Disodium 2-oxoglutarate

Toxicity to aquatic plants other than algae

Administrative data

Endpoint:

toxicity to aquatic plants other than algae

Type of information:

experimental study

Adequacy of study:

key study

Study period:

10 May - 31 August 2017

Reliability:

1 (reliable without restriction)

Rationale for reliability incl. deficiencies:

guideline study

Data source

Reference

Reference Type:

study report

Title:

Unnamed

Year:

2017

Report date:

2017

Materials and methods

Test guideline

Qualifier:

according to guideline

Guideline:

OECD Guideline 221 (Lemna sp. Growth Inhibition Test)

Version / remarks:

23 March 2006

Deviations:

no

GLP compliance:

yes (incl. QA statement)

Test material

Test material information

Constituent 1

Reference substance name:

Disodium 2-oxoglutarate

EC Number:

206-167-8

EC Name:

Disodium 2-oxoglutarate

Cas Number:

305-72-6

Molecular formula:

C5H6O5.2Na

IUPAC Name:

disodium 2-oxopentanedioate

impurity 1

Reference substance name:

Water

EC Number:

231-791-2

EC Name:

Water

Cas Number:

7732-18-5

Molecular formula:

H2O

IUPAC Name:

Oxidane

Test material form:

solid: crystalline

Remarks:

Crystalline powder

Details on test material:

Batch No: 14152300
Storage: 15 – 25 °C

Sampling and analysis

Analytical monitoring:

yes

Test solutions

Vehicle:

no

Details on test solutions:

PREPARATION AND APPLICATION OF TEST SOLUTION
The following test vessels were set up:
- Test solution (Tn); containing test medium, test item and duckweed (six replicates)
- Blank control (Bn); containing test medium and duckweed (six replicates)
- Since the test item is soluble, the test solutions were prepared by directly dissolving the test item in Swedish Standard (SIS) medium. The resulting solution was used as the test solution in the toxicity test. Pure SIS medium served as blank controls. The pH of the SIS medium was adjusted before the test to pH 6.5±0.2.
- Young and rapidly growing plants without visible lesions or discoloration (chlorosis) from an exponentially growing monoculture of Lemna minor were distributed into the test vessels (9-12 fronds per replicate, 2-4 fronds per plant). The vessels were randomly placed in the incubation chamber at the start of the test and randomly repositioned after the observations made at days 3 and 5.
- Semi static exposure conditions: New test solutions were prepared similarly at days 3 and 5.
- Evidence of undissolved material: Not relevant. Since the test item is soluble, the test solutions were prepared by directly dissolving the test item in SIS medium. The resulting solution was used as the test solution in the toxicity test.

Test organisms

Test organisms (species):

Lemna minor

Details on test organisms:

TEST ORGANISMS
- Test organism: Exponential growing plant monoculture of Lemna minor (Umweltbundesamt, FGIII 2.5, Überwachungsverfahren Abwasserentsorgung, Schichauweg 58, D-12307 Berlin)
- Culture: All-glass vessel containing sterile Swedish Standard (SIS) medium and the exponentially growing plant monoculture.
- Illumination: Continuous (6500–10000 lux) from Osram Fluora L18W77 (Osram AG, Winterthur, Switzerland) and CH Lighting F18T8/6500K EUP (CH Lighting CO., Ltd., China)
- Temperature: 24±2°C
- Control of sensitivity: Yearly, with 3,5-dichlorophenol

Study design

Test type:

semi-static

Water media type:

freshwater

Limit test:

yes

Total exposure duration:

7 d

Test conditions

Hardness:

not reported

Test temperature:

Min.: 22.2 ; Max.: 23.2 ; Mean: 22.7

pH:

6.3 - 7.1

Dissolved oxygen:

not reported

Salinity:

not reported

Conductivity:

not reported

Nominal and measured concentrations:

The test concentrations of a-Ketoglutarate disodium salt during the 7-day test period were determined by HPLC at the beginning of the test, as well as after 3 (old and fresh solutions), 5 (old and fresh solutions) and 7 days of exposure. These analyses confirmed the right dosage of the test item and showed that the test concentrations decreased very rapidly. Indeed, while in all fresh solutions (day 0, 3 and 5) the concentration was within the expected range (97.2, 95.8 and 95.4 mg/l, respectively), the concentration was below the LOQ of 1.05 mg/l in all old solutions (day 3, 5 and 7). Therefore, the toxicity was assessed based on the measured concentration, calculated as the arithmetic mean (AM) of the three geometric means (GMs) of each respective test medium renewal period; this arithmetic mean was 7.0 mg/l.

Details on test conditions:

TEST SYSTEM
- Test vessel: 400 ml beakers, all-glass, with 200 ml of test medium. The beakers were covered with black paper up until the 200 ml mark to ensure that illumination comes only from above and not from the sides.
- Test medium: Recommended Swedish Standard (SIS) medium, prepared with ultra-pure water
- Preculture: Exponentially growing plant monoculture of Lemna minor
- Illumination: Source: continuous (6500–10000 lux) from Osram Fluora L18W77 (Osram AG, Winterthur, Switzerland) and CH Lighting F18T8/6500K EUP (CH Lighting CO., Ltd., China)
- Homogeneity: the light intensity is maintained within ±15% from the average light intensity over the incubation area.
- Incubation: Beakers were incubated on a black non-reflecting surface
- pH: 6.5 at the beginning of the test; the pH value in the blank control should not increase by more than 1.5 units during the test
- Temperature: 24±2°C
- Test type: Semi-static exposure conditions over a period of 7 days
- Test duration: 7 d

RANGE-FINDING STUDY
Prior to the definitive test a non-GLP range finding test with the nominal concentrations of 10, 100 and 500 mg/l of a-Ketoglutarate disodium salt was performed under static conditions.

EFFECT PARAMETERS MEASURED
Frond Number, Growth Rate
Frond Number, Yield
Dry Weight, Growth Rate
Dry Weight, Yield

Reference substance (positive control):

not required

Results and discussion

Effect concentrations

Effect concentrations 1

Key result

Duration:

7 d

Dose descriptor:

EC50

Effect conc.:

> 7 mg/L

Nominal / measured:

meas. (geom. mean)

Conc. based on:

test mat.

Basis for effect:

frond number

Effect concentrations 2

Key result

Duration:

7 d

Dose descriptor:

NOEC

Effect conc.:

7 mg/L

Nominal / measured:

meas. (geom. mean)

Conc. based on:

test mat.

Basis for effect:

frond number

Effect concentrations 3

Duration:

7 d

Dose descriptor:

EC50

Effect conc.:

> 7 mg/L

Nominal / measured:

meas. (geom. mean)

Conc. based on:

test mat.

Basis for effect:

frond number

Effect concentrations 4

Duration:

7 d

Dose descriptor:

NOEC

Effect conc.:

7 mg/L

Nominal / measured:

meas. (geom. mean)

Conc. based on:

test mat.

Basis for effect:

frond number

Effect concentrations 5

Duration:

7 d

Dose descriptor:

EC50

Effect conc.:

> 7 mg/L

Nominal / measured:

meas. (geom. mean)

Conc. based on:

test mat.

Basis for effect:

other: Dry Weight

Effect concentrations 6

Duration:

7 d

Dose descriptor:

NOEC

Effect conc.:

7 mg/L

Nominal / measured:

meas. (geom. mean)

Conc. based on:

test mat.

Basis for effect:

other: Dry Weight

Effect concentrations 7

Duration:

7 d

Dose descriptor:

EC50

Effect conc.:

> 7 mg/L

Nominal / measured:

meas. (geom. mean)

Conc. based on:

test mat.

Basis for effect:

other: Dry Weight

Effect concentrations 8

Duration:

7 d

Dose descriptor:

NOEC

Effect conc.:

7 mg/L

Nominal / measured:

meas. (geom. mean)

Conc. based on:

test mat.

Basis for effect:

other: Dry Weight

Details on results:

Effects on frond number
With respect to both endpoints growth rate and yield inhibition, no significant effects were observed compared to the untreated controls. Based on these observations and the measured concentrations of the test item, the median effect concentration with respect to the frond number’s growth rate (frond number ErC50) and yield (frond number EyC50) of a-Ketoglutarate disodium salt to Lemna minor was estimated to be >7.0 mg/l and the NOErC was determined to be 7.0 mg/l.

Effects on dry weight
With respect to both endpoints growth rate and yield inhibition, no significant effects were observed compared to the untreated controls. Based on these observations and the measured concentrations of the test item, the median effect concentration with respect to the dry weight’s growth rate (dry weight ErC50) and yield (dry weight EyC50) of a-Ketoglutarate disodium salt to Lemna minor was estimated to be >7.0 mg/l and the NOErC was determined to be 7.0 mg/l.

Reported statistics and error estimates:

The statistical significance of differences between effects observed for control and test item treatments were determined by Student’s t-test using the software ToxRat® Standard Version 3.2.1

Applicant's summary and conclusion

Validity criteria fulfilled:

yes

Conclusions:

The acute toxicity of a-Ketoglutarate disodium salt (CAS no. 305-72-6) to duckweed Lemna minor was determined in a 7 day semi-static test according to OECD guideline 221. The median effect concentrations (EC50) of a-Ketoglutarate disodium salt on duckweed
Lemna minor were estimated to be > 7 mg/l measured concentration after 7 d of exposure. The NOEC values were determined to be 7 mg/l measured concentration after 7 d of exposure. The results of the test can be considered reliable without restriction.

Executive summary:

The growth inhibitory effects of a-Ketoglutarate disodium salt (CAS no. 305-72-6) to the duckweed Lemna minor were investigated according to test guideline OECD 221, under semi-static conditions over a period of 7 days.

The test item a-Ketoglutarate disodium salt is solid, has a high solubility (>=511 g/l at pH 7.5 and at 23 °C) and is 99.3% pure. The test media were prepared by directly dissolving the test item in Swedish Standard medium.

The single investigated nominal concentration was 100 mg/l.

Six parallel test vessels were used for each test concentration of the test item and six vessels for the blank controls. Since the test item is unstable, fresh test solutions were prepared after 3 and 5 days.

The test concentrations of a-Ketoglutarate disodium salt during the 7-day test period were determined by HPLC at the beginning of the test, as well as after 3 (old and fresh solutions), 5 (old and fresh solutions) and 7 days of exposure. These analyses confirmed the right dosage of the test item and showed that the test concentrations decreased very rapidly indicating the test item being unstable under the conditions of the presented study (high light intensity and an aquatic environment). Indeed, while in all fresh solutions the concentration was within the expected value, the concentration was below the LOQ of 1.05 mg/l in all old solutions (after 2-3 days). Therefore, the toxicity was assessed based on measured concentrations, calculated as the arithmetic mean (AM) of the three geometric means (GMs) of each respective test medium renewal period; this arithmetic mean was 7.0 mg/l.

The two endpoints frond number and biomass (dry weight) were investigated at days 3, 5 and 7 and each of them were assessed as growth rate and yield. No significant toxic effects were observed for any of the endpoints considered.

The results of the toxicity assessment of a-Ketoglutarate disodium salt to the duckweed Lemna minor are summarized in the following table showing ECx values based on measured concentrations:

Parameter, expressed as Measured concentration (mg/l)	EC50	NOEC
Frond Number, Growth Rate	>7.0	7.0
Frond Number, Growth Rate	>7.0	7.0
Frond Number, Yield	>7.0	7.0
Dry Weight, Growth Rate	>7.0	7.0
Dry Weight, Growth Rate	>7.0	7.0
Dry Weight, Yield	>7.0	7.0

The 7-day ErC50 (based on frond number) of a-Ketoglutarate disodium salt (CAS no. 305-72-6) on the duckweed Lemna minor is >7.0 mg/l. This value is based on the measured concentrations.

The validity criterion was fulfilled.