Sodium ascorbate

Toxicity to aquatic algae and cyanobacteria

Link to relevant study record(s)

Reference

Endpoint:

toxicity to aquatic algae and cyanobacteria

Type of information:

experimental study

Adequacy of study:

key study

Study period:

2011-12-30 to 2012-01-09

Reliability:

1 (reliable without restriction)

Rationale for reliability incl. deficiencies:

guideline study

Qualifier:

according to guideline

Guideline:

OECD Guideline 201 (Alga, Growth Inhibition Test)

Deviations:

no

Qualifier:

according to guideline

Guideline:

EU Method C.3 (Algal Inhibition test)

Deviations:

no

GLP compliance:

yes (incl. QA statement)

Specific details on test material used for the study:

- CAS Number: 134-03-2
- Batch Number.: TL01101901

Analytical monitoring:

yes

Details on sampling:

- Concentrations: 220 mg/L. The samples from the lower test concentrations were not analyzed, since these concentrations were below the NOEC determined in this test.
- Sampling method: For measurement of the actual concentrations of the test item, duplicate samples were taken from the test media of all test concentrations and the control after 0, 3, 9, 24, and 72 hours of test duration. For sampling at 3, 9, and 24 hours, additional flasks containing the test medium with algae were incubated for each treatment under the test conditions. For sampling at the end of the test, the test medium of the treatment replicates was pooled.
- Sample storage conditions before analysis: Immediately after sampling, 4% oxalic acid solution was added to each sample (1 mL oxalic acid per mL sample volume) in order to stabilize the latter during the storage period. Thereafter, all samples were stored deep-frozen (at about -20 °C) under nitrogen gas.

Vehicle:

no

Details on test solutions:

PREPARATION AND APPLICATION OF TEST SOLUTION (especially for difficult test substances)
- Method: The highest nominal concentration of 220 mg/L was prepared by dissolving 220.40 mg of the test item completely in 1000 mL of test water stirring intensely for 10 minutes at room temperature in the dark. The test medium of the highest test concentration was diluted with adequate volumes of test water to prepare the lower concentrated test media. The test media were prepared as far as possible in the dark just before the start of the test.
- Differential loading: No
- Controls: Test water without test item

Test organisms (species):

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

Details on test organisms:

TEST ORGANISM
- Common name: algae
- Strain: Strain No. 61.81 SAG
- Source (laboratory, culture collection): Collection of Algal Cultures (SAG, Institute for Plant Physiology, University of Göttingen, 37073 Göttingen / Germany)
- Age of inoculum (at test initiation): 4 days
- Method of cultivation: The algae were cultivated under the test conditions. The inoculum culture was diluted threefold one day before the start of the test to ensure that the algae were in the exponential growth phase when used to inoculate the test solutions.

ACCLIMATION
- Acclimation period: 4 days
- Culturing media and conditions (same as test or not): same
- Any deformed or abnormal cells observed: no

Test type:

static

Water media type:

freshwater

Limit test:

no

Total exposure duration:

72 h

Hardness:

0.15 mmol/L

Test temperature:

23 °C

pH:

Start of test: 7.8 - 8.0
End of test: 8.2 - 8.5

Nominal and measured concentrations:

Nominal: 4.6, 10, 22, 46, 100 and 220 mg/L
Measured concentrations (nominal 220 mg/L):
0 h: 206 mg/L
3 h: 179 mg/L
9 h: 151 mg/L
24 h: 85.8 mg/L
72 h: 15.5 mg/L

Details on test conditions:

TEST SYSTEM
- Test vessel: 50 mL Erlenmeyer flasks
- Type: Covered with a glass dish
- Aeration: During exposure, the test solutions were continuously stirred by magnetic stirrers.
- Initial cells density: 10000 cells/mL
- No. of vessels per concentration (replicates): 3
- No. of vessels per vehicle control (replicates): 6

GROWTH MEDIUM
- Standard medium used: Yes

TEST MEDIUM / WATER PARAMETERS
- Source/preparation of dilution water: Reconstituted test water (AAP Medium) prepared according to the test guidelines
NaHCO3: 15 mg/L
K2HPO4: 1.044 mg/L
MgSO4 × 7 H2O: 14.6 mg/L
MgCl2 × 6 H2O: 12.16 mg/L
CaCl2 × 2 H2O: 4.41 mg/L
NaNO3: 25.5 mg/L
H3BO3: 186.0 μg/L
MnCl2 × 4 H2O: 415.0 μg/L
ZnCl2: 3.27 μg/L
CoCl2 × 6 H2O: 1.43 μg/L
CuCl2 × 2 H2O: 0.012 μg/L
Na2MoO4 × 2 H2O: 7.26 μg/L
FeCl3 × 6 H2O: 160.0 μg/L
Na2EDTA × 2 H2O: 300.0 μg/L
- Culture medium different from test medium: No


OTHER TEST CONDITIONS
- Sterile test conditions: No
- Adjustment of pH: No
- Photoperiod: Continuous illumination
- Light intensity and quality: Fluorescent tubes, 4900 Lux ± 15%

EFFECT PARAMETERS MEASURED:
- Determination of cell concentrations: Fluorescence measurement

TEST CONCENTRATIONS
- Spacing factor for test concentrations: 2.1
- Range finding study: The selection of the test concentrations was based on the results of a range-finding test (non-GLP). No details on the range finding study are reported.

Reference substance (positive control):

yes

Remarks:

Potassium dichromate is tested as a positive control twice a year. The result of the latest positive control test performed in October 2011 showed that the sensitivity of the test system was within the internal historical range.

Key result

Duration:

72 h

Dose descriptor:

EC50

Effect conc.:

> 74 mg/L

Nominal / measured:

meas. (TWA)

Conc. based on:

test mat.

Basis for effect:

growth rate

Duration:

72 h

Dose descriptor:

EC50

Effect conc.:

> 74 mg/L

Nominal / measured:

meas. (TWA)

Conc. based on:

test mat.

Basis for effect:

biomass

Key result

Duration:

72 h

Dose descriptor:

NOEC

Effect conc.:

>= 74 mg/L

Nominal / measured:

meas. (TWA)

Conc. based on:

test mat.

Basis for effect:

growth rate

Duration:

72 h

Dose descriptor:

NOEC

Effect conc.:

>= 74 mg/L

Nominal / measured:

meas. (TWA)

Conc. based on:

test mat.

Basis for effect:

biomass

Details on results:

- Exponential growth in the control (for algal test): Yes
- Observation of abnormalities (for algal test): No
- Unusual cell shape: No
- Flocculation: No
- Adherence to test vessels: No
- Any observations (e.g. precipitation) that might cause a difference between measured and nominal values: No

Results with reference substance (positive control):

Not specified.

Reported statistics and error estimates:

Not specified.

Table: Results for the growth inhibition test with Pseudokirchneriella subcapitata.

Parameter (0-72 h)	Growth rate	Yield
EC50 (mg/L)	>74	>74
NOEC (mg/L)	≥74	≥74

Validity criteria fulfilled:

yes

Conclusions:

The test item sodium ascorbate had no statistically significant inhibitory effect on the growth of the algae (average growth-rate and yield) at the highest tested concentration of mean measured 74 mg/L.

Executive summary:

The influence of the test item on the growth of the freshwater green algal species Pseudokirchneriella subcapitata was investigated in a 72-hour static test according to OECD TG 201. The nominal concentrations of the test item of 4.6, 10, 22, 46, 100 and 220 mg/L were tested in parallel with a control. The measured concentrations of the test item in the test media of the nominal test concentration of 220 mg/L were 94, 81, 69 and 39% of the nominal value at test start (0 hours), after 3, 9 and 24 hours, respectively. At the end of the test, 7% of the nominal value was found. Thus, during the test period of 72 hours the test item concentration in the test medium decreased. The mean measured test concentration (time-weighted mean) was calculated to be 74 mg/L, corresponding to 33% of the nominal value. The biological results were as follows (on the basis of the mean measured concentration of the test item): 72-hour EC50 > 74 mg/L for the growth rate and yield, respectively; 72-hour NOEC ≥ 74 mg/L both for the growth rate and yield. The test item had no statistically significant inhibitory effect on the growth of the algae (average growth-rate and yield) at the highest tested concentration of mean measured 74 mg/L. All validity criteria of the test guidelines were fulfilled

Description of key information

The test item sodium ascorbate had no statistically significant inhibitory effect on the growth of the algae (average growth-rate and yield) at the highest tested concentration of mean measured 74 mg/L. On the basis of the mean measured concentration of the test item, the 72-hour EC50 was determined to be > 74 mg/L for the growth rate and yield, respectively; the 72-hour NOEC was determined to be ≥ 74 mg/L both for the growth rate and yield.

Key value for chemical safety assessment

EC50 for freshwater algae:

74 mg/L

EC10 or NOEC for freshwater algae:

74 mg/L

Additional information

The influence of the test item on the growth of the freshwater green algal species Pseudokirchneriella subcapitata was investigated in a 72-hour static test according to OECD TG 201. The nominal concentrations of the test item of 4.6, 10, 22, 46, 100 and 220 mg/L were tested in parallel with a control. The measured concentrations of the test item in the test media of the nominal test concentration of 220 mg/L were 94, 81, 69 and 39% of the nominal value at test start (0 hours), after 3, 9 and 24 hours, respectively. At the end of the test, 7% of the nominal value was found. Thus, during the test period of 72 hours the test item concentration in the test medium decreased. The mean measured test concentration (time-weighted mean) was calculated to be 74 mg/L, corresponding to 33% of the nominal value. The biological results were as follows (on the basis of the mean measured concentration of the test item): 72-hour EC50 > 74 mg/L for the growth rate and yield, respectively; 72-hour NOEC ≥ 74 mg/L both for the growth rate and yield. The test item had no statistically significant inhibitory effect on the growth of the algae (average growth-rate and yield) at the highest tested concentration of mean measured 74 mg/L. All validity criteria of the test guidelines were fulfilled