2H-Pyran, tetrahydro-3-(phenylmethyl)-

Short-term toxicity to fish

Link to relevant study record(s)

Reference

Endpoint:

short-term toxicity to fish

Type of information:

experimental study

Adequacy of study:

key study

Study period:

From January 16, 2012 to January 20, 2012

Reliability:

2 (reliable with restrictions)

Rationale for reliability incl. deficiencies:

guideline study with acceptable restrictions

Remarks:

This study was performed according to national and international guidelines and is considered reliable with restrictions. No dose-response curve was observed. The LC50 value was determined as a geometric mean of LC0 and LC100. Test fish was a bit bigger than recommended : 2,6 +/- 1,7 cm (recommended 2 +/- 1 cm).The Certificate of Analysis of the test substance was not provided in the study report. Moreover, the substance is adequately identified, but details on composition are missing.

Qualifier:

according to guideline

Guideline:

other: 203 Fish Acute toxicity test. China Environmental Science Press

Version / remarks:

State Environmental Protection Administration of China. The Guidelines for the Testing of Chemicals, 203 Fish Acute toxicity test. China Environmental Science Press. 2004.

Deviations:

no

Qualifier:

according to guideline

Guideline:

OECD Guideline 203 (Fish, Acute Toxicity Test)

Version / remarks:

OECD. OECD Guidelines for the Testing of Chemicals, 203 Fish Acute Toxicity Test. Paris: OECD, Adopted 17 July 1992.

Deviations:

no

GLP compliance:

yes

Analytical monitoring:

yes

Details on sampling:

- Sampling method: The sample of blank control and 6.87mg/L, 11.0mg/L, 17.6mg/L test groups were directly analyzed after it were filtered through 0.45μm membrane. And the test solutions of 28.1mg/L and 45.0mg/L test groups were filtered by 0.45μm filter membrane, the filtrate diluted 2 times were analyzed. These samples were analyzed immediately. Otherwise the samples were stored at 4°C until analysis.

Vehicle:

no

Details on test solutions:

Preparation of test stock solution : 0.500g test substance was dissolved in some test water with ultrasonic treatment for 10min. Then the solution was transferred into 5L-jar, diluted with the test water to 5000mL and was stirred with a magnetic stirrer for 20min. Filtered with 0.45μm filter membrane. Abandoned the suspended fraction, the saturation concentration of filtered test solution of nominal concentration of 100mg/L test substance solution was obtained. The saturation solution was prepared every day (Three copies of the saturation solution were prepared at 0h after start of the test; one saturation solution was respectively prepared at 24h, 48h and 72h after start of the test).
The actual concentration of the saturation solution prepared every day was measured, the data of the actual concentrations were 85.42mg/L at 0h after start of the test, 84.82mg/L at 24h after start of the test, 79.55mg/L at 48h after start of the test and 73.72mg/L at 72h after start of the test. Then it was used to prepare the test solutions of each exposure level of test substance.

- Test concentrations : Five test concentrations of 6.87 mg/L, 11.0 mg/L, 17.6 mg/L, 28.1 mg/L and 45.0 mg/L as measured concentration of stock solution with a geometric series of a factor 1.6 (6.040 mg/L, 9.769 mg/L, 15.78 mg/L, 25.56 mg/L and 40.37mg/L as measured concentration in geometric mean). One blank control containing no test substance was set to the test series, too.

- The dissolved oxygen concentration in the test solutions was kept at least 60 percent of the air saturation value throughout the test.

Test organisms (species):

Danio rerio (previous name: Brachydanio rerio)

Details on test organisms:

TEST ORGANISM
- Source : Guangzhou Yang Shi Breeding Farm
- Common name: Danio rerio (previous name: Brachydanio rerio)
- Length at study initiation (length definition, mean, range and SD): 10 fish were randomly selected and measured on January 11, 2012. The average and standard deviation of the population from which the fish were selected were 26.04 mm ± 1.71 mm at total length
- Weight at study initiation (mean and range, SD): the average and standard deviation were 0.186 g ± 0.053 g at weight.

ACCLIMATION
- Acclimation period: 12 days
- Acclimation conditions (same as test or not): under the same conditions of water quality (Dechlorinated tap water), temperature (23 ± 2 °C), 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.
- Feeding frequency during acclimation: The test fish were fed daily until 24 hours before the start of the test.
- Health during acclimation (any mortality observed): The mortality was 0% during 7 days before the start of the test, which met the requirement.


FEEDING DURING TEST
- No feeding during the test

Test type:

semi-static

Water media type:

other: dechlorinated tap water

Limit test:

no

Total exposure duration:

96 h

Hardness:

141 mg/L (expressed as CaCO3)

Test temperature:

23 +/- 2°C

pH:

pH range : 7.96 - 8.43

Dissolved oxygen:

60 to 84 %

Salinity:

Not applicable

Conductivity:

Not available

Nominal and measured concentrations:

Nominal concentrations: 6,87 mg/l, 11,0 mg/L, 17,6 mg/L, 28,1 mg/L and 45,0 mg/L
Measured concentrations (geometric means): 6.040 mg/L, 9.769 mg/L, 15.78 mg/L, 25.56 mg/L and 40.37 mg/L

Details on test conditions:

TEST SYSTEM
- Type (delete if not applicable): open test vessels
- Material, size, headspace, fill volume: 3L / test vessel
- Aeration: no aeration
- Renewal rate of test solution (frequency/flow rate): every 24h
- No. of organisms per vessel: 7 fish / test vessel
- No. of vessels per concentration (replicates): 3 replicates per test group
- No. of vessels per control (replicates): 1 blank control

OTHER TEST CONDITIONS
- Photoperiod: 14h light / 10h darkness
- Light type: Artificial light of white fluorescent lamp
- The dissolved oxygen concentration in the test solutions was kept at least 60 percent of the air saturation value throughout the test.

EFFECT PARAMETERS MEASURED (with observation intervals if applicable) : Mortality and visible abnormality of the test fish were recorded at 3h, 24h, 48h, 72h and 96h after the start of the test during test period. The temperature, pH and dissolved oxygen of each test group were measured at 0h, 96h and 24h, 48h, 72h (before and after the renewal) after the start of the test. The actual concentrations of test substance of blank control and setting concentration 6.87 mg/L, 11.0 mg/L, 17.6 mg/L, 8.1 mg/L and 45.0 mg/L test groups were measured at 0h, 96h and 24h, 48h, 72h (before and after the renewal) by a High Performance Liquid Chromatography (HPLC).

TEST CONCENTRATIONS
- Spacing factor for test concentrations: 1.6
- Test concentrations: Five test concentrations : 6,87 mg/l, 11,0 mg/L, 17,6 mg/L, 28,1 mg/L and 45,0 mg/L (measured concentration of stock solution, factor 1,6).

Reference substance (positive control):

yes

Remarks:

Potassium dichromate was used as reference substance.

Key result

Duration:

96 h

Dose descriptor:

LC50

Effect conc.:

20.1 mg/L

Nominal / measured:

meas. (geom. mean)

Conc. based on:

test mat.

Basis for effect:

mortality (fish)

Remarks on result:

other: 95% CL not available

Duration:

96 h

Dose descriptor:

LC0

Effect conc.:

15.78 mg/L

Nominal / measured:

meas. (geom. mean)

Conc. based on:

test mat.

Basis for effect:

mortality (fish)

Duration:

96 h

Dose descriptor:

LC100

Effect conc.:

25.56 mg/L

Nominal / measured:

meas. (geom. mean)

Conc. based on:

test mat.

Basis for effect:

mortality (fish)

Details on results:

- Mortality of control: The mortalities of the controls were less than 10% at the end of the test.
- Abnormal responses: No abnormal response was observed in blank control. Observed abnormal responses during the test were reduced activity, overturn and loss of equilibrium and immobilized at the bottom; lying on the bottom; at the surface; swimming at the bottom; spasmodic swimming.
Observed abnormal responses are shown in Table 2.

Results with reference substance (positive control):

The test result of Zebra fish (Brachydanio rerio) sensitivity test revealed that 24h-LC50 of the reference substance K2Cr2O7 was 307 mg/L with 95% confidence limits (L95) of 273 mg/L - 346 mg/L, within the range of 200 mg/L-400 mg/L. See Overall remarks and attachments.

Sublethal observations / clinical signs:

Concentration of test substance in test solution:

The actual concentration of test substance in blank control were all <0.01647mg/L (LOD) at 0h, 96h and 24h, 48h, 72h (before and after the renewal) after the start of the test. Analysis of the test solutions with concentration of 6.87 mg/L, 11.0 mg/L, 17.6 mg/L, 28.1 mg/L and 45.0 mg/L at 0h, 96h and 24h, 48h, 72h (before and after the renewal) after the start of the test. The measured concentrations of test substance in the test solution at the start of the exposure and after the renewal ranged from 5.898 mg/L to 42.72 mg/L (86% to 95% of the setting concentration of test substance). That before the renewal and at the end of the exposure ranged from 5.484 mg/L to 38.14 mg/L (80% to 85% of the setting concentration of test substance). During the test, the actual concentration of each exposure level can be maintained within 80%-120% of the setting concentration of test substance.

Table 1 : Cumulative mortality (%)

Settingconcentration (mg/L)	Number of test fish	Cumulative mortality（%）
		3h	24h	48h	72h	96h
Blank control	21	0	0	0	0	0
6.87	21	0	0	0	0	0
11.0	21	0	0	0	0	0
17.6	21	0	0	0	0	0
28.1	21	48	100	100	100	100
45.0	21	100	100	100	100	100

 

Table 2 : Observed abnormal response

Settingconcentration (mg/L)	Rep- licate	Result of observation (Left column: Number of affected fish / Total survival number, Right column: Symptom detail）
		3h		24h		48h		72h		96h
Blank control	1	0/7	N	0/7	N	0/7	N	0/7	N	0/7	N
	2	0/7	N	0/7	N	0/7	N	0/7	N	0/7	N
	3	0/7	N	0/7	N	0/7	N	0/7	N	0/7	N
6.87	1	0/7	N	0/7	N	0/7	N	0/7	N	0/7	N
	2	0/7	N	0/7	N	0/7	N	0/7	N	0/7	N
	3	0/7	N	0/7	N	0/7	N	0/7	N	0/7	N
11.0	1	0/7	N	0/7	N	0/7	N	7/7	IMM(4) RA(3)	7/7	IMM(2) RA(5)
	2	0/7	N	0/7	N	0/7	N	7/7	IMM(7)	7/7	IMM(6) RA(1)
	3	0/7	N	0/7	N	0/6	N	7/7	IMM(3) RA(4)	7/7	IMM(2) RA(5)
17.6	1	7/7	IMM(1) RA(6)	7/7	IMM(7)	7/7	IMM(5) OLE(2)	7/7	IMM(5) OLE(2)	7/7	IMM(6) OLE(1)
	2	7/7	IMM(2) RA(5)	7/7	IMM(7)	7/7	IMM(7)	7/7	IMM(7)	7/7	IMM (7)
	3	7/7	OLE(1) IMM(2) RA(4)	7/7	IMM(7)	7/7	IMM(6) OLE(1)	7/7	IMM(5) OLE(2)	7/7	IMM(5) OLE(2)
28.1	1	5/5	LB(5)	—	—	—	—	—	—	—	—
	2	2/2	LB(2)	—	—	—	—	—	—	—	—
	3	4/4	SS(2) LB(2)	—	—	—	—	—	—	—	—
45.0	1	—	—	—	—	—	—	—	—	—	—
	2	—	—	—	—	—	—	—	—	—	—
	3	—	—	—	—	—	—	—	—	—	—

Note: “N”— Normal; “—”: All test fish died;“RA”: Reduced activity; “OLE”: overturn andloss of equilibrium;“IMM”:Immobilized at the bottom; “LB”:lying on the bottom; “AS”:at the surface；“AB”swimming at the bottom；“SS”:spasmodic swimming.

Validity criteria fulfilled:

yes

Remarks:

0% mortality in the blank control at the end of the test and dissolved oxygen concentrations were kept at least 60 percent of the air saturation value.

Conclusions:

Under the conditions of the test, the 96h-LC50 of test substance (based on the actual concentration) was 20.1 mg/L for Zebra fish (Brachydanio rerio) and the 95% confidence limit is not available. As no dose-response curve was observed, the LC50 value was determined as a geometric mean of LC0 and LC100.
The highest concentration causing 0% mortality at 96h (LC0) was 15.78 mg/L (based on geometric mean measured concentration of 17.6 mg/L test group).
The lowest concentration causing 100% mortality at 96h (LC100) was 25.56 mg/L (based on geometric mean measured concentration of 28.1 mg/L test group).

Executive summary:

The acute toxicity of the test substance to Danio rerio was assessed in a study following three guidelines : (1) State Environmental Protection Administration of China, The Guidelines for the Testing of Chemicals, 203 Fish Acute toxicity test, China Environmental Science Press, 2004 ; (2) OECD Guidelines for the Testing of Chemicals, 203 Fish Acute Toxicity Test. Paris: OECD, Adopted 17 July 1992 and (3) State Environmental Protection Administration of China, The Guidelines for the test of chemical (HJ/T 153-2004), Beijing, China Environmental Science Press, 2004.

The following five concentrations were tested : 6,87 mg/l, 11,0 mg/L, 17,6 mg/L, 28,1 mg/L and 45,0 mg/L (measured concentration of stock solution, factor 1.6).

10 fish were randomly selected and measured (at study initiation), the average and standard deviation of the population from which the fish were selected were 26.04 mm ± 1.71 mm total length. They were also weighed, the average and standard deviation were 0.186 g ± 0.053 g.

The test was performed at 23 +/- 2°C, with 7 fish / test vessel and 3 replicates were used per test group.

The recordred 96h-LC50 is 20.1 mg/L, based on measured concentrations.

This study is considered reliable with restrictions. No dose-response curve was observed. The LC50 value was determined as a geometric mean of LC0 and LC100. In addition, test fish was a bit bigger than recommended : 2,6 +/- 1,7 cm (recommended 2 +/- 1 cm)

Description of key information

Chinese 203 Fish Acute toxicity test (2004) and OECD 203 Fish Acute Toxicity Test (1992), Guangdong detection center of microbiology, GLP, 2012:

96h-LC50 (Danio rerio) = 20.1 mg/L (based on measured concentrations)

Key value for chemical safety assessment

Fresh water fish

Fresh water fish

Effect concentration:

20.1 mg/L

Additional information

The acute toxicity of the test substance to Danio rerio was assessed in a study following three guidelines : (1) State Environmental Protection Administration of China, The Guidelines for the Testing of Chemicals, 203 Fish Acute toxicity test, China Environmental Science Press, 2004 ; (2) OECD Guidelines for the Testing of Chemicals, 203 Fish Acute Toxicity Test. Paris: OECD, Adopted 17 July 1992 and (3) State Environmental Protection Administration of China, The Guidelines for the test of chemical (HJ/T 153-2004), Beijing, China Environmental Science Press, 2004. The test was performed according to GLP and validity criteria were fulfilled. However, this study was considered reliable with restrictions as no dose-response curve was observed. The LC50 value was determined as a geometric mean of LC0 and LC100. In addition, test fish was a bit bigger than recommended : 2,6 +/- 1,7 cm (recommended 2 +/- 1 cm).

The following five concentrations were tested : 6,87 mg/l, 11,0 mg/L, 17,6 mg/L, 28,1 mg/L and 45,0 mg/L (measured concentration of stock solution, factor 1.6).

10 fish were randomly selected and measured (at study initiation), the average and standard deviation of the population from which the fish were selected were 26.04 mm ± 1.71 mm total length. They were also weighed, the average and standard deviation were 0.186 g ± 0.053 g. The test was performed at 23 +/- 2°C, with 7 fish / test vessel and 3 replicates were used per test group.

The recorded 96h-LC50 is 20.1 mg/L, based on measured concentrations.