Reactive Blue F08-0170

Short-term toxicity to fish

Administrative data

Endpoint:

short-term toxicity to fish

Type of information:

experimental study

Adequacy of study:

key study

Study period:

11 July to 15 July 2011

Reliability:

1 (reliable without restriction)

Rationale for reliability incl. deficiencies:

other: Study conducted to OECD test guidance in compliance with GLP.

Data source

Reference

Reference Type:

study report

Title:

Unnamed

Year:

2011

Report date:

2012

Materials and methods

Test guideline

Qualifier:

according to guideline

Guideline:

OECD Guideline 203 (Fish, Acute Toxicity Test)

Deviations:

no

GLP compliance:

yes

Test material

Test material information

Constituent 1

Details on test material:

Reactive Blue F08-0170

Specific details on test material used for the study:

Details on properties of test surrogate or analogue material (migrated information):
Not applicable.

Sampling and analysis

Analytical monitoring:

yes

Details on sampling:

The actual concentrations of nominal concentration of 370 mg/L test substance solution and blank control was measured at 0h, 96h and 24h, 48h, 72h (after and before the renewal) after the start of the test by a High Performance Liquid Chromatography (HPLC). See below for details.

Test solutions

Vehicle:

no

Details on test solutions:

Test water
The chlorine of drinking water was removed by aeration.
Hardness: 112mg/L (expressed as CaCO3)
pH: 7.81

Preparation of test solution
1.850g test substance was dissolved in a certain amount of test water with ultrasonic treatment for 3 min. Then the solution was diluted with the test water to 5000mL and was stirred with a magnetic stirrer for 1h. A nominal concentration of 370 mg/L test solution (102 mg/L calculated based on 27.48% purity of main the constituent VS/VSE) was obtained. The test item Reactive Blue F08-0170 is a multiconstituent composed of 6 constituents. VS/VSE as the most important constituent was chosen as analytical lead substance for monitoring the stability of the test substance in the test medium.

Test organisms

Test organisms (species):

Danio rerio (previous name: Brachydanio rerio)

Details on test organisms:

Reason for selection of species
The test species is recommended by "The Guidelines for the Testing of Chemicals"
Supplier: Kang Da Breeding Farm
Acclimation:
Zebra fish (Brachydanio rerio) was obtained on April 26, 2011.
Before the start of the test, Zebra fish (Brachydanio rerio) were acclimated for 17 days by flow-through under the same conditions of water quality (Dechlorinated tap water), temperature (23℃±2℃), light and dark period (14h light / 10h dark). The dissolved oxygen concentration during acclimation was kept at least 80 % of the air saturation value. The test fish were fed daily until 24 hours before the start of the test.
The mortality was 0% during 7 days before the start of the test, which met the requirement.
Size: 10 fish were randomly selected and measured on May 18, 2011. The average and standard deviation were 26.84 mm±1.97 mm at total length and the average and standard deviation were 0.295 g±0.117 g at weight in the population from which the fish were selected.
Allocation to the test groups: Zebra fish (Brachydanio rerio) were allocated at random to each test group within 30 minutes.

Study design

Test type:

semi-static

Water media type:

freshwater

Limit test:

yes

Total exposure duration:

96 h

Post exposure observation period:

None

Test conditions

Hardness:

112mg/L (expressed as CaCO3)

Test temperature:

24.0℃ to 24.9℃

pH:

7.51 to 8.4

Dissolved oxygen:

74% to 96%. The test solutions were not aerated during the exposure. The dissolved oxygen concentration in the test solutions during the test was kept at least 60 % of the air saturation value.

Salinity:

Not applicable.

Nominal and measured concentrations:

Nominal concentration of 370 mg/L of test substance solution (102 mg/L calculated based on 27.48% purity of main constituent VS/VSE).

Details on test conditions:

Test conditions:
Conditions of exposure
Type of test: Semi-static test (renewal at every 24h)
Exposure duration: 96hr
Test concentration: Nominal concentration of 370 mg/L of test substance solution (102 mg/L calculated based on 27.48% purity of main constituent VS/VSE).
Control: A blank control (test water without test substance) was also run in parallel to the test concentration.
Replicate: No replicate
Number of fish: 10 fish / test group
Volume of test solution: 3L / test vessel

Conditions of test environment:
The test was performed without adjustment of pH.
Light: Artificial light of white fluorescent lamp, 14h light / 10h darkness
Feeding: The fish were not fed during the exposure.

Reference substance (positive control):

no

Results and discussion

Effect concentrations

Effect concentrations 1

Duration:

96 h

Dose descriptor:

LC50

Effect conc.:

> 148.2 mg/L

Nominal / measured:

meas. (geom. mean)

Conc. based on:

test mat.

Basis for effect:

mortality (fish)

Effect concentrations 2

Duration:

96 h

Dose descriptor:

NOEC

Effect conc.:

> 148.2 mg/L

Nominal / measured:

meas. (geom. mean)

Conc. based on:

test mat.

Basis for effect:

mortality (fish)

Details on results:

Mortality
Cumulative mortality data of each group at 24h, 48h, 72h and 96h after the start of the test are shown in Table 1. below.
The cumulative mortality of blank control was 0% at the end of the test, which met the criterion for the validity of the test .
No deaths occurred in either test group.

Observed abnormal response
Data on abnormal response observed during the test are shown in Table 2 below.
No abnormal response was observed in blank control and test substance group.

Measurement of temperature, pH and dissolved oxygen of test solution
The temperature, pH and dissolved oxygen of each test group at 0h, 96h and 24h, 48h, 72h (after and before the renewal) after the start of the test are shown in Tables 3 - 5 below.
During the test, the measurement values of pH, temperature and dissolved oxygen ranged from 7.51 to 8.40, 24.0℃ to 24.9℃, and 74% to 96%,respectively. The dissolved oxygen concentrations were kept at least 60 percent of the air saturation value, which met the criterion for the validity of the test.

Concentration of main constituent VS/VSE in test solution
The measured concentrations of main constituent VS/VSE in blank control was all < 0.06410mg/L(LOD) at 0h, 96h and 24h, 48h, 72h (after and before the renewal) after the start of the test.
The measured concentrations of main constituent VS/VSE in nominal concentration of 370 mg/L test substance solutions at 0h, 96h and 24h, 48h, 72h (after and before the renewal) after the start of the test were 144.6 mg/L (0h), 153.4mg/L (24h old), 146.6mg/L (24h new), 143.4mg/L (48h old), 138.6mg/L (48h new), 162.8mg/L (72h old), 146.4mg/L (72h new) and 151.1mg/L (96h), which were exceeded ±20% of the nominal concentration of main constituent VS/VSE (102 mg/L).

Median Lethal Concentration (LC50)
The LC50 at 24h, 48h, 72h and 96h after the start of the test are shown in Table 6.
At the end of the test, the cumulative mortality of nominal concentration of 370 mg/L test substance group was 0%, so the 96h-LC50 of test substance to Zebra fish (Brachydanio rerio) was greater than nominal concentration of 370 mg/L test substance solution (148.2 mg/L calculated based on the actual concentration of main constituent VS/VSE).

Results with reference substance (positive control):

Not applicable.

Reported statistics and error estimates:

None.

Any other information on results incl. tables

Sublethal observations / clinical signs:

Table 1 Cumulative mortality(%)

Nominal concentration (mg/L)	Number of test fish	Cumulative mortality（%）
		24h	48h	72h	96h
Blank control	10	0	0	0	0
370	10	0	0	0	0

 

Table 2 Observed abnormal response

Nominal concentration (mg/L)	Result of observation (Left column: Number of affected fish / Total survival number, Right column: Symptom detail)
	24h		48h		72h		96h
Blank control	0/10	N	0/10	N	0/10	N	0/10	N
370	0/10	N	0/10	N	0/10	N	0/10	N

Note: N—Normal

Table 3 Temperature of test solutions (°C)

Nominal concentration (mg/L)	0h	24h		48h		72h		96h
		old	new	old	new	old	new
Blank control	24.1	24.1	24.1	24.4	24.4	24.2	24.7	24.3
370	24.0	24.3	24.0	24.6	24.9	24.2	24.5	24.3

 

Table 4 pH of test solutions

Nominal concentration (mg/L)	0h	24h		48h		72h		96h
		old	new	old	new	old	new
Blank control	8.40	8.04	7.97	7.55	8.36	7.96	8.36	7.80
370	7.93	7.63	7.51	7.83	8.08	7.83	8.05	7.85

Table 5 Dissolved oxygen concentration of test solutions

Nominal concentration (mg/L)	0h	24h		48h		72h		96h
		old	new	old	new	old	new
Blank control	94	80	93	74	89	87	91	85
370	93	79	96	85	94	87	96	86

 

Table 6 Median lethal concentration (LC₅₀) and the 95% confidence limits

Exposure duration	LC50(mg/L)^1	95% confidence limits
24 h	＞148.2	—
48 h	＞148.2	—
72 h	＞148.2	—
96 h	＞148.2	—

Note:1 — Calculated based on geometric mean of actual concentration of one of the 6 constituents of the test item.

Applicant's summary and conclusion

Validity criteria fulfilled:

yes

Conclusions:

Under the conditions of the test, the 96h-LC50 of test substance to Zebra fish (Brachydanio rerio) was greater than nominal concentration of 370 mg/L test substance solution (148.2 mg/L calculated based on the geometric mean of actual concentration of main constituent VS/VSE). 96h Lowest Observed Effect Concentration (LOEC) was greater than nominal concentration of 370 mg/L test substance solution (148.2 mg/L calculated based on the geometric mean of actual concentration of main constituent VS/VSE), 96h No Observed Effect Concentration (NOEC) was nominal concentration of 370 mg/L test substance solution (148.2 mg/L calculated based on the geometric mean of actual concentration of main constituent VS/VSE).

Executive summary:

An acute fish toxicity test of Reactive Blue F08 -0170 was conducted according to OECD Guidelines for the Testing of Chemicals, 203 Fish Acute Toxicity Test. Paris: OECD, Adopted 17 July 1992.

Under the conditions of the test, the 96h-LC50 of test substance to Zebra fish (Brachydanio rerio) was greater than nominal concentration of 370 mg/L test substance solution (148.2 mg/L calculated based on the geometric mean of actual concentration of main constituent VS/VSE). 96h Lowest Observed Effect Concentration (LOEC) was greater than nominal concentration of 370 mg/L test substance solution (148.2 mg/L calculated based on the geometric mean of actual concentration of main constituent VS/VSE), 96h No Observed Effect Concentration (NOEC) was nominal concentration of 370 mg/L test substance solution (148.2 mg/L calculated based on the geometric mean of actual concentration of main constituent VS/VSE).