and mixtures, OECD series on testing and
assessment number 23, December 14, 2000.
GLP compliance:
Analytical monitoring:
Details on sampling:
During the final test singular samples were taken from all test concentrations and the blank control at the start of exposure (fresh) and after 24 hours (spent). In addition, samples were taken after 48 hours from the spent solution containing 56% filtrate. After 72 and 96 hours of exposure samples were takend from the three lowest test concentrations and the blank control, respectively fresh and spent solutions. 2 ml samples were removed from the center of the test vessels at each sampling event and were analyzed on the day of sampling.
Details on test solutions:
Identity and concentration of auxiliary solvent for dispersal: Preparation of test solutions started with a solution of 100
mg/l applying a short mixing period in order to reduce the
expected degradation due to reaction with oxygen. A total of
5 minutes magnetic stirring resulted in a clear solution
with a few floating particles and negligible precipitate of
unknown origin. As a result, the solution was coarsely
filtered through a paper filter (Schleicher and Schuell 604)
to remove any larger (> ca. 5 µm) undissolved particles.
The lower test concentrations were prepared by subsequent
dilutions of the filtrate in test medium. The final test
solutions were all clear and colourless.

Part of the test solutions was used for the simultaneously
performed acute Daphnia magna toxicity test.
Test organisms (species):
Cyprinus carpio
Details on test organisms:
Source: Zodiac, proefacc, "De Haar Vissen", Wageningen University and Research Centre, the Netherlands
Mean length = Range-finding test: 2.8 +/- 0.2 cm, final test: 2.6 +/- 0.1 cm
Mean weight = Range-finding test: 0.62 +/- 0.11 g, final test: 0.52 +/- 0.11 g
Total fish used =54
Fish were held for 12 days after delivery in ISO medium.
Water quality parameters were kept within the optimum limits for the respective fish species.
Fish were fed daily with pelleted fish food and/or artemia until 24 hours before the test is started.
In the batch of fish used for the test, mortality during the seven days prior to the start of the test was less than 5%.
Test type:
Water media type:
Limit test:
Total exposure duration:
96 h
Post exposure observation period:
Not applicable
250 mg CaCO3/L
Test temperature:
21.1 - 22.2 throughout the test
7.3 - 8.0 throughout the test
Dissolved oxygen:
>60% saturation throughout the test
Not applicable
Nominal and measured concentrations:
Nominal concentrations: 0, 10, 18, 32, and 56% of the filtrate
Measured concentrations were 91 - 105% of nominal.
Details on test conditions:
- Test vessel: 10.5 litres
- Material, size, headspace, fill volume: all-glass, containing 10 litres of test solution
- Aeration: the test media were not aerated during the test
- Renewal rate of test solution (frequency/flow rate): daily
- No. of organisms per vessel: 7
- No. of vessels per concentration (replicates): 1
- No. of vessels per control (replicates): 1
- Biomass loading rate: 0.36 g fish/litre i.e. 7 fish per 10 litres of test medium

ISO-medium, aerated until the dissolved oxygen concentration had reached saturation and the pH had stabilised. After aeration the hardness was 250 mg CaCO3 per litre and the pH was 7.9-8.0

- Photoperiod: 16 hours daily

- Test concentrations: A flitrate prepared at a loading rate of 100 mg/l and dilutions containing 10, 18, 32 and 56% of the filtrate.
Reference substance (positive control):
24 h
Dose descriptor:
Effect conc.:
56 mg/L
Basis for effect:
mortality (fish)
48 h
Dose descriptor:
Effect conc.:
42 mg/L
Basis for effect:
mortality (fish)
72 h
Dose descriptor:
Effect conc.:
32 mg/L
Basis for effect:
mortality (fish)
96 h
Dose descriptor:
Effect conc.:
26 mg/L
Basis for effect:
mortality (fish)
Remarks on result:
other: 24 -33 mg/L
96 h
Dose descriptor:
Effect conc.:
10 mg/L
Reported statistics and error estimates:
Mortality in carp at 56 and 100 mg/l at 24 hours.
Mortality in carp at 32 mg/l at 72 hours.

Clinical Effects:
No abnormalities in carp at 0 or 10 mg/l
Immobilisation observed in carp at 18, 32, 56 and 100 mg/l
Discoloration observed in carp at 32, 56, and 100 mg/l
One carp at 32 mg/l observed snapping and swimming upright at surface at 96 hrs.
Sublethal observations / clinical signs:


 Concentration (mg/l)  N

             Cumulative Mortality

 Total Mortality (%)
 2 1/4h  24h  48h  72h  96h  
 Blank-control  7  0  0  0  0  0  0
 10  7  0  0  0  0  0  0
 18  7  0  0  0  0  0  0
 32  7  0  0  0  3  6  86
 56  7  0  7  7  7  7  100
 100  7  0  7  7  7  7  100


 Concentration (mg/l)  Time (hours)  Effects  Relative Number
 Blank-control, 10  0 -96  No abnormalities  7/7
 18  0 -72  No abnormalities  7/7
   96  No abnormalities  6/7
     Immobilised  1/7
 32  0 -24  No abnormalities  7/7
   48  No abnormalities  3/7
     Immobilised  2/7
     Discoloured and swimming at the surface  2/7
   72  No abnormalities  1/4
     Immobilised  2/4
     Discoloured and swimming at the surface  1/4
   96  Snapping and swimming upright at the surface  1/1
 56  0 -2 1/4  No abnormalities  7/7
   24  Immobilised  2/4
     Discoloured  2/4
 100  2 1/4  Discoloured  7/7
Validity criteria fulfilled:
Under the conditions of the present test, Unoxol 3,4 Dialdehyde induced no visible effects in carp at 10 mg/L (NOEC). The 96-hour LC50 was 26 mg/L based on measured concentrations (95% confidence interval between 24 and 33 mg/L).
Executive summary:

Carp (Cyprinus carpio) were exposed to Unoxol 3,4 Dialdehyde under semi-static conditions with daily renewals for 96 hours. Measured concentrations were close to the nominal loading rate. Temperature, pH, and dissolved oxygen levels were within required specifications for the test organism. Under the conditions of the present test, Unoxol 3,4 Dialdehyde induced no visible effects in carp at 10 mg/L (NOEC). The 96-hour LC50 was 26 mg/L based on measured concentrations (95% confidence interval between 24 and 33 mg/L).

Description of key information

The 96-hour LC50 value of 26 mg/L for Unoxol 3,4-Dialdehyde was taken from a valid, acute, exposure study with carp (Cyprinus carpio).

Key value for chemical safety assessment

Fresh water fish

Fresh water fish
Effect concentration:
26 mg/L

Additional information

Two acute freshwater fish studies were assessed for this endpoint. One study was conducted with carp (Cyprinus carpio), and this study was determined to be acceptable for use in risk assessment (Klimisch score = 1). The other study was conducted with Chinese rare minnows (Gobiocyprus rarus), and was determined to be acceptable for use in risk assessment (Klimisch score = 1). Both fish acute studies exposed the test organisms to a series of five test concentrations and a negative (dilution water) control for 96 hours in a static-renewal exposure system. In both studies, test solutions were renewed every 24 hours. Observations of mortality and other signs of toxicity were made at or shortly after test initiation, as well as 24, 48, 72 and 96 hours after test initiation. The 96-hour LC50 was determined to be 26 mg/L in the fish acute study with carp. The 96-hour LC50 was determined to be 65.1 mg/L in the fish acute study with the Chinese rare minnows. Based on these results, the carp study was selected as the key study since the LC50 value was lower for this species.