2-ethyl-3-hydroxy-4-pyrone

Toxicity to microorganisms

Link to relevant study record(s)

Reference

Endpoint:

toxicity to microorganisms, other

Type of information:

experimental study

Adequacy of study:

key study

Study period:

15 January 1993 - 12 February 1993

Reliability:

2 (reliable with restrictions)

Rationale for reliability incl. deficiencies:

guideline study without detailed documentation

Remarks:

replicates are not given, no CoA, no calculations to verify

Reason / purpose for cross-reference:

reference to same study

Qualifier:

according to guideline

Guideline:

other: OECD Guideline 301 B (Ready Biodegradability: CO2 Evolution Test)

Version / remarks:

1981

Deviations:

yes

Remarks:

Adaptations in line with Birch, R. R., & Fletcher, R. J. (1991)

Principles of method if other than guideline:

Closed system was used in accordance with the publication of Birch, R. R., & Fletcher, R. J. (1991) The application of dissolved inorganic carbon measurements to the study of aerobic biodegradability. Chemosphere, 23, 855–872.

GLP compliance:

yes

Analytical monitoring:

yes

Remarks:

CO2 production (Inorganic carbon)

Details on sampling:

SAMPLING
- Sampling frequency: day 3, 7, 10, 14, 17, 21, 24, and 28
- Sampling method: Carbon dioxide was measured in the head space gas and inorganic carbon was measured in the liquid medium of 1 vessel. On the final test day more than one vessel is analysed for both the test compound and the control.

Vehicle:

no

Test organisms (species):

activated sludge

Details on inoculum:

Secondary effluent from an unacclimatised activated sludge plant at URL North. The inoculum was 10% by volume and was filtered through a Whatman filter paper to remove the coarse particulate matter. The level of dissolved inorganic carbon was reduced by sparging the filtered effluent with nitrogen at pH 6.5.

Test type:

static

Water media type:

freshwater

Limit test:

yes

Total exposure duration:

28 d

Test temperature:

18 - 21 °C

Nominal and measured concentrations:

9.19 mg/L organic carbon.

Details on test conditions:

TEST CONDITIONS
- Preparation stock solutions:
Stock solutions of the test substances with the exception of benzyl alcohol, were prepared by dissolving an appropriate mass in warm distilled water (approx. 55 °C) After shaking for 3 h in a closed vessel the stock solutions were filtered (Whatman 541 filter) and the total carbon (TC) concentration of the filtrate determined using an Ionics total carbon analyser. Since the test substance is volatile the acidification and sparging step in the analysis were omitted. Therefore, in order to determine the dissolved organic carbon concentration of the stock solutions, a water blank was also analysed by the same procedure to give an estimate of the total inorganic carbon content of the water used to prepare the stock solutions (assuming a minimal DOC content). The DOC concentrations of the stock solutions were determined by subtractions of the water blank value from the TC values of the stock solutions. A 5 mL volume of each test substance stock solution was added to the appropriate test vessels. Benzyl alcohol was added directly using a high precision volumetric syringe. (1.2 μL)

TEST SYSTEM
- Culturing apparatus: 160 mL sealed vessels hypovials containing 100 mL mineral salts medium in
oculated with secondary effluent and the respective test or reference substance. Vessels were incubated at 20°C on a rotary shaker.
- Number of culture flasks/concentration: 5 vessels per test/reference substance
- Method used to create aerobic conditions: The headspace in each vessel had a volume of 60 mL and when filled with air contains approximately 6 times the mass of oxygen required for the complete oxidation of the substance.
- Measuring equipment: Ionics 555 Inorganic Carbon Analyser
- Test performed in closed vessels due to significant volatility of test substance: yes, sealed with butyl rubber septum and an aluminium crimp seal.

SAMPLING
- Sampling frequency: day 3, 7, 10, 14, 17, 21, 24, and 28
- Sampling method: Carbon dioxide was measured in the head space gas and inorganic carbon was measured in the liquid medium of 1 vessel. On the final test day more than one vessel is analysed for both the test compound and the control.

CONTROL AND BLANK SYSTEM
The control vessels are identical to the test vessel except for the omission of the test substance.

STATISTICAL METHODS:
The percentage biodegradation is given by:
(TICt – TICc) /TC * 100%

TICt = μg Total Inorganic Carbon in test bottle
TICc = μg Total Inorganic Carbon in control bottle
TC = μg organic carbon added to test vessel as test compound.

The total inorganic carbon is the summation of the results of the analysis of the liquid and gas phases of each bottle after reference to the appropriate inorganic standard curve. On the final day more than 1 test vessel was analysed for both the test substance and reference substance. In this case the TICc is the arithmetic mean mass of inorganic carbon in the control vessels. The percentage biodegradation is calculated for each individual test vessel and the mean percentage biodegradation and standard deviation are determined and the results are expressed as:

Mean % biodegradation ± (S.t)/√n
S = the standard deviation
t = the ‘t’alue (two tailed test) at n-degrees of freedom at the 0.05 significance level
n = the number of results used to obtain the standard deviation

Reference substance (positive control):

yes

Remarks:

benzyl alcohol

Key result

Duration:

28 d

Dose descriptor:

NOEC

Effect conc.:

15.5 mg/L

Nominal / measured:

nominal

Conc. based on:

test mat.

Basis for effect:

inhibition of total respiration

Details on results:

The air temperature over the 28 day test period was in the range of 18 - 21 °C.
Benzyl alcohol in combination with the test susbtance achieved 95.8% biodegradation after 28-days and the 60% "pass" level was reached within 10 days of exceeding the 10% level. Biodegradation of Benzyl alcohol was not inhibited, confirming that the microorganisms of the innoculum are not effected by the test substance at a concentrations of 9.19 mg/L Organic carbon.

Results with reference substance (positive control):

n.a.

Validity criteria fulfilled:

not applicable

Conclusions:

The microorganisms in the inoculum are not affected by the test substance at a concentration of 9.19 mg/L organic carbon. The NOEC was determined to be 9.19 mg/L based on organic carbon, which would be 15.5 mg/L Ethyl Maltol.

Executive summary:

A ready biodegradability test was used to address the toxicity of Ethyl Maltol to microorganisms. A CO2 Evolution Test in accordance with OECD Guideline for Testing of Chemicals 301 B was performed under closed conditions in compliance with GLP. Carbon dioxide was measured in the head space gas and inorganic carbon was measured in the liquid medium on day 3, 7, 10, 14, 17, 21, 24, and 28. Benzyl alcohol was used as a reference substance and achieved 95.8% biodegradation after 28-days. The 60% "pass" level was reached within 10 days of exceeding the 10% level, confirming the suitability of the inoculum and the test conditions. Based on these results it can be concluded that the microorganisms in the inoculum are not affected by the test substance at a concentration of 9.19 mg/L organic carbon. The NOEC was determined to be 9.19 mg/L. Due to the organic carbon content being 59.4% in Ethyl Maltol (as determined in the daphnia study), this would correspond to 15.5 mg/L test substance.

Description of key information

A ready biodegradability test was used to address the toxicity of Ethyl Maltol to microorganisms. A CO2 Evolution Test in accordance with OECD Guideline for Testing of Chemicals 301 B was performed under closed conditions in compliance with GLP. Carbon dioxide was measured in the head space gas and inorganic carbon was measured in the liquid medium on day 3, 7, 10, 14, 17, 21, 24, and 28. Benzyl alcohol was used as a reference substance and achieved 95.8% biodegradation after 28-days. The 60% "pass" level was reached within 10 days of exceeding the 10% level, confirming the suitability of the inoculum and the test conditions. Based on these results it can be concluded that the microorganisms in the inoculum are not affected by the test substance at a concentration of 9.19 mg/L organic carbon. The NOEC was determined to be 9.19 mg/L. Due to the organic carbon content being 59.4% in Ethyl Maltol (as determined in the daphnia study), this would correspond to 15.5 mg/L test substance.

Key value for chemical safety assessment

EC10 or NOEC for microorganisms:

15.5 mg/L

Additional information

In the study the NOEC = 9.19 mg/L was based on organic carbon content, but for the key value in this dossier was recalculated as 15.5 mg Ethyl Maltol/L.