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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
2 (reliable with restrictions)
Rationale for reliability incl. deficiencies:
other: Not all details available on chemical analysis method and results.
according to guideline
OECD Guideline 203 (Fish, Acute Toxicity Test)
GLP compliance:
GLP issued by State Environmental Protection Administration (SEPA) of the People’s Republic of China
Analytical monitoring:
Details on sampling:
Samples were taken (at least in duplicate) from each concentration during the Limit Test at 0, 24, 48, 72 and 96 h. On each occasion, one sample was analysed after pre-treatments; the remaining samples were retained in case further analysis would be required.

Pre-treatment method of the sample
10.0 ml collected water samples were diluted to 50.0 mL by 40% methanol solution (methanol: water = 40:60 (v:v)) and filtrated by 0.22 μm millipore membrane (discarding the initial 10mL filtrates), then the filtrates were analysed by UPLC.
Details on test solutions:
In the range-finding test and limit test, the test solutions of meta-Diisopropanolbenzene were prepared by directly dissolving appropriate amount of meta-Diisopropanolbenzene in dilution medium and then facilitating its dispersion by ultrasonication for 20 min.
Test organisms (species):
Danio rerio (previous name: Brachydanio rerio)
Details on test organisms:
The test species of Zebra fish, Brachydanio rerio (Batch No. F20111115), were obtained as fry from a commercial fish supplier of Guangzhou Hongfa Aquatic Animals Culture Company in China.
Feeding fry were held for 45 days in holding tanks supplied with a continuous flow of aerated water before being used for testing. Fish to be used in the test was held for 7 days in water of the quality and temperature to be used in the test.
A photoperiod of 16 hours light, provided by overhead fluorescent tubes, and 8 hours dark was maintained. The oxygen concentration was more than 90% of the air saturation value.
The fish was fed daily during the holding period on proprietary fish food. They were held without food for approximately 24 hours before being placed into the test vessels. The ingredients of the fish food are given as follows:
Crude Protein >49.0%
Crude Fat > 5.0%
Crude Fibre < 3.0%
Crude Ash < 12.0%
Moisture < 10.0%
During the holding period the tanks was inspected daily and any debris or unhealthy or dead fish removed.
After the 48 hour settling-in period, no mortality was observed in the following 7 days. So this batch of fish (Batch No. F20111115) was accepted.
The wet-weight of the fish used to the test was 0.20 g in average, with an average length of 2.30 cm.
Test type:
Water media type:
Limit test:
Total exposure duration:
96 h
119 to 125 mg (CaCO3)/L
Test temperature:
23.0 to 23.3 °C
7.40 to 7.72
Dissolved oxygen:
85% to 100% of the air saturation
Nominal and measured concentrations:
Nominal: 100 and 150 mg/L
Measured: 93.9 and 147 mg/L
Details on test conditions:
- Test vessel: Tanks made of chemically inert material, with a sealable inert lid, and with a capacity of approximately 8 L
- Aeration: no
- No. of organisms per vessel: 10
- No. of vessels per concentration (replicates): 3
- No. of vessels per control (replicates): 3
- Biomass loading rate: 0.2 g to 0.6 g fish (wet weight) per litre of test medium
- Light: 16 hours photoperiod daily (light intensity: 1000 to 1500 lux)
- Feeding: none

Good quality tap water which had been dechlorinated for at least 24 hours was used. The total hardness of the test water was 122.2 mg (CaCO3)/L, oxygen concentration was about 98% of the air saturation value and pH was 7.55 at room temperature. Characteristics of the dilution water are measured at least twice a year by Jiangsu Provincial Center for Disease Prevention and Control.

In the range-finding test, groups of fish (5 per group) were exposed to the test solutions with nominal concentration of 1, 10 and 100 mg/L. Synchronously one control group was conducted in test water without the test item. For each test tank 5 L test solution was filled in. No replicates were used.
The test fish were be randomly chosen and put in different test solutions after the temperature has been adjusted to the required value. This was done in 30 min.
Test duration was 96 hours and no fish died in any of the concentrations.
Reference substance (positive control):
96 h
Dose descriptor:
Effect conc.:
> 150 mg/L
Nominal / measured:
Conc. based on:
test mat.
Basis for effect:
mortality (fish)
96 h
Dose descriptor:
Effect conc.:
>= 150 mg/L
Nominal / measured:
Conc. based on:
test mat.
Basis for effect:
mortality (fish)
Details on results:
The results indicated that concentration of meta-Diisopropanolbenzene was stable (within 80% of the initial concentration) in the water during the test period. Thus static method used in the Limit Test was reasonable.

All fish in the control groups and treated groups were alive and appeared normal.
Results with reference substance (positive control):
24 h-LC50 was 284 mg/L
Validity criteria fulfilled:
The study was performed under Chinese GLP. This study can be used for risk assessment and classification and labelling, with some restrictions.
Executive summary:

Under static conditions, the acute toxicity of test item (meta-Diisopropanolbenzene) to zebra fish (Brachydanio rerio) was conducted according to“The guidelines for the testing of chemicals”, SEPA(HJ/T 153-2004); and with reference toProcedure 203 of the “Guidelines for Testing of Chemicals” of the OECD: "Fish, Acute Toxicity Test” (1992) etc. A range-finding test and then a limit test were performed respectively. Nominal concentrations of 1, 10 and 100 mg/L were used in the range-finding test, while nominal concentrations of 100 mg/L and 150 mg/L were used in the limit test. Water samples taken from the control and the test treatments in the limit test were analysed. The mean analyzed concentrations were 93.9 mg/L and 147 mg/L respectively. The analytical results showed that the concentration of the test item is consistent in the test medium throughout the 96-hour test period (deviation within 20%). Thus a static procedure was reasonable.In the range-finding test, 5 fishes per treatment and no replicates were used, while 10 fishes per treatment and 3 replicates in the limit test.The test fishes were exposed for 96 hours to the testsolution.During the whole test period, the pH values of the control mediums and test mediums were between 7.40 and 7.72, and the Dissolved Oxygen (DO) values varied from 85% to 100% of the air saturation at the test temperature, and the total hardness was in the range of 119 to 125 mg(CaCO3)/L. During the test, the temperature of the test mediums were maintained in the range of 23.0oC to 23.3oC, and all fishes in the control group were normal. With the same conditions, K2Cr2O7 was used as the negative control substance, and the resulting 24 h-LC50was 284 mg/L.So the study met the acceptability criteria prescribed by the protocol (pH:6.0~8.5; dissolved oxygen concentration: > 60% of the air saturation value;total hardness:10~250 mg(CaCO3)/L; temperature: (23±2)oC;24 h-LC50ofK2Cr2O7in the range of 200 to 400 mg/L). Therefore the test was considered valid.During the whole test period, all fish in the control groups and treated groups were alive and appeared normal.The results showed that under valid static test conditions, the maximum concentration causing no mortality and the 96 h-LC50of the test item to zebra fish are both greater than the nominal concentration of 150mg/L(measured concentration 147 mg/L).According to “The guidelines for the hazard evaluation of new chemical substances (HJ/T 154 -2004)”, the test substance of meta-Diisopropanolbenzene belongs to low toxic chemical for fish (zebra fish, Brachydanio rerio).

Description of key information

The results showed that under valid static test conditions, the maximum concentration causing no mortality and the 96 h-LC50of the test item to zebra fish are both greater than the nominal concentration of 150mg/L(measured concentration 147 mg/L)

Key value for chemical safety assessment

Additional information

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