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Short-term toxicity to fish

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Reference
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: GLP study, OECD 203 compliant
Qualifier:
according to
Guideline:
OECD Guideline 203 (Fish, Acute Toxicity Test)
Deviations:
no
GLP compliance:
yes (incl. certificate)
Analytical monitoring:
yes
Details on sampling:
For the purpose of chemical analysis, duplicate aliquots (ca 20 mL) were removed from test and control WAF at the commencement of the test (0 h) and at 72 h as ‘fresh’ solution and at 24 and 96 h from test and control tanks as ‘expired’ solution.
Vehicle:
no
Test organisms (species):
Danio rerio (previous name: Brachydanio rerio)
Details on test organisms:
Zebra fish (Brachydanio rerio) in the size range 1-3 cm were used for this test and were received from Water Wonders, Edinburgh on 17 July 2009. All fish were in good health and free from any apparent malformation. All fish were acclimatised to laboratory conditions for a minimum of 14 days prior to commencement of the test. During holding, the fish were fed daily on a suitable standard fish diet. The fish were not fed for a period of 24 h prior to test commencement or throughout the duration of the test.
Test type:
semi-static
Water media type:
freshwater
Limit test:
yes
Total exposure duration:
96 h
Post exposure observation period:
no
Hardness:
82.8 mg CaCO3/L
Test temperature:
22.0 - 23.0 °C
pH:
Fresh medium: 6.3 -6.7
Prior to medium renewal: 5.7 - 6.3
Dissolved oxygen:
Fresh medium: 86 - 92 % air saturation value
Prior to medium renewal: 57 - 67 % asv
Nominal and measured concentrations:
Saturated solutions were prepared with nominal concentrations of 5 mg/L. Measured concentrations for fresh solutions were equal to 379 µg/L (T = 0 h) and 334 µg/L (T = 72 h). Measured concentrations for expired solutions were equal to 172 µg/L (T = 24 h) and 235 µg/L (T = 96 h). This means that deviation of expired wrt fresh concentrations were -55 and -30%. Overall geometric mean of exposure concentration for this limit test was 267 µg/L
Details on test conditions:
TEST SYSTEM
Tanks of ca 20 L capacity of moulded glass construction and sealed with glass lids to prevent a head-space of air in the tank
One tank was prepared at the initial loading rate of 1 mg dibenzyltoluene/L and one control, containing control WAF.
Seven zebra fish were added to each tank within 30 min of preparation.
Fish were transferred to freshly prepared test solutions at 24 h intervals.

TEST MEDIUM / WATER PARAMETERS
Reconstituted freshwater (RFW) prepared at Charles River, with high grade salts and reverse osmosis grade water was used during the holding and testing phases of the study. The salts were dissolved in a volume of deionised water (500 L) and this solution was metered into a flow of deionised water, to produce RFW according to the formula recommended by OECD Guideline 203.

Reference substance (positive control):
no
Duration:
96 h
Dose descriptor:
LC50
Effect conc.:
> 267 µg/L
Nominal / measured:
meas. (geom. mean)
Conc. based on:
test mat.
Basis for effect:
mortality
Remarks on result:
other: no effect at solubility limit, ca. 267 µg/L
Duration:
96 h
Dose descriptor:
LC0
Effect conc.:
267 µg/L
Nominal / measured:
meas. (arithm. mean)
Conc. based on:
test mat.
Basis for effect:
mortality
Remarks on result:
other: no effect at solubility limit, ca. 267 µg/L
Details on results:
The fish in the limit test were observed after 1, 4, 24, 48, 72 and 96 h.
At the termination of the test, the length (from the tip of the snout to the caudal peduncle) and weight of each surviving fish was recorded.
Analytical samples taken at 0 and 24 h and at 72 and 96 h.

All test solutions appeared clear and colourless throughout the test period. No precipitation of benzyltoluene was observed.

There was no mortality recorded during the test. All fish appeared active and healthy throughout.

The length and weight of fish after 96 h exposure were measured. Fish were within the range 26-30 mm in length and 0.217-0..391 g in weight.

Reported statistics and error estimates:
Statistical analysis was not conducted on the data as there was no mortality.

Table 1: Recorded mortality of fish throughout the study.

Observation Time

Number of dead fish

Control

Saturated solution

0 h

0

0

24 h

0

0

48 h

0

0

72 h

0

0

96 h

0

0

 

 Table 2:  Measured and mean estimates of test item concentration throughout the study in the exposed group.

Time (h)

Measured concentrations (µg/L)

Geometrical mean (µg/L)

Control

Saturated solution

Saturated solution

Global : 267 µg/L

0

ND

379

255

24

ND

172

72

ND

334

280

96

ND

235

ND : not detected

Validity criteria fulfilled:
yes
Conclusions:
The 96 h LC50 of dibenzyltoluene to zebra fish is concluded to be greater than the maximum solubility of dibenzyltoluene in water, under the conditions of the test. The 96 h NOEC is concluded to be the maximum solubility of dibenzyltoluene in test water.

Executive summary:

The acute toxicity of benzyltoluene to zebra fish (Brachydanio rerio)was assessed by conducting a limit test at the maximum solubility of benzyltoluene under test conditions. The test was conducted over a 96 h period under semi-static conditions, in accordance with OECD (1992) Guideline 203, following procedures in OECD (2000) Guidance Document No. 23 on Testing Difficult Substances.

Test solutions were prepared as Water Accommodated Fractions (WAF’s) from an initial loading rate of 5 mg benzyltoluene/L. The method of test solution preparation was selected to maximise the solubility of benzyltoluene under test exposure conditions, while minimising exposure to insoluble fractions. A control of untreated test water was also included.

One test tank (containing saturated solution of benzyltoluene) and one control tank were prepared and 7 zebra fish were added to each. Fish were transferred to freshly prepared solutions at 24 h intervals. No mortality or other adverse effects were noted throughout the test period. Analytical samples taken at 0 and 24 h and at 72 and 96 h indicated that benzyltoluene is only partially soluble in test water with concentrations ranging from 172 to 379 µg/L.

Measured concentrations of “expired” test solutions at 24 and 96 h, showed that initial concentrations of benzyltoluene were not maintained. The loss over the 24 h period between test solution renewals ranged from 30-55%. Due to the significant loss in concentration over the period between test solution renewal, the geometric mean measured concentration was calculated. The overall (0, 24, 72 and 96 h) geometric mean measured concentration of benzyltoluene in the test was 267 µg/L. 

The 96 h LC50of benzyltoluene to zebra fish is concluded to be greater than the maximum solubility of benzyltoluene in water, under the conditions of the test. The no observed effect concentration (NOEC, 0% mortality) is concluded to be the maximum solubility of benzyltoluene in test water.

Description of key information

The 96 h LC50 of benzyltoluene to zebra fish is concluded to be greater than the maximum solubility of benzyltoluene in water, under the conditions of the test. The no observed effect concentration (NOEC) is concluded to be the maximum solubility of benzyltoluene in test water

Key value for chemical safety assessment

Additional information

The acute toxicity of benzyltoluene to zebra fish (Danio rerio)was assessed by conducting a limit test at the maximum solubility of benzyltoluene under test conditions. The test was conducted over a 96 h period under semi-static conditions, in accordance with OECD (1992) Guideline 203, following procedures in OECD (2000) Guidance Document No. 23 on Testing Difficult Substances. Test solutions were prepared as Water Accommodated Fractions (WAF’s). The method of test solution preparation was selected to maximise the solubility of benzyltoluene under test exposure conditions, while minimising exposure to insoluble fractions. Fish were transferred to freshly prepared solutions at 24 h intervals. No mortality or other adverse effects were noted throughout the test period. The overall (0, 24, 72 and 96 h) geometric mean measured concentration of benzyltoluene in the test was 267 µg/L. The 96 h LC50 of benzyltoluene to zebra fish is concluded to be greater than the maximum solubility of benzyltoluene in water, under the conditions of the test. The no observed effect concentration (NOEC) is concluded to be the maximum solubility of benzyltoluene in test water.