Ozonized olive oil

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:

Study initiation date 7 March 2019
Experimental starting date 8 March 2019
Experimental completion date 15 March 2019

Reliability:

1 (reliable without restriction)

Rationale for reliability incl. deficiencies:

guideline study

Qualifier:

according to guideline

Guideline:

OECD Guideline 201 (Alga, Growth Inhibition Test)

Version / remarks:

Guideline 201: Freshwater Alga and Cyanobacteria, Growth Inhibition Test, adopted March 2006, Annex 5 corrected July 2011.

Deviations:

no

GLP compliance:

yes

Analytical monitoring:

yes

Details on sampling:

Samples were taken from all test media including the control at the start of the test (without algae), after 24, 48 and 72 hours of exposure (containing algae). For sampling at the start of the test the samples were taken from the freshly prepared test solutions as well as from the control. For all subsequent sampling time points, the replicates per treatment were pooled before sampling. Algal cells were removed by centrifugation (4500 g, 10 min) prior to analysis.
The samples were analysed without further storage.

Vehicle:

yes

Details on test solutions:

Since the test item is a poorly soluble UVCB, the test solution consisted of the Water Soluble Fraction, prepared by addition of a stock solution of test item in hexane to sterile glassware, evaporation of the hexane, addition of sterile test water (section 4.2), stirring for about 96 hours, followed by filtration with glass fiber filter (MILLIPORE AP15). 100 ml of the resulting test solution were added to the sterile test flasks and inoculated with an exponentially growing preculture of algae.

Test organisms (species):

Desmodesmus subspicatus (previous name: Scenedesmus subspicatus)

Details on test organisms:

The test organism used for the study was Desmodesmus subspicatus 86.81 SAG, supplied by the SAG Culture Collection of Algae at Göttingen University, 37073 Göttingen, Germany.
Test conditions
Test vessel: 250 ml flasks, all-glass, with 100 ml of test medium, shaken (130 rpm), capped with air permeable stoppers.
Preculture: Exponentially growing liquid culture of Desmodesmus subspicatus kept under the same environmental conditions as in the test.
Illumination: Continuous, 4440 – 8880 lux ± max. 15% variation, source: Osram Fluora L 18W77 and Osram Daywhite L 18W840 (Osram AG, Winterthur, Switzerland).
Temperature: 21–24 °C maintained at ±2 °C in a thermo-controlled room.
pH: The pH of the control should not increase by more than 1.5 units during the test.
Nominal initial cell density: 6000 cells/ml corresponding to 0.005 OD680 units; all test media contain the same initial density of algal cells.

Test type:

static

Water media type:

other: Reconstituted water (OECD medium) prepared according to the test guidelines was used for algal cultivation and testing. Analytical grade salts were dissolved in sterile purified water

Limit test:

no

Total exposure duration:

72 h

Hardness:

The calculated water hardness of the test water is 24.2 mg/l as CaCO3.

Test temperature:

The calculated water hardness of the test water is 24.2 mg/l as CaCO3.

pH:

pH= 8.0,
The pH of the control should not increase by more than 1.5 units during the test.

Nominal and measured concentrations:

Nominal cocentration =100mg/L
Calculated test item concentration=2,62mg/L

Details on test conditions:

Test conditions
Test vessel: 250 ml flasks, all-glass, with 100 ml of test medium, shaken (130 rpm), capped with air permeable stoppers.
Preculture: Exponentially growing liquid culture of Desmodesmus subspicatus kept under the same environmental conditions as in the test.
Illumination: Continuous, 4440 – 8880 lux ± max. 15% variation, source: Osram Fluora L 18W77 and Osram Daywhite L 18W840 (Osram AG, Winterthur, Switzerland).
Temperature: 21–24 °C maintained at ±2 °C in a thermo-controlled room.
pH: The pH of the control should not increase by more than 1.5 units during the test.
Nominal initial cell density: 6000 cells/ml corresponding to 0.005 OD680 units; all test media contain the same initial density of algal cells.

Reference substance (positive control):

yes

Remarks:

For evaluation of the algal quality and the test procedure, potassium dichromate is tested as a positive control twice a year.

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

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

Details on results:

The microscopic examination of the algal cells at the end of the test showed no abnormal appearance of the algae growing at the loading rate of 100 mg/l. The appearance of the algal cells was not affected by the test item up to at least this concentration.
No remarkable observations were made concerning the appearance of the test media.

Reported statistics and error estimates:

Student T-test, one-sided smaller, alpha = 0.05

Validity criteria fulfilled:

yes

Conclusions:

The 72-hour EC50 based on average specific growth rate of Reaction products of olive oil and ozone (NOVOX) to Desmodesmus subspicatus was therefore estimated to be >100 mg/l, while the NOEC was determined to be 100 mg/l.

Executive summary:

The growth inhibitory effects of Reaction products of olive oil and ozone (NOVOX) (EC no. 919-979-9) to the green alga Desmodesmus subspicatus were investigated according to

OECD guideline 201 over a period of 72 hours. The test item Reaction products of olive oil and ozone (NOVOX) is a solid and poorly soluble UVCB substance.

The test solution consisted of the Water Soluble Fraction, prepared by addition of a stock solution of test item in hexane to sterile glassware, evaporation of the hexane, addition of sterile test water, stirring for 96 hours and finally filtration.

The single loading rate tested was 100 mg/l. Six parallel test vessels were used for the test item and six for the control.

The concentrations of Reaction products of olive oil and ozone (NOVOX) in the test media were analysed by DOC analytics at the beginning and after 24, 48 and 72 hours of exposure. The DOC determinations confirmed the very poor solubility of the test item. Conclusions about its stability cannot be drawn, since this type of analytics is non-specific. Therefore, the effective concentrations ECx were assessed based on the loading rate of the test item, which was 100 mg/l.

With respect to the endpoint average specific growth rate no significant effects were observed at 100 mg/l loading rate compared to the control.

With respect to the endpoint yield no significant effects were observed at 100 mg/l loading rate compared to the control.

The results of the average specific growth rate and yield inhibition of Reaction products of olive oil and ozone (NOVOX) to the green alga Desmodesmus subspicatus are summarized in the following table based on the loading rate of the test item:

Parameter (0-72 h)	Growth rate [mg/l]	Yield [mg/l]
EC50	>100	>100
NOEC	100	100

In conclusion, Reaction products of olive oil and ozone (NOVOX) (EC no. 919-979-9) had no significant inhibitory effects on the green algae Desmodesmus  ubspicatus up to its solubility limit in test water under the conditions of the test.

Description of key information

The growth inhibitory effects of Reaction products of olive oil and ozone (NOVOX) (EC no. 919-979-9) to the green alga Desmodesmus subspicatus were investigated according to

OECD guideline 201 over a period of 72 hours. The test item Reaction products of olive oil and ozone (NOVOX) is a solid and poorly soluble UVCB substance.

The test solution consisted of the Water Soluble Fraction, prepared by addition of a stock solution of test item in hexane to sterile glassware, evaporation of the hexane, addition of sterile test water, stirring for 96 hours and finally filtration.

The single loading rate tested was 100 mg/l. Six parallel test vessels were used for the test item and six for the control.

The concentrations of Reaction products of olive oil and ozone (NOVOX) in the test media were analysed by DOC analytics at the beginning and after 24, 48 and 72 hours of exposure. The DOC determinations confirmed the very poor solubility of the test item. Conclusions about its stability cannot be drawn, since this type of analytics is non-specific. Therefore, the effective concentrations ECx were assessed based on the loading rate of the test item, which was 100 mg/l.

With respect to the endpoint average specific growth rate no significant effects were observed at 100 mg/l loading rate compared to the control.

With respect to the endpoint yield no significant effects were observed at 100 mg/l loading rate compared to the control.

The results of the average specific growth rate and yield inhibition of Reaction products of olive oil and ozone (NOVOX) to the green alga Desmodesmus subspicatus are summarized in the following table based on the loading rate of the test item:

Parameter (0-72 h)	Growth rate [mg/l]	Yield [mg/l]
EC50	>100	>100
NOEC	100	100

In conclusion, Reaction products of olive oil and ozone (NOVOX) (EC no. 919-979-9) had no significant inhibitory effects on the green algae Desmodesmus  ubspicatus up to its solubility limit in test water under the conditions of the test.

Key value for chemical safety assessment

EC10 or NOEC for freshwater algae:

100 mg/L

Additional information

The substance is very poorly soluble in water and contains unstable peroxide that hydrolize in contact with water.

The 72-hour EC50 based on average specific growth rate of Reaction products of olive oil and ozone (NOVOX) to Desmodesmus subspicatus was therefore estimated to be >100 mg/l, while the NOEC was determined to be 100 mg/l.