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Ecotoxicological information

Toxicity to aquatic algae and cyanobacteria

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Administrative data

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Endpoint:
toxicity to aquatic algae and cyanobacteria
Type of information:
experimental study
Adequacy of study:
key study
Study period:
not detailed
Reliability:
1 (reliable without restriction)
Rationale for reliability incl. deficiencies:
test procedure in accordance with national standard methods
Reason / purpose for cross-reference:
reference to same study
Qualifier:
according to guideline
Guideline:
other: EPA-600/9-78-018
GLP compliance:
yes
Specific details on test material used for the study:
Bisphenol A with a purity of 99.93 %.
Analytical monitoring:
yes
Vehicle:
no
Test organisms (species):
Raphidocelis subcapitata (previous names: Pseudokirchneriella subcapitata, Selenastrum capricornutum)
Details on test organisms:
The test alga, Selenastrum capricornutum Printz, is a freshwater, unicellular, green algae (Chlorophaceae) of the order Chlorococcales. The culture was obtained from the Environmental Research Laboratory - U.S. EPA, Corvallis, Oregon and received December31, 1979. The algal stock culture was maintained by means of weekly aseptic transfers to fresh sterile algal assay medium. Old cultures were diluted approximately 1:400 (e.g. 0.5 mL of old culture in 200 mL fresh medium).
Test type:
static
Water media type:
freshwater
Limit test:
no
Total exposure duration:
96 h
Hardness:
no data provided
Test temperature:
no data provided
pH:
no data provided
Dissolved oxygen:
no data provided
Salinity:
no data provided
Conductivity:
no data provided
Nominal and measured concentrations:
Nominal Test Concentration of 0 (control), 0.78, 1.30, 2.16, 3.60, 6.00, and 10.0 mg/L, corresponded to analyzed test concentrations of ND (0.05)+/-0.00, 0.69+/-0.02, 1.17+/-0.10, 1.99+/-0.16, 3.41+/-0.13, 5.86+/-0.11, and 9.74+/-0.06 mg/L.
Reference substance (positive control):
no
Key result
Duration:
96 h
Dose descriptor:
EC10
Effect conc.:
1.41 mg/L
Basis for effect:
biomass
Key result
Duration:
96 h
Dose descriptor:
EC50
Effect conc.:
3.1 mg/L
Basis for effect:
other: cell volume
Key result
Duration:
96 h
Dose descriptor:
EC50
Effect conc.:
2.73 mg/L
Basis for effect:
biomass
Remarks:
cell count
Key result
Duration:
96 h
Dose descriptor:
EC10
Effect conc.:
1.36 mg/L
Basis for effect:
biomass
Remarks:
cell count and cell volume
Remarks on result:
other:
Remarks:
The EC10 was delineated in the EU Risk Assessment Report of BPA, 2008, not in the study report.
Results with reference substance (positive control):
No positive control used.
Reported statistics and error estimates:
Percent inhibition values and log10 of the concentration were used to estimate the EC50 concentration values, using non-linear interpolation between the two concentrations which bracketed the 50 percent inhibition response. Unweighted replicate means of 96 hour cell count and total cell volume data were evaluated by one-way analysis of variance procedure. The Dunnett's one-tailed t-test was used to compare treatment means to control means at a = 0.05.

The 96-hr EC50 for cell count was 2.73 mg/L and the 96-hr EC50 for total cell volume was 3.10 mg/L.

Validity criteria fulfilled:
yes
Conclusions:
The study followed a validated test method and was GLP documented. The toxicity of Bisphenol A to, the freshwater green algae, Selenastrum capricornutum Printz was determined. The EC50 values are based on 50% inhibition of' cell count and total cell volume compared to the controls. The 96 hour EC50 values based on cell count and total cell volume using analyzed test concentrations were 2.73 mg/L and 3.10 mg/L. Bisphenol A, respectively. Algal growth was inhibited at concentrations of 1.99 mg/L and higher. Therefore, this study is considered relevant and reliable for regulatory purposes. Based on the dose-response data reported by the authors, a robust EC10 was calculated using the Trap software, which was determined to be 1.41 mg/l. This EC10 is considered to be valid and robust and was thus used for evaluation under the SSD approach. EC10 values were derived in accordance with Moermond et al. (2016) and ECHA (2008).
Executive summary:

The acute toxicity to the planktonic alga Selenastrum capricornutum was determined in a 4 day growth test using a validated test system and GLP documentation. Nominal concentrations ranged from 0 to 1000 mg/L and as BPA concentrations were stable results are expressed as nominal concentrations. Flasks containing 100 mg/L and greater were excluded from the assessment due to a complete loss of algae in the test system. Cell number in the 1.0, 2.2, 4.6, 10, 22, and 46 mg/L nominal test concentration flasks, reported as a % of mean control cell number on day 4 were 91, 50, 26, 1, 0, and 0, respectively. The 96 hr EC50, based on cell counts on day 4 was calculated to be 2.5 mg/L (95 % confidence limits 2.2 - 2.8 mg/L). The study is considered sufficiently valid to be ranked as Klimisch 1 and adequate to be used for regulatory purposes. Based on the dose-response data reported by the authors, a robust EC10 was calculated using the Trap software, which was determined to be 1.41 mg/l. This EC10 is considered to be valid and robust and was thus used for evaluation under the SSD approach. For the purpose of developing a SSD in support of PNEC setting, the toxicity values for the most sensitive endpoint are selected. The toxicity values are determined on the basis of the geometric mean if more than one value for the same endpoint and exposure time is available. EC10 values were derived in accordance with Moermond et al. (2016) and ECHA (2008).

Endpoint:
toxicity to aquatic algae and cyanobacteria
Type of information:
experimental study
Adequacy of study:
key study
Reliability:
1 (reliable without restriction)
Reason / purpose for cross-reference:
reference to same study
Qualifier:
according to guideline
Guideline:
other: EPA-560/6-82-002
GLP compliance:
yes
Analytical monitoring:
yes
Vehicle:
no
Test organisms (species):
Skeletonema costatum
Test type:
static
Water media type:
saltwater
Limit test:
no
Total exposure duration:
96 h
Reference substance (positive control):
no
Key result
Duration:
96 h
Dose descriptor:
EC50
Effect conc.:
1.1 mg/L
Nominal / measured:
meas. (arithm. mean)
Conc. based on:
act. ingr.
Basis for effect:
other: cell count and relative fluorescence (measure of chlorophyll a)
Remarks on result:
other: The EC50 in the original study was 1.0 mg/L. In the EU risk assessment report 2003 the data was statistically re-evaluated based on more appropriate statistical methods and the EC50 was determined to be 1.1 mg/L.
Duration:
96 h
Dose descriptor:
EC50
Effect conc.:
1.4 mg/L
Nominal / measured:
meas. (arithm. mean)
Conc. based on:
act. ingr.
Basis for effect:
other: chlorophyll A inhibition
Remarks on result:
other: The EC50 in the original study was 1.0 mg/L. In the EU risk assessment report 2003 the data was statistically re-evaluated based on more appropriate statistical methods and the EC50 was determined to be 1.4 mg/L.
Key result
Duration:
96 h
Dose descriptor:
EC10
Effect conc.:
0.4 mg/L
Nominal / measured:
meas. (arithm. mean)
Conc. based on:
act. ingr.
Basis for effect:
biomass
Remarks:
cell number
Remarks on result:
other: In the EU risk assessment report 2003 the data was statistically re-evaluated and additionally an EC10 of 0.4 mg/L was derived.

Nominal Test Concentrations were Control, 0.72, 1.1, 1.9, 3.2, 5.4, 9.0, and 15 mg/L which corresponded to analysed test concentrations of 0.66, 1.1, 1.6, 2.8, 5.2, 8.8, and 15 mg/L. Test concentrations were found to be stable over the course of the study and the blind tests showed that satisfactory control over the accuracy and precision of the Bisphenol A concentration measurements were achieved. Chlorophyll A concentration and cell counts were significantly reduced at all Bisphenol A test concentrations. Based on the analysed concentration and the observed effects, the EC50s for cell count inhibition and chlorophyll A inhibition, calculated by probit analysis, were 1.1 mg/L and 1.4 mg/L, respectively.

Validity criteria fulfilled:
yes
Conclusions:
Based on the analysed concentration and the observed effects, the EC50s for cell count inhibition and chlorophyll A inhibition, calculated by probit analysis, were 1.1 mg/L and 1.4 mg/L, respectively.
Executive summary:

The marine algae Skeletonema costatum was tested in a 96-h test with nominal concentrations of Bisphenol A of 0, 0.72, 1.1, 1.9, 3.2, 5.4, 9.0, and 15 mg/L. These corresponded to analysed test concentrations of 0, 0.66, 1.1, 1.6, 2.8, 5.2, 8.8, and 15 mg/L, respectively. At the end of the exposure, chlorophyll A concentration and cell counts were significantly reduced at all Bisphenol A test concentrations. Based on the analysed concentration and the observed effects, the EC50s for cell count inhibition and chlorophyll A inhibition, calculated by probit analysis, were 1.1 mg/L and 1.4 mg/L, respectively.

Description of key information

There are two key studies for exposure of algae to Bisphenol A (both rated Klimisch 1, i.e., reliable without restriction) for both compartments, freshwater and marine water. The key study by Alexander et al. (1985; published in Alexander 1988) which was conducted in accordance with EPA-600/9-78-018 test method reported the 96-h EC50 value of 2.73 mg/L for the freshwater algae Pseudokirchneriella subcapitata based on cell count. Furthermore, an EC10 of 1.36 mg/L was derived for this study. This was further cemented by data from van Sprang et al. (2022), who calculated an EC10 value of 1.41 mg/L for the data set as reported by Alexander et al based on the dose-response data.  The other key study conducted by Suprenant (1985; published as Alexander et al., 1988) was performed on marine Skeletonema costatum following test method EPA 560/6-82-002. At the end of the exposure, chlorophyll A concentration and cell counts were significantly reduced at all Bisphenol A test concentrations. Based on the analysed concentration and the observed effects, the EC50s for cell count and chlorophyll A content, calculated by probit analysis (re-analysed by the UK rapporteur in the EU Risk Assessment update in 2008 in accordance with the OECD Guideline), were 1.1 mg/L and 1.4 mg/L, respectively. The 96-hr EC10 for the most sensitive endpoint, cell count, was calculated by probit analysis to be 0.4 mg/L.


 


Furthermore, there are two Klimisch 2 studies with algae, which support the Klimisch 1 studies.



  • Stephenson, 1983, reports an EC50 of 2.5 mg/L based on growth rate in a study conducted according to test method E09-04 with the fresh water species Pseudokirchneriella subcapitata.

  • Liu et al., 2010, report an EC50 of 3.73 mg/L based on growth rate in a study performed similar to OECD 201 with the marine species Navicula incerta.


 


Concluding, the lowest EC50 of 2.73 mg/L and EC10 of 1.41 mg/L for fresh water algae was reported in Alexander et al., 1985. The lowest EC50 of 1.1 mg/L and the lowest EC10 of 0.4 mg/L for marine water was reported in Suprenant, 1985.

Key value for chemical safety assessment

EC50 for freshwater algae:
2.73 mg/L
EC50 for marine water algae:
1.1 mg/L
EC10 or NOEC for freshwater algae:
1.41 mg/L
EC10 or NOEC for marine water algae:
0.4 mg/L

Additional information

There are two key studies for exposure of algae to Bisphenol A (both assigned to Reliability 1, i.e. reliable without restriction) for both compartments, freshwater and marine water.

The freshwater algae, Pseudokirchneriella subcapitata (formerly known as Selenastrum capricornutum), was exposed for 96 h to nominal concentrations of Bisphenol A of 0, 0.78, 1.30, 2.16, 3.6, 6.0, and 10.0 mg/L (Alexander et al., 1985; published in Alexander et al., 1988). These corresponded to mean measured concentrations of 0, 0.69, 1.17, 1.99, 3.41, 5.86, and 9.74 mg/L, respectively. The 96-h EC50 for cell count was 2.73 mg/L and the 96-h EC50 for total cell volume was 3.10 mg/L. The EC10 for the most sensitive endpoint, cell count, was calculated by probit analysis to be 1.36 mg/L.

The marine diatom, Skeletonema costatum, was exposed for 96 h to nominal concentrations of Bisphenol A of 0, 0.72, 1.1, 1.9, 3.2, 5.4, 9.0, and 15 mg/L (Suprenant, 1985c; published as Alexander, 1988). These corresponded to analysed test concentrations of 0, 0.66, 1.1, 1.6, 2.8, 5.2, 8.8, and 15 mg/L, respectively. At the end of the exposure, chlorophyll A concentration and cell counts were significantly reduced at all Bisphenol A test concentrations. Based on the analysed concentration and the observed effects, the EC50s for cell count and chlorophyll A content, calculated by probit analysis, were 1.1 mg/L and 1.4 mg/L, respectively. The 96-hr EC10 for the most sensitive endpoint, cell count, was calculated by probit analysis to be 0.4 mg/L.

Two supporting studies rated as Klimisch 2 (reliable with restrictions) were identified (Stephenson et al., 1983; Liu et al, 2010). The study by Stephenson et al. was conducted in accordance with E09-04 test method with freshwater algae Pseudokirchneriella subcapitata and determined an EC50 of 2.5 mg/L. Liu et al. used a method similar to OECD 201 on the marine Navicula incerta and determined an EC50 of 3.73 mg/L based on growth rate.