Dipotassium oxalate

Toxicity to microorganisms

Link to relevant study record(s)

Reference

Reference 1

Endpoint:

activated sludge respiration inhibition testing

Type of information:

read-across from supporting substance (structural analogue or surrogate)

Adequacy of study:

key study

Reliability:

2 (reliable with restrictions)

Rationale for reliability incl. deficiencies:

study well documented, meets generally accepted scientific principles, acceptable for assessment

Justification for type of information:

REPORTING FORMAT FOR THE ANALOGUE APPROACH

1. HYPOTHESIS FOR THE ANALOGUE APPROACH
This read-across hypothesis is based on transformation of the target and source substances to common compounds (scenario1 of the RAAF). The target substance Dipotassium oxalate is a salt of the free acid Oxalic acid. The source substance Dipotassium oxalate dissociates in aqueous solutions thereby forming Oxalic acid and potassium ions. Both constituents are ubiquitously occurring and also an essential part of the intermediary metabolism. Based on the available data especially repeated dose toxicity or genetic toxicity, reproductive toxicity for Oxalic acid and potassium ions it can be deduced that Oxalic acid is the toxicity determining constituent. Potassium ions are considered to be less toxic due to their role in cell metabolism and the well established mechanisms to maintain the cellular ion equilibrium.

2. SOURCE AND TARGET CHEMICAL(S) (INCLUDING INFORMATION ON PURITY AND IMPURITIES)
Source: Oxalic acid, CAS 144-62-7
Target: Dipotassium oxalate, CAS 583-52-8

3. ANALOGUE APPROACH JUSTIFICATION
Please refer to the justification for read-across attached to Chapter 13

4. DATA MATRIX
Please refer to the justification for read-across attached to Chapter 13

Reason / purpose for cross-reference:

read-across source

Key result

Duration:

16 h

Dose descriptor:

other: Toxicity threshold

Effect conc.:

1 550 mg/L

Nominal / measured:

nominal

Conc. based on:

test mat.

Basis for effect:

growth inhibition

Details on results:

- Any observations (e.g. precipitation) that might cause a difference between measured and nominal values: not reported
- Effect concentrations exceeding solubility of substance in test medium:
- Adsorption (e.g. of test material to the walls of the test container): not reported

Validity criteria fulfilled:

not applicable

Conclusions:

There are no data available for the target substance dipotassium oxalate, however, there are some information about the source substance oxalic acid. A justification for the use of the read-across approach is attached to Chapter 13. In the present test Pseudomonar putida were incubated with serial dilutions of Oxalic acid for 16h under static conditions. The toxicity threshold was determined to be 1550 mg/L for Pseudomonas putida. Based on this result the EC50 is above the limit concentrations of 100 mg/L. Oxalic acid is not considered to mediate any inhibitory effect on bacterial growth.

Executive summary:

No data are available for the target substance dipotassium oxalate, however, data are available for the structurally similar substance oxalic acid. A justification for read-across is attached to Chapter 13 of this IUCLID.

 

Reference 2

Endpoint:

activated sludge respiration inhibition testing

Type of information:

experimental study

Adequacy of study:

key study

Reliability:

2 (reliable with restrictions)

Rationale for reliability incl. deficiencies:

study well documented, meets generally accepted scientific principles, acceptable for assessment

Principles of method if other than guideline:

- Principle of test: The multiplication of bacterial cells of the genus Pseudomonas is inhibited by dissolved toxic water ingredients. After a certain period the increase in the number of cells in a test culture free from toxic influence and with a defined standardized offer of nutrients will exceed that observed in a teat culture containing dissolved toxic substances and kept under identical conditions. The concentration of the bacterial suspension is measured turbidimetrically (while diffused light is screened off); it is expressed by the extinction of the primary light of the monochromatic radiation at 436 nm for a layer of 10 mm thickness. The concentration at which the inhibitory action of a pollutant starts will be present in that step of a dilution series of the pollutant having an extinction value at the end of the test period that is ≥ 3 % below the mean value of extinction for non-toxic dilutions of the test cultures.
- Short description of test conditions: Determination of the extinction of the monochromatic radiation at 436nm for a 10 mm layer of the bacterial suspension by photoelectric measurement. On the basis of the values thus measured, the final turbidity value of the bacterial suspension are adjusted by means of sterile saline in such a way that the extinction value for a measuring sample that has been subject to onward dilution 1 + 9 with saline will correspond to the extinction value of a Formazin standard suspension TE/F/436 nm = 10.
Prior to the preparation of test cultures the pollutant solution of a known content is neutralized in sterile double-distilled water to be tested, the concentration of the acid or alkaline solution to be added is selected in such a way that the volume added is kept as small as possible. pH is not adjusted, the effect of the pH of the pollutant solution to be studied is to be included in the test. From the pollutant solution prepared as described or the sewage and sterile double-distilled water are prepared, four parallel dilution series in 300-mL Erlenmeyer flasks, stoppered with cotton-lined plastic caps. Each of the dilutions contains 1 part v/v of the pollutant solution in 20 to 214 parts v/v of mixture. The dilution series are prepared as follows: the first flask of each series contains 160 mL of pollutant solution at the start. Starting from this flask prepare the subsequent dilution stein at a constant dilution ratio by consistently mixing 80 mL of preliminary pollutant dilution and 80 mL double distilled water. Consequently, each flask contains 80 mL of culture liquid at the start. Make up each flask of the three dilution series to be inoculated to 100 mL by adding 5 mL each of stock solution 1.5 mL each of stock solution II and 10 mL each of the prepared bacterial suspension from the preliminary culture having a known adjusted extinction value. Following inoculation, the extinction value of the monochromatic radiation at 436 nm for a 10-mm layer of the bacterial suspension of the test cultures will correspond to the extinction value of the Formazin standard suspension TE/F/ 436 nm = 10. Make up the flasks of the dilution series that are not inoculated to 100 mL by adding 5 mL each of stock solution 1,5 mL each of stock solution II, and 10 mL of saline. Leave both inoculated and non-inoculated dilution series at 25°C for 16 h. After termination of the test period measure the extinction of the monochromatic radiation at 436 nm in a 10-mm layer in the inoculated dilution series. If after termination of the test period, colouration or turbidity occurs in the dilution series for chemical-physical reasons use the analogous steps of dilution of the non-inoculated series as photometric blank values for turbidimetry of the inoculated dilution series. For turbidimetry of heavily coloured waters it may be necessary in very rare cases to adjust the wavelength of the monochromatic radiation used for determination of the extinction of the dilutions to the spectral transmissivity of the substance to be measured by using monochromatic radiation at 578 nm or 691 nm instead of 436 nm. This does not apply to the wave-length of the monochromatic radiation used to determine the extinction of the bacterial inoculum.

- Parameters analysed / observed: Toxicity threshold

GLP compliance:

no

Analytical monitoring:

no

Vehicle:

no

Details on test solutions:

Trace elements solution (in grams per liter of double- distilled water)
0.055 AI2 (SO4)3*18 H2O; 0.028 K J, A.R. ; 0.028 KBr, A.R.; 0.055 TiO2 LAB; 0.028 SnCl2* 2 H2O, A.R. ; 0.028 LiCI, A.R.; 0.389 MnCI2*4 H2O, A.R.; 0.614 H3BO3, A.R.; 0.055 ZnSO4*7 H2O, A.R.; 0.055 CuSO4*5 H2O, A.R.; 0.059 NiSO4*6 H2O, A.R.; 0.055 Co(NO3)2*6 H2O, A.R.
Vitamin solution
0.2 mg biotin (as D + biotin); 2.0 nag nicotinic acid 1.0 mg thiamine (as thiamine HCI); 1.0 rag p-aminobenzoic acid; 0.5 mg panthothenic acid (as D-panthothenic acid, Na-salt); 5 mg pyridoxamine (as pyridoxamine dihydrochloride); 2.0 mg cyanocobalamin (vitamin B 12); 100 mL double-distilled water. Fill 6 mL each of the nutrient medium into culture tubes, sterilize the latter in a steam sterilizer by fractionated sterilization (three times) for 30min. Let solidify in slant position.
Stock solution I
20.000 g D(+) glucose (for biochemical and microbiological purposes); 4.240 g sodium nitrate, NaNO3, A.R.; 2.400 g dipotassium hydrogen phosphate, K2HPO4 anhydrous, high purity; after which add 3 mL of vitamin solution. 1.200 g potassium dihydrogen phosphate, KH2PO4, A.R.; 30 mL trace elements solution. Dissolve glucose and nutrient salts separately in 500 mL double-distilled water each, sterilize in a steam sterilizer for 30 min and unite solutions when cooled.
Stock solution II
Dissolve: 0.200 g ferrous sulphate, FeSO4*7H2O, A.R.; 4.000 g magnesium sulphate, MgSO4*7H2O, A.R. in 1000 mLsterile double-distilled water. Saline Dissolve: 0.500 g sodium chloride, NaCI, A.R. in 1000 mL double-distilled water. Sterilize solution in a steam sterilizer for 30 min.

Test organisms (species):

Pseudomonas putida

Details on inoculum:

- Laboratory culture: yes, not further described
- Method of cultivation: Keep stock cultures of the test strain, Pseudomonas putida, on the nutrient for stock and preliminary cultures in agar slant tubes.
- Preparation of inoculum for exposure: Prepare, for onward culturing of the test strain, new stock cultures at intervals of 1 week each. Incubate the inoculated stock cultures at 25°C for 24 h and keep in stock. If needed, prepare preliminary cultures from stock cultures on the above mentioned nutrient medium in agar slant tubes and incubate at 25°C for 24 h. Then, wash off the cell material with sterile saline.

Test type:

static

Water media type:

saltwater

Limit test:

no

Total exposure duration:

16 h

Key result

Duration:

16 h

Dose descriptor:

other: Toxicity threshold

Effect conc.:

1 550 mg/L

Nominal / measured:

nominal

Conc. based on:

test mat.

Basis for effect:

growth inhibition

Details on results:

- Any observations (e.g. precipitation) that might cause a difference between measured and nominal values: not reported
- Effect concentrations exceeding solubility of substance in test medium:
- Adsorption (e.g. of test material to the walls of the test container): not reported

Validity criteria fulfilled:

not applicable

Conclusions:

In the present test Pseudomonas putida were incubated with serial dilutions of Oxalic acid for 16h under static conditions. The toxicity threshold was determined to be 1550 mg/L for Pseudomonas putida. Based on this result the EC50 is above the limit concentrations of 100 mg/L. Oxalic acid is not considered to mediate any inhibitory effect on bacterial growth.

Executive summary:

In the present publication of Bringmann and Kühn (1978) the toxicity to microorganisms was determined according to the following method:

The multiplication of bacterial cells of the genus Pseudomonas is inhibited by dissolved toxic water ingredients. After a certain period the increase in the number of cells in a test culture free from toxic influence and with a defined standardized offer of nutrients will exceed that observed in a teat culture containing dissolved toxic substances and kept under identical conditions. The concentration of the bacterial suspension is measured turbidimetrically (while diffused light is screened off); it is expressed by the extinction of the primary light of the monochromatic radiation at 436 nm for a layer of 10 mm thickness. The concentration at which the inhibitory action of a pollutant starts will be present in that step of a dilution series of the pollutant having an extinction value at the end of the test period that is ≥ 3 % below the mean value of extinction for non-toxic dilutions of the test cultures.

The toxicity threshold after 16 h of incubation was reported for each substance tested. For Pseudomonas putida the toxicity threshold was determined to be 1550 mg/L. Based on this result the EC50 is above the limit concentrations of 100 mg/L. Oxalic acid is not considered to mediate any inhibitory effect on bacterial growth.

Description of key information

-published data by Brinkmann and Kühn, measurement of the inhibitory effect of several substances on Pseudomonas bacterial cells. Pseudomonas putida were eposed to serial dilutions of oxalic acid (exact values not reported) for 16h under static conditions in saltwater. The toxicit threshold was determined to be 1550 mg/L for Pseudomonas putida, read-across, oxalic acid, RL2

Key value for chemical safety assessment

EC10 or NOEC for microorganisms:

1 550 mg/L

Additional information

In the present publication of Bringmann and Kühn (1978) the toxicity to microorganisms was determined according to the following method:

The multiplication of bacterial cells of the genus Pseudomonas is inhibited by dissolved toxic water ingredients. After a certain period the increase in the number of cells in a test culture free from toxic influence and with a defined standardized offer of nutrients will exceed that observed in a teat culture containing dissolved toxic substances and kept under identical conditions. The concentration of the bacterial suspension is measured turbidimetrically (while diffused light is screened off); it is expressed by the extinction of the primary light of the monochromatic radiation at 436 nm for a layer of 10 mm thickness. The concentration at which the inhibitory action of a pollutant starts will be present in that step of a dilution series of the pollutant having an extinction value at the end of the test period that is ≥ 3 % below the mean value of extinction for non-toxic dilutions of the test cultures.

The toxicity threshold after 16 h of incubation was reported for each substance tested. For Pseudomonas putida the toxicity threshold was determined to be 1550 mg/L. Based on this result the EC50 is above the limit concentrations of 100 mg/L. Oxalic acid is not considered to mediate any inhibitory effect on bacterial growth.