Reaction mass of (9-acetoxy-3,8,10-triethyl-7,8,10-trimethyl-1,5-dioxa-9-azaspiro[5.5]undec-3-yl)methyl palmitate and (9-acetoxy-3,8,10-triethyl-7,8,10-trimethyl-1,5-dioxa-9-azaspiro[5.5]undec-3-yl)methyl stearate

Toxicity to microorganisms

Link to relevant study record(s)

Reference

Endpoint:

activated sludge respiration inhibition testing

Type of information:

experimental study

Adequacy of study:

key study

Study period:

2003

Reliability:

1 (reliable without restriction)

Rationale for reliability incl. deficiencies:

other: GLP-guideline study with analytical monitoring

Qualifier:

according to guideline

Guideline:

OECD Guideline 209 (Activated Sludge, Respiration Inhibition Test

Deviations:

no

Qualifier:

according to guideline

Guideline:

EU Method C.11 (Biodegradation: Activated Sludge Respiration Inhibition Test)

Deviations:

no

GLP compliance:

yes (incl. QA statement)

Analytical monitoring:

no

Vehicle:

no

Details on test solutions:

The final nominal concentrations tested were 6.25, 12.5, 25, 50 and 100 mg/L. Concentrations exceeding 100 mg test item per liter were not tested.

According to the results of a solubility pre-test, no stock solution could be prepared due to the limited water solubility.

Therefore, test item amounts of 3.15, 6.39,12.37, 25.47 and 52.05 mg were first weighed (by means of an analytical balance) onto glass slides and thereafter transferred into the respective test flasks with 284 mL of tap water. The test item was mixed into the tap water by ultrasonic treatment for
ten minutes and intense stirring for 24 hours at room temperature in the dark to dissolve a maximum amount of the test item and/or disperse it as
homogeneously as possible. Then, 16 mL of synthetic wastewater and 200 mL of the activated sludge inoculum were added.

Additionally, two controls (tap water, synthetic wastewater and activated sludge inoculum, without test item) and the reference item
3,5-dichlorophenol (positive control) at nominal concentrations of 5, 16 and 50 mg/L were tested in parallel under otherwise identical conditions
of the test.

For test media containing the reference item, a stock solution of 3,5-dichlorophenol was prepared according to the test guidelines: 0.5 g of
3,5-dichlorophenol was dissolved in 10 mL 1 M NaOH and diluted to about 30 mL with purified water. Excess of NaOH was neutralized with 0.5 mol/L H2SO4 to the point of incipient precipitation. Thereafter, the mixture was made up to one liter with purified water. The pH was determined to be 7.5.
The final concentration of 3,5-dichlorophenol amounted to 500 mg/L. Aliquots of this 3,5-dichlorophenol stock solution were mixed with synthetic
wastewater and tap water in the respective test flasks to obtain the desired concentrations.

Test organisms (species):

activated sludge of a predominantly domestic sewage

Details on inoculum:

The study was performed with aerobic activated sludge from a wastewater treatment plant (ARA Ergolz II, Fullinsdorf, Switzerland) treating
predominantly domestic wastewater. The sludge was washed twice with tap water by centrifugation and the supernatant liquid phase was decanted.
A homogenized aliquot of the final sludge suspension was weighed, thereafter dried and the ratio of wet to dry weight was calculated.

Test type:

static

Water media type:

freshwater

Limit test:

no

Total exposure duration:

3 h

Nominal and measured concentrations:

6.25, 12.5, 25, 50 and 100 mg/L (nominal)

Details on test conditions:

The test was performed in 1000 mL glass flasks. The test flasks were labeled with the necessary infonnation to ensure unmistakable identiflcation.

At the start of the test, 200 mL activated sludge inoculum with a sludge concentration of 2.7 g/L dry weight (corresponding to about 1.1 g dry
material per liter test medium) was added. The sludge was added in time intervals of 15 minutes (an arbitrary but convenient interval) flrst to a
control, then to the test solutions of the reference item, then to the test solutions of the test item, and finally to the second control.

During the incubafion period of exactly 3 hours all test media and the controls were continuously aerated with compressed air at a flow of
approximately one liter per minute. The concentrafion of dissolved oxygen did not drop below 2.5 mg/L during the incubation period,
and just before measurement of the respiration rates the dissolved oxygen concentrafions were at least 8.4 mg/L. The temperature in the test
media, measured in one control, was 20 °C at the start and at the end of the incubafion period.

Duration:

3 h

Dose descriptor:

NOEC

Effect conc.:

100 mg/L

Nominal / measured:

nominal

Conc. based on:

test mat.

Basis for effect:

inhibition of total respiration

Remarks:

respiration rate

Duration:

3 h

Dose descriptor:

EC50

Effect conc.:

> 100 mg/L

Nominal / measured:

nominal

Conc. based on:

test mat.

Basis for effect:

inhibition of total respiration

Remarks:

respiration rate

Description of key information

The inhibition of the degradation activity of activated sludge is not anticipated when introduced in appropriate low concentrations.

Key value for chemical safety assessment

EC50 for microorganisms:

100 mg/L

EC10 or NOEC for microorganisms:

100 mg/L

Additional information

According to regulation (EC) No 1907/2006, Annex XI, paragraph 1.5., substances whose physicochemical, toxicological and ecotoxicological properties are likely to be similar or follow a regular pattern as a result of structural similarity may be considered as a group, or "category" of substances. Environmental effects or environmental fate may be predicted from data for reference substance(s) within the group by interpolation to other substances in the group (read-across approach). The substance consists of two main components, a component with a C18 chain (60%) and C16 chain (30%) component. The structure of both substances is the same except the chain length.

The substance with C18 is already notified under EC 445-990-9. It is expected that the structural difference between the octadecanoate and the hexadecanoate does not affect the physico-chemical properties, the toxicological and ecotoxicologocal profile as well as the environmental fate. Furthermore, it can be assumed that the degradation products of EC 445-990-9 are very similar to the degradation products of the mixture containing the octadecanoate and the hexadecanoate. Therefore read-across to EC 445-990-9 is scientifically justified for physico-chemical properties, the toxicological and ecotoxicologocal profile as well as environmental fate.