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Ecotoxicological information

Toxicity to aquatic algae and cyanobacteria

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Link to relevant study record(s)

Reference
Endpoint:
toxicity to aquatic algae and cyanobacteria
Type of information:
experimental study
Adequacy of study:
key study
Reliability:
1 (reliable without restriction)
Rationale for reliability incl. deficiencies:
guideline study
Remarks:
guideline study conducted under GLP with chemical analysis. Raw data available.
Qualifier:
equivalent or similar to guideline
Guideline:
OECD Guideline 201 (Alga, Growth Inhibition Test)
Principles of method if other than guideline:
The test was conducted according to the notice of the Director General of the Pharmaceutical and Food Safety Bureau, Director General of the Manufacturing Industries Bureau, and Director General of the Environmental Policy Bureau, Ministry of the Environment, 'Partial Revision of 'Methods for Testing New Chemical Substances' (Pharmaceutical and Food Safety Bureau, MHLW No. 1120001 dated November 20, 2006, Manufacturing Industries Bureau, METI No. 2 dated November 13, 2006, and Environmental Policy Bureau, MOE No. 061120001.
GLP compliance:
yes
Analytical monitoring:
yes
Details on sampling:
- Concentrations: 0.005, 0.01, 0.02, 0.04, 0.1, 0.25, 0.63, 1.6, and 3.9 mg/L (nominal)
- Sampling method: Immediately after sample collection (e.g. micropipette) phosphate solution was added
- Sample storage conditions before analysis: the sample was stored in a refrigerator at 4°C until analysis
Vehicle:
no
Details on test solutions:
PREPARATION AND APPLICATION OF TEST SOLUTION (especially for difficult test substances)
- Method: Stock solutions were prepared. The desired test concentrations were obtained by pipetting a fixed volume of the stock solution into the test vessels containing the OECD culture medium.
- Chemical name of vehicle (organic solvent, emulsifier or dispersant): no vehicle used
Test organisms (species):
Raphidocelis subcapitata (previous names: Pseudokirchneriella subcapitata, Selenastrum capricornutum)
Details on test organisms:
TEST ORGANISM
- Common name: green algae
- Strain: ATCC 22662
- Source (laboratory, culture collection): American Type Culture Collection
- Age of inoculum (at test initiation): 3 d
- Method of cultivation: after obtaining the culture, a subculture was cultivated aseptically using a slant medium with ATCC culture medium 625. A pre-preculture was prepared by transplanting the test algae from the slant medium to the ATCC culture medium used for subculture and was cultured under stationary conditions for 8 d. A preculture was prepared by culturing the test algae in OECD medium under environmental conditions similar to those for testing for 3 d. The algae showed exponential growth.

ACCLIMATION
- Acclimation period: 3 d
- Culturing media and conditions (same as test or not): yes
- Any deformed or abnormal cells observed: not reported
Test type:
static
Water media type:
freshwater
Limit test:
no
Total exposure duration:
72 h
Hardness:
no data
Test temperature:
22.6-23.0°C
pH:
at test start: 7.3-7.7
at test end: 7.8-8.1
Dissolved oxygen:
no data
Salinity:
no data (fresh water test)
Nominal and measured concentrations:
nominal: 0.040, 0.10, 0.25, 0.63, 1.6, and 3.9 mg/L
measured initial: 0.0033, 0.10, 0.240, 0.61, 1.5, and 3.9 mg/L
mean measured (0-72 h geom. mean): 0.0057, 0.018, 0.056, 0.11, 0.24, 0.45 mg/L

additional test:
nominal (additional test): 0.005, 0.01, and 0.02 mg/L
measured initial: 0.0048, 0.0085, and 0.019 mg/L
mean measured (0-72 h geom. mean; additional test: 0.00085, 0.0015, and 0.0033 mg/L
Details on test conditions:
TEST SYSTEM
- Test vessel: 300 mL Erlenmeyer flask
- Type (delete if not applicable): open (the test vessels were capped with a breathable silicon plug)
- Material, size, headspace, fill volume: glass/300 mL/'200 mL'/100 mL
- Aeration: not reported
- Type of flow-through (e.g. peristaltic or proportional diluter): test conducted under static conditions
- Initial cells density: 0.5 x 10E4 cells/mL
- Control end cells density: 896256 cells/mL (additional test: 900943 cells/mL)
- No. of vessels per concentration (replicates): 3
- No. of vessels per control (replicates): 6

GROWTH MEDIUM
- Standard medium used: yes

TEST MEDIUM / WATER PARAMETERS
- Source/preparation of dilution water: OECD culture medium was used
- Culture medium different from test medium: preculture and culture used for the test were cultivated in OECD medium
- Intervals of water quality measurement: Temperature and light flux density inside the incubator were measured at the beginning and every 24 h until the end of exposure. The pH of the test solutions were measured at the beginning and at the end of exposure.


OTHER TEST CONDITIONS
- Sterile test conditions: yes
- Adjustment of pH: no
- Photoperiod: continuous lighting
- Light intensity and quality: 60-90 µmol/m² x s (fluorescent lamp)

EFFECT PARAMETERS MEASURED (with observation intervals if applicable) : growth rate, biomass
- Determination of cell concentrations: cell count was measured via a particle counter (Beckman Coulter)

TEST CONCENTRATIONS
- Spacing factor for test concentrations: 2.5 (additional test: 2)
- Range finding study
- Test concentrations: 0.1, 0.3, 0.9, and 2.7 mg/L (additional range finding test: 0.025, 0.050 mg/L
- Results used to determine the conditions for the definitive study: yes
Reference substance (positive control):
yes
Remarks:
potassium dichromate
Duration:
72 h
Dose descriptor:
EC50
Effect conc.:
0.33 mg/L
Nominal / measured:
meas. (initial)
Conc. based on:
test mat.
Basis for effect:
growth rate
Remarks on result:
other: 95% c.i.: 0.243-0.452 mg/L; recalculated
Duration:
72 h
Dose descriptor:
EC10
Effect conc.:
0.034 mg/L
Nominal / measured:
meas. (initial)
Conc. based on:
test mat.
Basis for effect:
growth rate
Remarks on result:
other: 95% c.i.: 0.015-0.057
Duration:
72 h
Dose descriptor:
NOEC
Effect conc.:
0.019 mg/L
Nominal / measured:
meas. (initial)
Conc. based on:
test mat.
Basis for effect:
growth rate
Remarks on result:
other: recalculated
Duration:
72 h
Dose descriptor:
EC50
Effect conc.:
0.053 mg/L
Nominal / measured:
meas. (geom. mean)
Conc. based on:
test mat.
Basis for effect:
growth rate
Remarks on result:
other: 95% c.i.: 0.045-0.061 mg/L
Duration:
72 h
Dose descriptor:
NOEC
Effect conc.:
0.002 mg/L
Nominal / measured:
meas. (geom. mean)
Conc. based on:
test mat.
Basis for effect:
growth rate
Duration:
72 h
Dose descriptor:
EC50
Effect conc.:
0.041 mg/L
Nominal / measured:
meas. (initial)
Conc. based on:
test mat.
Basis for effect:
other: yield
Remarks on result:
other: 95% c.i.: 0.025-0.074 mg/L; recalculated
Duration:
72 h
Dose descriptor:
EC10
Effect conc.:
0.009 mg/L
Nominal / measured:
meas. (initial)
Conc. based on:
test mat.
Basis for effect:
other: yield
Remarks on result:
other: 95% c.i.: 0.0016-0.016 mg/L
Duration:
72 h
Dose descriptor:
NOEC
Effect conc.:
0.009 mg/L
Nominal / measured:
meas. (initial)
Conc. based on:
test mat.
Basis for effect:
other: yield
Remarks on result:
other: recalculated
Duration:
72 h
Dose descriptor:
EC50
Effect conc.:
< 0.033 mg/L
Nominal / measured:
meas. (geom. mean)
Conc. based on:
test mat.
Basis for effect:
other: yield
Duration:
72 h
Dose descriptor:
NOEC
Effect conc.:
< 0.033 mg/L
Nominal / measured:
meas. (geom. mean)
Conc. based on:
test mat.
Basis for effect:
other: yield
Results with reference substance (positive control):
ErC50 (0-72 h)=0.89 mg/L
Reported statistics and error estimates:
The growth inhibition rate was calculated by comparison with the growth rate (velocity method) which was used to calculate the ErC50 using the probit method (EcoTox-Statics was used for calculation). The NOEC was calculated using the EcoTox-Statics, the Bartlett method, ANOVA, the Dunett method, and the Kruskal-Wallis rank sum test. The recalculation was done using probit and Welch-t-test (yield) and Williams Multiple t-test (growth rate) using the programme ToxRat V. 2.10.

The toxicity of hydroquinone to algae was investigated in a GLP-study conducted similar to OECD Guideline 201 (Alga, Growth Inhibition Test). The authors determined the 72 h NOEC and EC50 (growth rate each) using mean measured concentrations to be 0.0015 mg/L and 0.053 mg/L, respectively. However, hydroquinone is known to be fast degraded to several toxic degradation products depending on the environmental conditions like oxygen, pH and light (e.g. p-benzoquinone). At test start the initial concentrations were between 82 and 100% of the nominal values and were between 1.4 and 5.2% at test end. The degradation products were not analyses in this study. Provided that the formed degradation products are also of high ecotoxicity the evaluation of the test based on the concentration of the parent compound alone may overestimate its toxicity. Provided further that the total amount of toxic equivalents is stable during the exposure period the initial concentration is regarded as relevant concentration. Therefore, the algal test was re-evaluated based on the measured initial concentrations. The effects on algae were tested in two separate tests. The cell numbers of the controls of the two tests were comparable and therefore the two tests were combined for the re-evaluation.

The 72 h ErC50 and NOEC (growth rate) were determined to be 0.330 mg/L and 0.019 mg/L (based on recalculated measured initial concentrations). The 72 h EyC50 and NOEC (yield) were determined to be 0.041 mg/L and 0.0085 mg/L (based on recalculated measured initial concentrations).

The validity criteria according to OECD Guideline 201 were fulfilled (biomass increase: > factor 16; mean coefficient of variation section-by section specific growth rates: <=35%, and coefficient of variation of average specific growth rates during the whole test period: <=7%).

Validity criteria fulfilled:
yes
Conclusions:
The toxicity of hydroquinone to algae was investigated in a GLP-study conducted equivalent to OECD Guideline 201 (Alga, Growth Inhibition Test). The 72 h ErC50 and NOEC (growth rate) were determined to be 0.330 mg/L and 0.019 mg/L (based on recalculated measured initial concentrations). The 72 h EyC50 and NOEC (yield) were determined to be 0.041 mg/L and 0.0085 mg/L (based on recalculated measured initial concentrations).
Executive summary:

The toxicity of hydroquinone to algae was investigated in a GLP-study conducted equivalent to OECD Guideline 201 (Alga, Growth Inhibition Test). Cultures of Pseudokirchneriella subcapitata were exposed to nominal concentrations of 0.005, 0.01, 0.02, 0.040, 0.10, 0.25, 0.63, 1.6, and 3.9 mg/L in two separate tests. The test substance concentrations were monitored by HPLC/UV. At test start the initial concentrations were between 82 and 100% of the nominal values and were between 1.4 and 5.2% at test end. The mean measured concentrations (0-72 h geom. mean) were 0.0057, 0.018, 0.056, 0.11, 0.24, 0.45 mg/L and 0.00085, 0.0015, and 0.0033 mg/L (additional test) for 72 h under static conditions. As a result of measuring the concentration after 72 h, the 0.04 mg/L test concentration was below the lower LOQ. Therefore, the decrease of concentration with time was determined using an initial nominal concentration of 0.63 mg/L. Based on the decay curve, concentrations below the LOQ were calculated. The 72 h NOEC and EC50 (growth rate each) were determined to be 0.0015 mg/L and 0.053 mg/L, respectively.

However, hydroquinone is known to be fast degraded to several toxic degradation products depending on the environmental conditions like oxygen, pH and light (e.g. p-benzoquinone). The degradation products were not analyses in this study. Provided that the formed degradation products are also of high ecotoxicity the evaluation of the test based on the concentration of the parent compound alone may overestimate its toxicity. Provided further that the total amount of toxic equivalents is stable during the exposure period the initial concentration is regarded as relevant concentration. Therefore, the algal test was re-evaluated based on the measured initial concentrations. The 72 h ErC50 and NOEC (growth rate) were determined to be 0.330 mg/L and 0.019 mg/L (based on recalculated measured initial concentrations). The 72 h EyC50 and NOEC (yield) were determined to be 0.041 mg/L and 0.0085 mg/L (based on recalculated measured initial concentrations). The validity criteria according to OECD Guideline 201 were fulfilled and this study is regarded as reliable without restriction.

For the hazard and risk assessment the effect concentrations for growth rate should be used: ErC50 and NOEC (growth rate) of 0.330 mg/L and 0.019 mg/L, respectively.

Description of key information

The toxicity of hydroquinone to algae was investigated in a study conducted equivalent or similar to OECD Guideline 201 (Alga, Growth Inhibition Test). The 72 h ErC50 and NOEC (growth rate) were determined to be 0.330 mg/L and 0.019 mg/L (based on recalculated measured initial concentrations). The 72 h EyC50 and NOEC (yield) were determined to be 0.041 mg/L and 0.0085 mg/L (based on recalculated measured initial concentrations).

Key value for chemical safety assessment

EC50 for freshwater algae:
0.33 mg/L
EC10 or NOEC for freshwater algae:
0.019 mg/L

Additional information

The toxicity of hydroquinone to algae was investigated in a GLP-study conducted equivalent to OECD Guideline 201 (Alga, Growth Inhibition Test). Cultures ofPseudokirchnerella subcapitatawere exposed to nominal concentrations of 0.005, 0.01, 0.02, 0.040, 0.10, 0.25, 0.63, 1.6, and 3.9 mg/L in two separate tests. The test substance concentrations were monitored by HPLC/UV. At test start the initial concentrations were between 82 and 100% of the nominal values and were between 1.4 and 5.2% at test end. The mean measured concentrations (0-72 h geom. mean) were 0.0057, 0.018, 0.056, 0.11, 0.24, 0.45 mg/L and 0.00085, 0.0015, and 0.0033 mg/L (additional test) for 72 h under static conditions. As a result of measuring the concentration after 72 h, the 0.04 mg/L test concentration was below the lower LOQ. Therefore, the decrease of concentration with time was determined using an initial nominal concentration of 0.63 mg/L. Based on the decay curve, concentrations below the LOQ were calculated. The 72 h NOEC and EC50 (growth rate each) were determined to be 0.0015 mg/L and 0.053 mg/L, respectively.

However, hydroquinone is known to be fast degraded to several toxic degradation products depending on the environmental conditions like oxygen, pH and light (e.g. p-benzoquinone). The degradation products were not analyses in this study. Provided that the formed degradation products are also of high ecotoxicity the evaluation of the test based on the concentration of the parent compound alone may overestimate its toxicity. Provided further that the total amount of toxic equivalents is stable during the exposure period the initial concentration is regarded as relevant concentration. Therefore, the algal test was re-evaluated based on the measured initial concentrations. The 72 h ErC50 and NOEC (growth rate) were determined to be 0.330 mg/L and 0.019 mg/L (based on recalculated measured initial concentrations). The 72 h EyC50 and NOEC (yield) were determined to be 0.041 mg/L and 0.0085 mg/L (based on recalculated measured initial concentrations). The validity criteria according to OECD Guideline 201 were fulfilled and this study is regarded as reliable without restriction.

For the hazard and risk assessment the effect concentrations for growth rate should be used: ErC50 and NOEC (growth rate) of 0.330 mg/L and 0.019 mg/L, respectively.