Glycollic acid

Toxicity to aquatic algae and cyanobacteria

Link to relevant study record(s)

Reference

Reference 1

Endpoint:

toxicity to aquatic algae and cyanobacteria

Type of information:

experimental study

Adequacy of study:

supporting study

Reliability:

2 (reliable with restrictions)

Rationale for reliability incl. deficiencies:

test procedure in accordance with national standard methods with acceptable restrictions

Qualifier:

according to guideline

Guideline:

ISO 10253 (Water quality - Marine Algal Growth Inhibition Test with Skeletonema costatum and Phaeodactylum tricornutum)

Deviations:

no

GLP compliance:

yes

Analytical monitoring:

no

Vehicle:

no

Test organisms (species):

Skeletonema costatum

Test type:

static

Water media type:

saltwater

Limit test:

no

Total exposure duration:

72 h

Reference substance (positive control):

yes

Remarks:

3,5-dichlorophenol

Duration:

72 h

Dose descriptor:

NOEC

Effect conc.:

99.8 mg/L

Nominal / measured:

nominal

Conc. based on:

test mat.

Basis for effect:

growth rate

Remarks on result:

other: NOEC equivalent to 69.9 mg a.s./L

Duration:

72 h

Dose descriptor:

EC50

Effect conc.:

190 mg/L

Nominal / measured:

nominal

Conc. based on:

test mat.

Basis for effect:

growth rate

Remarks on result:

other: EC50 equivalent to 133 mg a.s./L

Validity criteria fulfilled:

yes

Conclusions:

Based on nominal exposure concentrations the 72 h EC50, based on growth rate, for Skeletonema costatum exposed to glycolic acid 70% solution, was 190 mg/L (equivalent to 133 mg a.s./L) the corresponding NOEC was 99.8 mg/L (equivalent to 69.9 mg a.s./L).

Executive summary:

The growth inhibition of the marine algae (Skeletonema costatum) by glycolic acid 70% solution was investigated according to ISO10253:1995 E. Based on nominal  exposure concentrations the 72 h EC50, for growth rate, of S. costatum exposed to glycolic acid 70% solution, was 190 mg/L (equivalent to 133 mg a.s./L) the corresponding NOEC was 99.8 mg/L (equivalent to 69.9 mg a.s./L).

Reference 2

Endpoint:

toxicity to aquatic algae and cyanobacteria

Type of information:

experimental study

Adequacy of study:

key study

Study period:

15 March to 27 May, 2021

Reliability:

1 (reliable without restriction)

Rationale for reliability incl. deficiencies:

guideline study

Qualifier:

according to guideline

Guideline:

OECD Guideline 201 (Freshwater Alga and Cyanobacteria, Growth Inhibition Test)

Version / remarks:

adopted March 23, 2006, corrected July 28, 2011

Deviations:

no

GLP compliance:

yes (incl. QA statement)

Analytical monitoring:

yes

Details on sampling:

Duplicate samples from the freshly prepared test media (without algae) of all test concentrations and from the control were taken at the start of the test. To determine test item stability under the test conditions and the maintenance of test item concentrations during the test period, duplicate samples from the test media of all test concentrations and the control (containing algae) were taken after 24- and 48-hours and at the end of the test (after the 72-hour test period) by pouring together the contents of the test beakers of each treatment. All samples remained undiluted until sample preparation. All samples were stored in a freezer (≤ - 20°C), protected from light until analysis was performed. Afterwards, the samples were again stored deep frozen.

Vehicle:

no

Details on test solutions:

The test medium of the nominal 100 mg/L test concentration was prepared by dissolving 96.4 mg test item into 964 mL test water by intense stirring for 20 minutes at room temperature. The pH of the highest test concentration was adjusted from 4.0 to 8.2 using 1 M NaOH. Adequate volumes of this test medium were diluted with test water to prepare the test media of the other desired test concentrations. The test media were prepared just before introduction of the algae (test initiation).

Test organisms (species):

Raphidocelis subcapitata (previous names: Pseudokirchneriella subcapitata, Selenastrum capricornutum)

Details on test organisms:

Species: Pseudokirchneriella subcapitata (KORSHIKOV), Strain No. 61.81 SAG;
Origin: The algae were originally supplied by the „Sammlung von Algenkulturen, Albrecht-von-Haller-Institut für Pflanzen-wissenschaften, Universität Göttingen", 37073 Göttingen, Germany;
Breeding Conditions: The algae were cultivated in the testing laboratory under standardised conditions according to the test guidelines.

Test type:

static

Water media type:

freshwater

Limit test:

no

Total exposure duration:

72 h

Remarks on exposure duration:

No further remarks

Post exposure observation period:

Not applicable

Hardness:

0.24 mmol/L (= 24 mg/L) as CaCO3

Test temperature:

21.6 to 21.8 °C

pH:

Control at test start: 8.0;
Control at test end: 9.0;
Test item treatments at test start: 8.0 to 8.2;
Test item treatments at test end: 8.9 to 9.1

Dissolved oxygen:

Not applicable

Salinity:

Not applicable

Conductivity:

Not reported

Nominal and measured concentrations:

Nominal: 0 (control), 100, 40, 16, 6.4 and 2.6 mg/L (pH adjusted)

Details on test conditions:

Tests were conducted in 50 mL Erlenmeyer flasks containing nominal 50 mL of test medium covered with sterile caps. All glassware was autoclaved prior to test start. Reconstituted water (OECD medium) was prepared with deionised water in which analytical grade salts were added at the appropriate nominal concentrations.

Pre-experiments were performed to determine a suitable concentration range and to establish suitable methods to prepare the test solutions.

The definitive test was started (0 hours) by inoculation of a biomass of nominal 5000 algal cells per mL test medium. Cells were taken from an exponentially growing pre-culture, set up 3 days prior to the test start under the same conditions as in the test. The test was performed with three replicates per test concentration and six replicates in the control. Test units were continuously stirred by magnetic stirrers. The flasks were incubated in a water bath, were placed in a random order and were repositioned each day to minimize differences in test conditions. Two additional replicates of each test item concentration and the control were set up for analytical dose verification after 24- and 48-hours. These additional replicates were incubated under the same conditions as described above. Additionally, one replicate of each test concentration and of the control was prepared without algae to provide a “blank” for the spectrophotometric measurements. The additional replicates were incubated under the same conditions as described above. The blank values were subtracted from the absorption measured in the samples containing algae to eliminate absorption caused by the test item.

Temperature was measured daily in an Erlenmeyer flask filled with water and incubated under the same conditions as the test flasks. The pH was measured in all test item concentrations and the control at the start and the end of the test. Test units were provided with continuous illumination, with light intensity measured once during the test at 6 positions distributed over the experimental area at the surface of the test media (mean light intensity was 5050 lux, and ranged from 4870 to 5210 lux).

The cell density on each observation time was determined by spectrophotometric measurement. Defined volumes of the algal suspensions from all replicates and from the blanks were sampled after 24-, 48- and 72-hours of exposure, and were not replaced. The algal cell densities were calculated by subtracting the absorption of the blanks, from each of the measured absorption of the test media (with algae). Based on the counted cell densities and the absorption from an algal suspension and its dilutions, a linear regression was performed for the calculation of the cell densities of the replicates during the test. To check for any effect of the test item on the morphology of the algal cells, at least one sample from all test concentrations and the control was taken after the 72-hour test period, and the shape of the treated algal cells compared to the control was examined microscopically.

Reference substance (positive control):

yes

Remarks:

Potassium dichromate (tested at least twice a year)

Duration:

72 h

Dose descriptor:

LOEC

Effect conc.:

> 100 mg/L

Nominal / measured:

nominal

Conc. based on:

test mat.

Basis for effect:

growth rate

Duration:

72 h

Dose descriptor:

NOEC

Effect conc.:

>= 100 mg/L

Nominal / measured:

nominal

Conc. based on:

test mat.

Basis for effect:

growth rate

Duration:

72 h

Dose descriptor:

EC10

Effect conc.:

> 100 mg/L

Nominal / measured:

nominal

Conc. based on:

test mat.

Basis for effect:

growth rate

Duration:

72 h

Dose descriptor:

EC20

Effect conc.:

> 100 mg/L

Nominal / measured:

nominal

Conc. based on:

test mat.

Basis for effect:

growth rate

Duration:

72 h

Dose descriptor:

EC50

Effect conc.:

> 100 mg/L

Nominal / measured:

nominal

Conc. based on:

test mat.

Basis for effect:

growth rate

Duration:

72 h

Dose descriptor:

LOEC

Effect conc.:

> 100 mg/L

Nominal / measured:

nominal

Conc. based on:

test mat.

Basis for effect:

yield

Duration:

72 h

Dose descriptor:

NOEC

Effect conc.:

>= 100 mg/L

Nominal / measured:

nominal

Conc. based on:

test mat.

Basis for effect:

yield

Duration:

72 h

Dose descriptor:

EC10

Effect conc.:

> 100 mg/L

Nominal / measured:

nominal

Conc. based on:

test mat.

Basis for effect:

yield

Duration:

72 h

Dose descriptor:

EC20

Effect conc.:

> 100 mg/L

Nominal / measured:

nominal

Conc. based on:

test mat.

Basis for effect:

yield

Duration:

72 h

Dose descriptor:

EC50

Effect conc.:

> 100 mg/L

Nominal / measured:

nominal

Conc. based on:

test mat.

Basis for effect:

yield

Details on results:

Control cultures showed a cell density increase of 251.1-fold within 72-hours, a coefficient of variation of sectional (daily) growth rates of 9.1%, and a coefficient of variation of average growth between control replicates of 2.0%, thus all validity criteria were met.

At the start of the test 107% of the nominal test concentrations were found (average of all test concentration). After 72-hours, 108% of the nominal value was determined (average of all test concentrations). During the test the algae were exposed to a mean of 107% of nominal. Results therefore refer to nominal values, since the concentrations of the test item were within ± 20% of the nominal concentrations during the test.

The 72-hour EyC50 and ErC50 values were calculated to be >100 mg/L (pH adjusted). The 72-hour NOEyC and NOErC were determined to be ≥100 mg/L (pH adjusted), the 72-hour LOEyC and LOErC were > 100 mg/L. The microscopic examination of the shape of the algal cells after 72 hours of test duration did not show any difference between the algae that had been growing up to a nominal test concentration of 100 mg test item/L and the algal cells in the control. Thus, the shape of the algal cells was not obviously affected up to this test concentration, the highest concentration tested.

Results with reference substance (positive control):

The most recent test with Pseudokirchneriella subcapitata performed with the reference substance (potassium dichromate) resulted in 72-hour EyC50, ErC50, and EbC50 values of 0.402, 0.878 and 0.449 mg/L, respectively. The 72-hour NOEC values for yield, growth rate and biomass were all 0.2 mg/L. The 72-hour LOEC values for yield, growth rate and biomass were all 0.63 mg/L. The results were consistent with the levels proposed by the OECD 201 guideline and thus demonstrate that the algae and test conditions were satisfactory.

Reported statistics and error estimates:

Based on the calculated cell densities, the 72-hour ErC50 and EyC50, the corresponding EC20 and EC10 values and where possible their 95%-confidence limits were calculated by a three parametric normal concentration distribution function (CDF). For 72-hour LOEC and NOEC determination, the calculated growth rates and yields at each test concentration were tested for significant differences compared to the control values by Dunnett’s t-test. The software used to perform the statistical analysis was ToxRat Professional (Version 3.3.0).

Influence of Glypure™ 99 on the Growth of Pseudokirchneriella subcapitata

Parameter	Yield [mg test item/L]	Growth rate [mg test item/L]
72-hour EC50	>100	>100
95% conf. interval	n.d.	n.d.
72-hour EC20	>100	>100
95% conf. interval	n.d.	n.d.
72-hour EC10	>100	>100
95% conf. interval	n.d.	n.d.
72-hour NOEC	≥100	≥100
72-hour LOEC	>100	>100

n.d.: not determinable

Values refer to nominal test concentrations

 

 

Summary of Analytical Results

Sample description [mg test item/L]	% of nominal^1	RSD [%]	n
Control	n.a.	n.a.	4
2.6	117	1	4
6.4	103	2	4
16	110	1	4
40	103	0	4
100	103	1	4

¹mean value of all measured samples per treatment group

RSD: relative standard deviation per treatment group

n: number of analysed samples

n.a.: not applicable

 

 

Temperature in the Test Media during the Test Period

Water temperature [°C]
0 h	24 h	48 h	72 h
21.8	21.7	21.6	21.7

 

 

pH-Values in the Test Media at the Start and End of the Test

Nominal concentration [mg test item/L]	pH value
	0 hours	72 hours
Control	8.0	9.0
2.6	8.0	8.9
6.4	8.0	9.1
16	8.0	9.1
40	8.1	9.0
100	8.2	9.0

 

Validity criteria fulfilled:

yes

Conclusions:

The 72-hour EyC50 and ErC50 values were calculated to be >100 mg/L (pH adjusted). The 72-hour NOEyC and NOErC were determined to be ≥100 mg/L (pH adjusted), the 72-hour LOEyC and LOErC were > 100 mg/L (pH adjusted). Results were based on nominal values since the concentrations of the test item were within ± 20% of the nominal concentrations during the test.

Executive summary:

The toxicity of Glypure™ 99 to the freshwater algae, Pseudokirchneriella subcapitata was assessed according to OECD Guideline 201.

 

Exponentially growing cultures of the algal were exposed to test concentrations of 100, 40, 16, 6.4 and 2.6 mg/L and a control. Given the acidic properties of the test item, the pH of the highest test concentration was adjusted from 4.0 to 8.2 using 1 M NaOH. The inhibition of growth in relation to control cultures was determined over a test period of 72-hours. Defined volumes of the algal suspensions were sampled after 24, 48 and 72-hours for determination of cell densities by spectrophotometric measurement.

 

At the start of the test 107% of the nominal test concentrations were found (average of all test concentration). After 72-hours, 108% of the nominal value was determined (average of all test concentrations). During the test the algae were exposed to a mean of 107% of nominal. Results therefore refer to nominal values, since the concentrations of the test item were within ± 20% of the nominal concentrations during the test.

 

Control cultures showed a cell density increase of 251.1-fold within 72-hours, a coefficient of variation of sectional (daily) growth rates of 9.1%, and a coefficient of variation of average growth between control replicates of 2.0%, thus all validity criteria were met.

 

The 72-hour EyC50 and ErC50 values were calculated to be >100 mg/L (pH adjusted). The 72-hour NOEyC and NOErC were determined to be ≥100 mg/L (pH adjusted), the 72-hour LOEyC and LOErC were > 100 mg/L (pH adjusted).

Reference 3

Endpoint:

toxicity to aquatic algae and cyanobacteria

Type of information:

experimental study

Adequacy of study:

key study

Study period:

15 March to 28 May, 2021

Reliability:

1 (reliable without restriction)

Rationale for reliability incl. deficiencies:

guideline study

Qualifier:

according to guideline

Guideline:

OECD Guideline 201 (Freshwater Alga and Cyanobacteria, Growth Inhibition Test)

Version / remarks:

adopted March 23, 2006, corrected July 28, 2011

Deviations:

yes

Remarks:

In contrast to the recommendations of the Guidelines for testing algae the test was started with a higher nominal algal cell density of 15.000 cells per mL test medium in order to improve the accuracy of cell density measurements after 24 h

GLP compliance:

yes (incl. QA statement)

Analytical monitoring:

yes

Details on sampling:

Duplicate samples from the freshly prepared test media (without algae) of all test concentrations and from the control were taken at the start of the test. To determine test item stability under the test conditions and the maintenance of test item concentrations during the test period, duplicate samples from the test media of all test concentrations and the control (containing algae) were taken after 24- and 48-hours and at the end of the test (after the 72-hour test period) by pouring together the contents of the test beakers of each treatment. All samples remained undiluted until sample preparation. All samples were stored in a freezer (≤ - 20°C), protected from light until analysis was performed. Afterwards, the samples were again stored deep frozen.

Vehicle:

no

Details on test solutions:

The test medium of the nominal 100 mg/L test concentration was prepared by dissolving 82.2 mg test item into 822 mL test water by intense stirring for 20 minutes at room temperature. The pH of the highest test concentration was adjusted from 7.0 to 7.4 using 1 M NaOH. Adequate volumes of this test medium were diluted with test water to prepare the test media of the other desired test concentrations. The test media were prepared just before introduction of the algae (test initiation).

Test organisms (species):

Anabaena flos-aquae

Details on test organisms:

Species: Anabaena flos-aquae, UTEX 1444
Origin: The algae were supplied by "The University of Texas at Austin, UTEX Culture Collection of Algae", 205 W. 24thSt., BIO 218, Austin, TX 78712, USA.
Breeding Conditions: The algae were cultivated in the testing laboratory under standardised conditions according to the test guidelines.

Test type:

static

Water media type:

freshwater

Limit test:

no

Total exposure duration:

72 h

Remarks on exposure duration:

No further remarks

Post exposure observation period:

Not applicable

Hardness:

3.0 mmol/L (= 300 mg/L) as CaCO3

Test temperature:

22.1 to 22.6 °C

pH:

Control at test start: 7.5; Control at test end: 8.9;
Test item treatments at test start: 7.4 to 7.5; Test item treatments at test end: 8.8 to 8.9

Dissolved oxygen:

Not applicable

Salinity:

Not applicable

Conductivity:

Not reported

Nominal and measured concentrations:

Nominal: 0 (control), 100, 40, 16, 6.4 and 2.6 mg/L (pH adjusted);
Geometric mean measured: 0 (control), 95.3, 39.2, 16.5, 6.36 and 2.28 mg/L (pH adjusted)

Details on test conditions:

Tests were conducted in 50 mL Erlenmeyer flasks containing nominal 50 mL of test medium covered with sterile caps. All glassware was autoclaved prior to test start. Reconstituted water (20X AAP medium) was prepared with deionised water in which analytical grade salts were added at the appropriate nominal concentrations.

Pre-experiments were performed to determine a suitable concentration range and to establish suitable methods to prepare the test solutions.

In contrast to the recommendations of the Guidelines for testing algae, the definitive test was started with a higher nominal algal cell density of 15000 cells per mL test medium in order to improve the accuracy of cell density measurements after 24 hours and to fulfil the validity criteria. These cells were taken from an exponentially growing pre-culture, which was set up 3 days prior to the test start under the same conditions as in the test. The test was performed with three replicates per test concentration and six replicates in the control. The flasks were incubated in an incubation chamber and were placed in a random order and were repositioned each day to minimize differences in test conditions. Two additional replicates of each test item concentration and the control were set up for analytical dose verification after 24- and 48-hours. These additional replicates were incubated under the same conditions as described above. Additionally, one replicate of each test concentration and of the control was prepared without algae to provide a “blank” for the spectrophotometric measurements. The additional replicates were incubated under the same conditions as described above. The blank values were subtracted from the absorption measured in the samples containing algae to eliminate absorption caused by the test item.

Temperature was measured daily in an Erlenmeyer flask filled with water and incubated under the same conditions as the test flasks. The pH was measured in all test item concentrations and the control at the start and the end of the test. Test units were provided with continuous illumination, with light intensity measured once during the test at 6 positions distributed over the experimental area at the surface of the test media (mean light intensity was 4142 lux and ranged from 3900 to 4420 lux).

The cell density on each observation time was determined by spectrophotometric measurement. Defined volumes of the algal suspensions from all replicates and from the blanks were sampled after 24-, 48- and 72-hours of exposure, and were not replaced. The algal cell densities were calculated by subtracting the absorption of the blanks, from each of the measured absorption of the test media (with algae). Based on the counted cell densities and the absorption from an algal suspension and its dilutions, a linear regression was performed for the calculation of the cell densities of the replicates during the test. To check for any effect of the test item on the morphology of the algal cells, at least one sample from all test concentrations and the control was taken after the 72-hour test period, and the shape of the treated algal cells compared to the control was examined microscopically.

Reference substance (positive control):

yes

Remarks:

3,5-dichlorophenol (tested at least twice a year)

Key result

Duration:

72 h

Dose descriptor:

EC50

Effect conc.:

> 100 mg/L

Nominal / measured:

nominal

Conc. based on:

test mat.

Basis for effect:

growth rate

Remarks on result:

other: Re-assessed by the study assessor: As the test item concentrations were maintained within 80-120% of nominal concentrations during the test, the result should be based on nominal values.

Key result

Duration:

72 h

Dose descriptor:

NOEC

Effect conc.:

>= 100 mg/L

Nominal / measured:

nominal

Conc. based on:

test mat.

Basis for effect:

growth rate

Remarks on result:

other: Re-assessed by the study assessor: As the test item concentrations were maintained within 80-120% of nominal concentrations during the test, the result should be based on nominal values.

Key result

Duration:

72 h

Dose descriptor:

EC10

Effect conc.:

> 100 mg/L

Nominal / measured:

nominal

Conc. based on:

test mat.

Basis for effect:

growth rate

Remarks on result:

other: Re-assessed by the study assessor: As the test item concentrations were maintained within 80-120% of nominal concentrations during the test, the result should be based on nominal values.

Duration:

72 h

Dose descriptor:

LOEC

Effect conc.:

> 95.3 mg/L

Nominal / measured:

meas. (geom. mean)

Conc. based on:

test mat.

Basis for effect:

growth rate

Duration:

72 h

Dose descriptor:

NOEC

Effect conc.:

>= 95.3 mg/L

Nominal / measured:

meas. (geom. mean)

Conc. based on:

test mat.

Basis for effect:

growth rate

Duration:

72 h

Dose descriptor:

EC10

Effect conc.:

> 95.3 mg/L

Nominal / measured:

meas. (geom. mean)

Conc. based on:

test mat.

Basis for effect:

growth rate

Duration:

72 h

Dose descriptor:

EC20

Effect conc.:

> 95.3 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.:

> 95.3 mg/L

Nominal / measured:

meas. (geom. mean)

Conc. based on:

test mat.

Basis for effect:

growth rate

Duration:

72 h

Dose descriptor:

LOEC

Effect conc.:

> 95.3 mg/L

Nominal / measured:

meas. (geom. mean)

Conc. based on:

test mat.

Basis for effect:

yield

Duration:

72 h

Dose descriptor:

NOEC

Effect conc.:

>= 95.3 mg/L

Nominal / measured:

meas. (geom. mean)

Conc. based on:

test mat.

Basis for effect:

yield

Duration:

72 h

Dose descriptor:

EC10

Effect conc.:

> 95.3 mg/L

Nominal / measured:

meas. (geom. mean)

Conc. based on:

test mat.

Basis for effect:

yield

Duration:

72 h

Dose descriptor:

EC20

Effect conc.:

> 95.3 mg/L

Nominal / measured:

meas. (geom. mean)

Conc. based on:

test mat.

Basis for effect:

yield

Duration:

72 h

Dose descriptor:

EC50

Effect conc.:

> 95.3 mg/L

Nominal / measured:

meas. (geom. mean)

Conc. based on:

test mat.

Basis for effect:

yield

Details on results:

Control cultures showed a cell density increase of 21.0-fold within 72-hours, a coefficient of variation of sectional (daily) growth rates of 27.9%, and a coefficient of variation of average growth between control replicates of 3.2%, thus all validity criteria were met.

At the start of the test 105% of the nominal test concentrations were found (average of all test concentration). After 72-hours, 89% of the nominal value was determined (average of all test concentrations). Overall, the algae were exposed to a mean of 97 % of the nominal concentrations during the test. However, according to the study report, all reported biological effect endpoints are related to geometric mean measured concentrations.

The 72-hour EyC50, ErC10, ErC50 and ErC10 values were all calculated to be >95.3 mg/L (pH adjusted). The 72-hour NOEyC and NOErC were determined to be ≥95.3 mg/L (pH adjusted), the 72-hour LOEyC and LOErC were >95.3 mg/L. The microscopic examination of the shape of the algal cells after 72 hours of test duration did not show any difference between the algae that had been growing up to a up to a geometric mean measured test concentration of 95.3 mg/L and the algal cells in the control. Thus, the shape of the algal cells was not obviously affected up to this test concentration, the highest concentration tested.

Results with reference substance (positive control):

The most recent test with Anabaena flos-aquae performed with the reference substance (3,5-Dichlorophenol) resulted in 72-hour EyC50, ErC50, and EbC50 values of 1.70, 3.19 and 1.98 mg/L, respectively. The 72-hour NOEC values for yield, growth rate and biomass were 0.32, 1.0 and 0.32 mg/L, respectively. The 72-hour LOEC values for yield, growth rate and biomass were 1.0, 3.2 and 1.0 mg/L, respectively. The results were consistent with the levels proposed by the OECD 201 guideline and thus demonstrate that the algae and test conditions were satisfactory.

Reported statistics and error estimates:

Based on the calculated cell densities, the 72-hour ErC50 and EyC50, the corresponding EC20 and EC10 values and where possible their 95%-confidence limits were calculated by a three parametric normal concentration distribution function (CDF). For 72-hour LOEC and NOEC determination, the calculated growth rates and yields at each test concentration were tested for significant differences compared to the control values by Dunnett’s t-test. The software used to perform the statistical analysis was ToxRat Professional (Version 3.3.0).

Influence of Glypure™ 99 on the Growth of Anabaena flos-aquae

Parameter	Yield [mg test item/L]	Growth rate [mg test item/L]
72-hour EC50	>95.3	>95.3
95% conf. interval	n.d.	n.d.
72-hour EC20	>95.3	>95.3
95% conf. interval	n.d.	n.d.
72-hour EC10	>95.3	>95.3
95% conf. interval	n.d.	n.d.
72-hour NOEC	≥95.3	≥95.3
72-hour LOEC	>95.3	>95.3

n.d.: not determinable

Values refer to geometric mean measured test concentrations

 

 

Summary of Analytical Results

Sample description [mg test item/L]	% of nominal^1	RSD [%]	n	Geometric mean concentration^2 [mg test item/L]
Control	n.a.	n.a.	4	n.a.
2.6	91	29	4	2.28
6.4	100	8	4	6.36
16	103	2	4	16.5
40	98	5	4	39.2
100	95	8	4	95.3

¹mean value of all measured samples per treatment group

²Calculated according to OECD 23 (2000), Annex II as referenced by OECD 201 and values were rounded to three significant digits

RSD: relative standard deviation per treatment group

n: number of analysed samples

n.a.: not applicable

 

 

Temperature in the Test Media during the Test Period

Water temperature [°C]
0 h	24 h	48 h	72 h
22.3	22.4	22.6	22.1

 

 

pH-Values in the Test Media at the Start and End of the Test

Geometric mean measured concentration [mg test item/L]	pH value
	0 hours	72 hours
Control	7.5	8.9
2.28	7.5	8.8
6.36	7.5	8.9
16.5	7.5	8.9
39.2	7.5	8.8
95.3	7.4	8.8

 

Validity criteria fulfilled:

yes

Conclusions:

According to the study report: The 72-hour EyC50 and ErC50 values were calculated to be >95.3 mg/L (pH adjusted). In the absence of any statistically significant differences from the untreated control, the 72-hour NOEyC and NOErC were determined to be ≥95.3 mg/L (pH adjusted), the 72-hour LOEyC and LOErC were > 95.3 mg/L (pH adjusted). All reported biological effect endpoints are related to geometric mean measured concentrations.

Re-assessed by the study assessor: As the test item concentrations were maintained within 80-120% of nominal concentrations during the test, the results should be based on nominal values. Therefore, the 72h-EC50 and EC10 values are >100 mg/L and the 72h-NOEC value is ≥100 mg/L.

Executive summary:

The toxicity of Glypure™ 99 to the freshwater algae, Anabaena flos-aquae, was assessed according to OECD Guideline 201.

 

Exponentially growing cultures of the algal were exposed to nominal test concentrations of 100, 40, 16, 6.4 and 2.6 mg/L (corresponding to geometric mean measured concentrations of 95.3, 39.2, 16.5, 6.36 and 2.28 mg/L) and a control. Given the acidic properties of the test item, the pH of the highest test concentration was adjusted from 7.0 to 7.4 using 1 M NaOH. The inhibition of growth in relation to control cultures was determined over a test period of 72-hours. Defined volumes of the algal suspensions were sampled after 24, 48 and 72-hours for determination of cell densities by spectrophotometric measurement.

 

Control cultures showed a cell density increase of 21.0-fold within 72-hours, a coefficient of variation of sectional (daily) growth rates of 27.9%, and a coefficient of variation of average growth between control replicates of 3.2%, thus all validity criteria were met.

 

At the start of the test 105% of the nominal test concentrations were found (average of all test concentration). After 72-hours, 89% of the nominal value was determined (average of all test concentrations). Overall, the algae were exposed to a mean of 97 % of the nominal concentrations during the test.

 

According to the study report, all reported biological effect endpoints are related to geometric mean measured concentrations. The 72-hour EyC50 and ErC50 values were calculated to be >95.3 mg/L (pH adjusted). In the absence of any statistically significant differences from the untreated control, the 72-hour NOEyC and NOErC were determined to be ≥95.3 mg/L (pH adjusted), the 72-hour LOEyC and LOErC were > 95.3 mg/L (pH adjusted).

 

However, according to the study assessor, the conclusion were re-assessed. As the test item concentrations were maintained within 80-120% of nominal concentrations during the test, the results should be based on nominal values. The key 72h-EC50 and EC10 values are >100 mg/L and the key 72h-NOEC value is ≥100 mg/L. 

Description of key information

Studies assessing the toxicity of glycolic acid to algae are available. The key 72-hour ErC50 and ErC10 values (due to being the most conservative of the available reliable data) were >100 mg/L (based on analytically confirmed nominal test concentrations), shown in the freshwater algae species, Anabaena flos-aquae (Bebon and Sonntag, 2021b) and Selenastrum capricornutum (Bebon and Sonntag, 2021c). These key studies were GLP compliant and met all validity criteria of the study guideline (OECD guideline 201).

Additional information

It is noted that although the most conservative reported ErC50 value is 31.2 mg a.s./L (Haskell Laboratory for Health and Environmental Sciencies - Sloman, 2002), this study is unreliable. Although results from this study were based on nominal test item concentrations, the chemical analysis demonstrated that concentrations decreased to levels below the limit of quantification (LOQ) at 72 hours, thus actual exposure concentrations are expected to be lower than the nominal test item concentrations. Moreover, reliable test item concentrations cannot be established because chemical analysis of the test solutions was only performed at days 0 and 3 of the test. Furthermore, the composition of the test material was not sufficiently reported (70% glycolic acid is stated), with information included on a few impurities, but not all. When considering OECD guideline 201 (2011) validity criteria were met, except for the mean coefficient of variation for section-by-section specific growth rates (days 0-1, 1-2 and 2-3, for 72-hour tests) in the control cultures, which was >35% (70%). Consequently, this study was considered unreliable.