Registration Dossier

Administrative data

Endpoint:
short-term toxicity to fish
Type of information:
experimental study
Adequacy of study:
key study
Study period:
From 2001-02-12 to 2001-04-30
Reliability:
2 (reliable with restrictions)
Rationale for reliability incl. deficiencies:
other: GLP study conducted in compliance with agreed test protocols following OECD Method 203. Acceptable restriction was that analytical measurements on the test material were not conducted.
Cross-referenceopen allclose all
Reason / purpose:
reference to same study
Reason / purpose:
reference to other study

Data source

Reference
Reference Type:
study report
Title:
Unnamed
Year:
2001
Report Date:
2001

Materials and methods

Test guidelineopen allclose all
Qualifier:
according to
Guideline:
OECD Guideline 203 (Fish, Acute Toxicity Test)
Deviations:
no
Qualifier:
according to
Guideline:
EU Method C.1 (Acute Toxicity for Fish)
Deviations:
no
Principles of method if other than guideline:
Not applicable.
GLP compliance:
yes (incl. certificate)
Remarks:
2000-04-26

Test material

Reference
Name:
Unnamed
Type:
Constituent
Details on test material:
- Substance type: brown viscous liquid
- Physical state: liquid
- Analytical purity: no data
- Impurities (identity and concentrations): no data
- Composition of test material, percentage of components: no data
- Isomers composition: no data
- Purity test date: no data
- Lot/batch No.: no data
- Expiration date of the lot/batch: no data
- Stability under test conditions: no data
- Storage condition of test material: at room temperature in the dark
- Other: none
Specific details on test material used for the study:
Details on properties of test surrogate or analogue material (migrated information):
None.

Sampling and analysis

Analytical monitoring:
no
Details on sampling:
- Concentrations: 0, and 1000 mg/L WAF loading rates. Limit test approach used based on range finding study. It was considered unnecessary and unrealistic to test at loading rates in excess of 1000 mg/L.
- Sample storage conditions before analysis: not applicable.

Test solutions

Vehicle:
no
Details on test solutions:
PREPARATION AND APPLICATION OF TEST SOLUTION (especially for difficult test substances)
- Method: The test material was prepared as a Water Accommodated Fraction. Using a syringe, an amount of test material (21 .OO g) was added onto the surface of dechlorinated tap water (2 1 litres) to give the 1000 mg/l loading rate. This was then stirred, by magnetic stirrer, using a stirring rate approximately equal to 30 rpm. This stirring rate ensured that there was no gross mixing of the test material. This was stirred for 24 hours. The stirring was stopped after 24 hours and the mixture allowed to settle for 2 hours.
A wide bore glass tube covered at one end with parafilm was submerged into the vessel, sealed end down, to a depth of approximately 5 cm from the bottom of the vessel. Microscopic observations made on the WAF indicated that a significant amount of dispersed test material was present in the water column and hence it was considered justifiable to remove undissolved material by filtering through a glass wool plug (2-4 cm in length) to trap microemulsions. Prior to use the glass wool was preconditioned by thoroughly soaking overnight in deionised reverse osmosis water, rinsing three times with deionised reverse osmosis water and dechlorinated tap water followed by autoclaving. This preconditioning of the glass wool was a deviation from the protocol however the use of unconditioned glass wool has been shown to signifcantly alter the pH of the dilution water. This deviation did not affect the results or integrity of the study. The aqueous phase or WAF was then removed by siphon (the first 75-100 mL discarded) and used for testing.
- Eluate: Laboratory tap water dechlorinated and partly softened.
- Differential loading: not applicable.
- Controls: Laboratory tap water dechlorinated and partly softened.
- Evidence of undissolved material (e.g. precipitate, surface film, etc): none.

Test organisms

Test organisms (species):
Oncorhynchus mykiss (previous name: Salmo gairdneri)
Details on test organisms:
TEST ORGANISM
- Common name: rainbow trout
- Strain: no data
- Source: Donnington Fish farm Upper Swell, Gloucestershire, UK
- Age at study initiation (mean and range, SD): no data
- Length at study termination (length definition, mean, range and SD): 4.5+/- 0.4 cm
- Weight at study termination (mean and range, SD): 0.77 +/- 0.2 g at the end of the study.

At the start of the study, 10 fish were placed in each test vessel at random, in the prepared test solutions. The test vessels were then covered to reduce evaporation and maintained at 14°C in a temperature controlled room with a photoperiod of 16:8 h, with 20 min dawn and dusk transition periods for a period of 96 h. The control group was maintained under identical conditions but not exposed to the test material.

ACCLIMATION
- Acclimation period: Fish were acclimatized to test conditions from 15 February 2001 to 1 March 2001
- Acclimation conditions (same as test or not): yes
- Type and amount of food: fish were fed with commercial trout pellets.
- Feeding frequency: Fish were fed until approximately 48 hours prior to the start of the definitive test.
- Health during acclimation (any mortality observed): Less than 1% mortality in 7 days prior to start of test.

Study design

Test type:
static
Water media type:
freshwater
Limit test:
yes
Total exposure duration:
96 h
Post exposure observation period:
none

Test conditions

Hardness:
100 mg/L as CaCO3.
Test temperature:
14.0°C
pH:
7.6-7.8
Dissolved oxygen:
8.2-9.7 mg O2/L.
Salinity:
not applicable.
Nominal and measured concentrations:
nominal concentrations: 0 and 1000 mg/L
Details on test conditions:
TEST SYSTEM
- Test vessel: 20 L glass
- Type (delete if not applicable): closed, static
- Material, size, headspace, fill volume: 20 L
- Aeration: by narrow bore glass tubes.
- No. of organisms per vessel: 10
- No. of vessels per concentration (replicates): 2
- No. of vessels per control (replicates): 1
- Biomass loading rate: 0.39 g bw/L


OTHER TEST CONDITIONS
- Adjustment of pH: no data
- Photoperiod: 1:8
- Light intensity: no data
- Stirring rate recorded periodically

TEST CONCENTRATIONS
- Range finding study
- Test concentrations: 0, 10, 100, 1000 mg/L
- Results used to determine the conditions for the definitive study: no effect, in the range-finding study.
Reference substance (positive control):
no

Results and discussion

Effect concentrationsopen allclose all
Duration:
96 h
Dose descriptor:
LL50
Effect conc.:
> 1 000 mg/L
Nominal / measured:
nominal
Conc. based on:
test mat.
Basis for effect:
mortality
Duration:
96 h
Dose descriptor:
NOEC
Effect conc.:
1 000 mg/L
Nominal / measured:
nominal
Conc. based on:
test mat.
Basis for effect:
mortality
Details on results:
There were no mortalities in 20 fish exposed to a 1000 mg/l loading rate WAF for a period of 96 hours.
Results with reference substance (positive control):
Not applicable
Reported statistics and error estimates:
None.

Any other information on results incl. tables

Table 1: Cumulative mortality data in the definitive study.

Nominal loading rate (mg/L)

Cumulative mortality (n = 10)

% mortality

 

3 h

6 h

24 h

48 h

72 h

96 h

96 h

Control

0

0

0

0

0

0

0

1000 R1

0

0

0

0

0

0

0

1000 R2

0

0

0

0

0

0

0

Applicant's summary and conclusion

Validity criteria fulfilled:
yes
Conclusions:
The 96-Hour LL50 based on nominal loading rates was greater than 1000 mg/L loading rate WAF and correspondingly the No Observed Effect Loading rate was 1000 mg/L loading rate WAF.
Executive summary:

Introduction: A study was performed to assess the acute toxicity of the test material to rainbow trout (Oncorhynchus mykiss). The method followed that described in the OECD Guidelines for Testing of Chemicals (1992) No 203, "Fish, Acute Toxicity Test'' referenced as Method C.l of Commission Directive 92/69/EEC (which constitutes Annex V of Council Directive 67/548/EEC), US CFR Title 40, Part 797, Section 1400 and US EPA Draft Ecological Effects Test Guideline OPPTS 850.1075.

Methods: Following a preliminary range-finding study fish were exposed, in two groups of ten, to a Water Accommodated Fraction (WAF) of the test material, at a single nominal loading rate of 1000 mg/L for a period of 96 hours under static test conditions. The number of mortalities and any sub-lethal effects of exposure in each test and control vessel were determined 3 and 6 hours after the start of exposure and then daily throughout the study until termination after 96 hours.

Results: The 96-Hour LL50* based on nominal loading rates was greater than 1000 mg/L loading rate WAF and correspondingly the No Observed Effect Loading rate was 1000 mg/L loading rate WAF.

It was considered unnecessary and unrealistic to test at loading rates in excess of 1000 mg/L.

*LL = Lethal Loading rate