2-Ethylhexyl trans-4-methoxycinnamate

Phototransformation in water

Link to relevant study record(s)

Reference

Endpoint:

phototransformation in water

Type of information:

experimental study

Adequacy of study:

key study

Study period:

20/06/2000 - 18/12/2002

Reliability:

1 (reliable without restriction)

Rationale for reliability incl. deficiencies:

other: Study according to international guideline (US-EPA and SETAC) under GLP.

Study type:

direct photolysis

Qualifier:

according to guideline

Guideline:

EPA Guideline Subdivision N 161-2 (Photodegradation Studies in Water)

Deviations:

no

Qualifier:

according to guideline

Guideline:

other: SETAC. procedures for assessing the environmental fate and ecotoxicity of pesticides. Aqueous photolysis (1995).

Deviations:

yes

Remarks:

1) 20% acetonitrile was used in the aquaous testmaterial solution to prevent hydrolysis, although the guideline specifies <1% organic solvent. 2) Less than 90% mass balance could be recovered. 3) The identity of not all metabolites >10% has been recovered

Qualifier:

equivalent or similar to guideline

Guideline:

OECD Guideline draft (Phototransformation of Chemicals in Water - Direct and Indirect Photolysis)

Version / remarks:

Guideline has been consulted

Deviations:

yes

Remarks:

1) 20% acetonitrile was used in the aquaous testmaterial solution to prevent hydrolysis, although the guideline specifies <1% organic solvent. 2) Less than 90% mass balance could be recovered. 3) The identity of not all metabolites >10% has been recovered

Principles of method if other than guideline:

According to SETAC guideline the study should be conducted in a buffer with <1 % organic solvent. However, incubation of the test material in aqueous buffer caused problems and the photolysis test had to be conducted with a test solution in 20 % acetonitrile and 80 % buffer solution.

GLP compliance:

yes

Radiolabelling:

yes

Analytical method:

other: TLC

Details on sampling:

- Sampling intervals for the parent/transformation products: The dark control was sampled at t = 0 and after 1.1; 2.8; 5.8 and 8.0 days of incubation. After 8 days of incubation had stopped and data from a hydrolysis study had to be used for dark control. The first irradiated solution was taken at t = 0; and after 1.1; 2.8; 5.8 and 8.0 days of incubation.
In a second experiment , a closed setup was used (second light incubation). Samples were taken at t=0; and after 2.5; 4.3; 5.3; 21.5; 28; 46; 52; 119; 214; 288 and 551 hours of incubation.

- Sampling method: Samples for LSC (200 µL duplicate samples) and TLC analysis (0.5 mL) were taken

- Sampling methods for the volatile compounds, if any: Humidified, CO2-free air passed over the test solution and after that through two traps (A and B) containing 100 mL of 2 M aqueous NaOH solution each to collect 14CO2 followed by one trap (C) containing 100 mL of 2-methoxyethanol to trap volatile organic compounds. The volatile organic compounds and 14CO2 traps were sampled and replaced by fresh traps at each sampling point. Radioactivity in each trap was determined by liquid scintillation counting of a 1 mL aliquot in Ultima Gold scintillation cocktail. This was done for the dark control and the first incubation in light only. No traps were used for the second incubation in light, since the radioactivity detected in the traps of the fist light incubated 2-Ethylhexyl 4-methoxycinnamate solution never exceeded 2 %.

- Sampling intervals/times for pH measurements: pH of the test solutions were measured at t = 0, and after 6 and 8 days of incubation in the dark control and at t = 0 and after 6 and 8 days of incubation in the first irradiated solution. For the second irradiated solution, the pH measurements were taken at t = 0 and after 23 days of incubation.

- Sampling intervals/times for sterility check: Sterility of the test solutions were measured at t = 0, and after 6 and 8 days of incubation for the dark control and at t = 0 and after 6 and 8 days of incubation for the first irradiated solution. For the second irradiated solution, the sterility measurements were taken at t = 0 and after 23 days of incubation.

- Sample storage conditions before analysis: Chromatographic analysis was performed on the sampled aliquots as such and they were not subjected to any extraction procedure.

Buffers:

- pH: 7
- Type and final molarity of buffer: Type: sterilised by autoclaving, molarity not calculated.
- Composition of buffer: 13.6 g KH2PO4 in 2 liter Milli-Q water, adjusted to pH 7 with NaOH

Light source:

Xenon lamp

Light spectrum: wavelength in nm:

>= 290 - <= 800

Relative light intensity:

ca. 0.9

Details on light source:

- Emission wavelength spectrum: 300 - 800 nm
- Filters used and their purpose: quartz filter, cut-off below 290 nm
- Light intensity at sample and area irradiated: 55.6 watt/m², 48 cm²
- Relative light intensity based on intensity of sunlight: one day of irradiation was equivalent to 0.093 days of natural sunlight at 40 degrees
- Duration of light/darkness: Continuous light for 8 or 23 days (first and second irradiation study respectively)
- Other: for calculations, the spectrum 290 - 400 nm was used, since the test material did not absorb above 400 nm. Light intensity of the lightsource was determined by using a ferri-oxalate actinometre. This reacts to Fe2+ when irradtiated with blue and UV light, its concentration being linearly proportional to the amount of blue and UV photons absorbed. The amount of Fe2+ was determined spectrophotometrically at 510 nmthe actinometre absorbs between 290 nm and 527.5 nm and its quantum yield is that range is 1.1.
Tabulated values of light intensity in summer at 40 degrees N from 300 - 400 nm were used to calculate the relative light intensity of the lamp versus sunlight.

Type of sensitiser:

not specified

Details on test conditions:

TEST SYSTEM
- Type, material and volume of test apparatus/vessels: Suntest CPS+ with xenon light source (simulating natural daylight). Double walled vessels where used, with quartz lid
- Sterilisation method: autoclave, 121 degrees at 1 bar, 20 minutes. air: 0.2 microm sterile filter. Pipette tips: rinsing in acetonitrile before use.
- Measures to saturate with oxygen: humidified CO2 free air was passed over the test solutions after passing: a 1 L flask with approx. 500 mL 2M NaOH solution(aq), a 250 mL gas washing bottle with approx. 100mL 0.1 M NaOH solution with phenolphthalein, an empty gas washing bottle.
- Details on test procedure for unstable compounds: 20 % acetonitrile was used in the buffer solution
- Details of traps for volatile, if any: In initial test: The volatile organic compounds and 14CO2 traps were sampled and replaced by fresh traps at each sampling point. Radioactivity in each trap was determined by liquid scintillation counting of a 1 mL aliquot in Ultima Gold scintillation cocktail. Radioactivity was determined in the dark control and the first incubation in light only. No traps were used for the second incubation in light, since the total radioactivity detected in the traps of the first light incubated 2-Ethylhexyl 4-methoxycinnamate solution never exceeded 2 %.
- If no traps were used, type of test system: Second test: closed system: No traps were used for the second incubation in light, since the total radioactivity detected in the traps of the fist light incubated 2-Ethylhexyl 4-methoxycinnamate solution never exceeded 2 %.
- Indication of test material adsorbing to the walls of test apparatus: yes, see Results and discussion: test performance.

TEST MEDIUM
- Volume used/treatment: 225 mL
- Kind and purity of water: Milli-Q water
- Preparation of test medium:

A stock solution (StA) of 14C-labelled 2-Ethylhexyl 4-methoxycinnamate was prepared in acetonitrile. Its radioactivity was determined to be 10.64 MBq/mL by LSC (n = 1). According to TLC analysis (I and II), the radiochemical purity was determined to be 93 - 95 %. Hence, the test substance was purified according to the following procedure (June 21, 2000): At least 0.5 % of StA was spotted (line) on a Si60 TLC plate and developed in hexane:tert-butylmethylether 2:1. v:v. After identification, test substance was scraped off the plate and vortexed with 1 mL acetonitrile during 2 minutes in a 7 mL picovial covered with aluminum foil. The supernatant was transferred to a reagent tube using a glass pipette. A second extraction of scrapped of SiO2 TLC plate was performed using 0.5 mL acetonitrile during 1 minute followed. Both supernatants obtained from 1st and 2nd extraction were combined. After centrifugation for 5 minutes by 2500 rpm (= 1181 g) the supernatant was transferred to a 7 mL picovial, and coded StB (5.65 MBq/mL, purity: 98.6 - 98.7 %).
An aliquot of two mL of acetonitrile was added to StB: radioactivity was 1.27 MBq/mL (triple analysis, relative standard deviation 1.87 %). An aliquot of 394 µL of StB was added to 100 mL acetonitrile in a sterile measuring flask of 500 mL. The solution was made up using sterile buffer pH 7 (1) to a volume of 500 mL. The flask, coded as StD, was covered in aluminum foil. (July 4th 2000, 14:15 hr, t=0 dark control). Triple LSC analysis gave 896 Bq/mL (= 0.11 mg/L, 3.6*10' M), 1.17 % relative standard deviation. The pH was 7.22.

A second incubation was performed later (closed set-up). The test solution for the closed set-up was prepared as follows: One mL of StA was spotted (5 cm) on a Si-60 TLC plate. The plate was eluted with 2:1 hexane: tert. Butylmethylether v:v. The test substance 2-Ethylhexyl 4-methoxycinnamate was scraped off the plate after identification and vortexed with 1 mL acetonitrile for 2 minutes. The supernatant was centrifuged at 2500 rpm (= 1181 g). The supernatant was transferred to a 7 mL picovial covered with aluminum foil. The extraction procedure was repeated and the extracts were combined: StE (5.70 MBq/mL, 100 % purity by TLC I and II).
An aliquot of 43.9 µL of StE was transferred into a 50 mL amber measuring flask (sterile) and made up with acetonitrile, coded StE* (5.06 kBq/mL, rel stdev 0.27 %, n = 3).
30.21 g of StE* was transferred to the sterilized test vessel. 160 mL buffer pH 7 (2) was added: test solution weighed 189.57 g (198 mL). Nominal radioactivity: 987 Bq/mL, 0.12 mg/L, 3.96*10' M. The radioactivity measured at t = 0 was 904 Bq/mL (91.7 % of applied, 0.11 mg/L, 3.63*10'M) n = 3, rel stdev 1.84 %.

- Identity and concentration of co-solvent: acetonitrile
- Concentration of solubilising agent: 20 %

REPLICATION
- No. of replicates (dark): 1 plus the prior hydrolysis test which was also used as a dark replicate.
- No. of replicates (irradiated): 2 (1 for 8 days, 1 for 23 days)

Duration:

8 d

Temp.:

25.4 °C

Duration:

23 d

Temp.:

23.2 °C

Initial conc. measured:

0.12 mg/L

Reference substance:

no

Dark controls:

yes

Remarks:

8 days, stable

Computational methods:

- Calculation of molar absorption coefficients (e?) utilizing Beer-Lambert¿s relationship: Yes
- Experimental photolysis reaction rate (direct and/or indirect photolysis): Based on chromatographic analysis of the second irradiation experiment (23 days) using TLC method 1. A lower limit based on the % distribution of radioactivity *on plate* and a upper limit based on the % distribution as *applied radioactivity* were calculated in view of the high adsorption of the substance onto the test materials. All results from the experiment where fitted to simple first order kinetics. This was done for the % degradation "on plate" and for the degradation expressed "as applied radioactivity". Since the dark control showed no degradation, subtracting this from the calculated rates was the same as the calculated photolysis rates.
- Quantum yield calculations (quantum yield should be expressed as a fraction and not as a percent): Quantum yield was obtained by dividing the initial photolytic rate constant by the total number of photons absorbed per second by the test solution: The [(rate/ (24*3600) * (3.96*10-7)] in (k/sec)*(mol/L)
- GCSOLAR or ABIWAS calculation of reaction rates at different latitudes, seasons, time of the day, water bodies: No
- Half-lives or DT50, DT75, and DT90 values and appropriate associated confidence intervals: Mt = M0 . e^-kt and DT50 = Ln(2)/k

Preliminary study:

According to the SETAC-guideline (and OECD draft guideline) an aquatic photo-degradation study should be conducted for a test compound in buffer with <1 % organic solvent. However, incubation of the test substance in aqueous buffer caused problems and the photolysis test had to be conducted with a test solution of 20 % acetonitrile and 80 % buffer.

Test performance:

Mass balance: rinsing the stirring bar with 10 mL acetonitrile detected 1 % and rinsing the teflon tape with cork using 5 mL acetonitrile detected 2.6 % of applied radioactivity. This illustrates some adsorption behaviour of 2-Ethylhexyl 4-methoxycinnamate onto test materials, which might have obstructed the acquirement of proper mass balance results, i.e. >90 %.
Temperature was monitored throughout the experiments for both irradiated solutions and dark control and remained 22.3 - 24.1 °C for the irradiated solution (second irradiation study) and 24.3 - 24.5 °C for the dark control.
Sterility and pH were monitored throughout the experiment at t = 0 and after 6 and 8 days for the dark control and at t = 0 and after 23 days for the second irradiation study. Sterility was tested by adding 0.5 - 2 mL of test solution to 2 - 5 mL of Brain heart Infusion medium followed by incubation of the mixture for 48 hours at 37 °C. Absence of turbidity in the solution was taken as proof of sterility.

Parameter:

max lambda

Value:

307.5 nm

Remarks:

see table 6 in attached background material

Parameter:

max epsilon

Remarks:

see figure 3 in attached background material

% Degr.:

> 99

Sampling time:

21.4 d

Test condition:

second irradiation study, result for degradation of trans-2-Ethylhexyl 4-methoxycinnamate

% Degr.:

50

St. dev.:

0.97

Sampling time:

8.43 d

Test condition:

second irradiation study, calculated % = on plate, parent compound: cis-and-trans 2-Ethylhexyl 4-methoxycinnamate corrected to 40degrees N sunlight

% Degr.:

50

St. dev.:

0.99

Sampling time:

6.91 d

Test condition:

second irradiation study, calculated % = on plate, parent compound: trans 2-Ethylhexyl 4-methoxycinnamate degradation to metabolites, corrected to 40degrees N sunlight

% Degr.:

50

St. dev.:

1

Sampling time:

5.06 d

Test condition:

second irradiation study, calculated % = of applied, parent compound: trans 2-Ethylhexyl 4-methoxycinnamate degradation to metabolites, corrected to 40degrees N sunlight

% Degr.:

50

St. dev.:

0.99

Sampling time:

5.83 d

Test condition:

second irradiation study, calculated % = of applied, parent compound: cis-and trans 2-Ethylhexyl 4-methoxycinnamate , corrected to 40degrees N sunlight

Test condition:

Please See Calculated Dissipation figure 14, 15,16 and table 20 in attached background material for background of this information.

Quantum yield (for direct photolysis):

0

Rate constant (for indirect photolysis):

>= 0.077 - <= 0.111 other: in 1/day. Even though it is direct photolysis, a rate constant was calculated: see table 20 in attached background materials

DT50:

>= 5 - <= 9 d

Test condition:

second irradiation study in 20 % acetonitrile. Rate based on degradation from trans and cis-2-Ethylhexyl 4-methoxycinnamate to metabolites. The trans-to-cis transformation had a DT50 of ca 0.05 days.

Transformation products:

yes

No.:

#1

No.:

#2

No.:

#3

No.:

#4

No.:

#5

No.:

#6

No.:

#6

No.:

#6

No.:

#7

Details on results:

Degradation products #4, #5 and #7 remained under 10 % of applied radioactvity throughout the incubation.
#1 reached a maximum of 36.3 % of applied after 23 days
#2 reached a maximum of 64.5 % of applied after 0.1 day
#3 reached a maximum of 11.8% of applied after 23 days.
The reaction mechanism proposed by the authors is rapid photolytic conversion of trans-2-Ethylhexyl 4-methoxycinnamate into cis-2-Ethylhexyl 4-methoxycinnamate (and possibly reverse) followed by further slower photolytic degradation into the metabolites.

Remark on reaction rates, DT50 and quantum yields in table 20 and figure 14, 15 and 16 (see attached background material). Results where calculated based on TLC I, second irradiation study results.A major part of the activity appeared to adsorb onto testing equipment.Part of the adsorbed activity could be resolved (rinsing with organic solvents), but the rinsates were not analysed on TLC to retreive distribution of parent and metabolites. Hence, in order to calculate the degradationrate of 2-Ethylhexyl 4-methoxycinnamate the following two approaches were used:

1) "On plate": (low limit): the adsorbed activity could show the same distribution as found in the test solution. It was calculated using % distribution on plate.

2) "Of applied": (high limit): The adsorbed activity could consist of metabolites only. In this case degradation rate was calculated using % distribution of applied.

According to the authors thus a lower limit and a higher limit could be found, in wich region the actual degradation rate can be found.

Since the dark control did not show degradation, the degradation rates as obtained were equal to the final photolytic rates.

The quantum yield reported was between 1*10 ^-5 and 2*10 ^-5.

Conclusions:

The authors conclude that according to the classification scheme by Mensink, trans-2-Ethylhexyl 4-methoxycinnamate can be regarded as being very rapidly degradable when transition to cis-2-Ethylhexyl 4-methoxycinnamate is taken into account. When trans-cis transition is left out, 2-Ethylhexyl 4-methoxycinnamate can be classified as fairly biodegradable with a DT50 in the range of ca. 5 - 9 days. The main photolytic degradation products are p-methoxyzimtsaure (34.8 % applied initial substance), unidentifiable substances M1 and M3 (according to TLC method I) or MA, MB and MD (according to TLC method II). The quantum yield for the trans-to-cis conversion was 0.0021. The quantum yield for the degradation into metabolites appeared to be in the range of 1*10-5 to 2*10-5.

Executive summary:

The photolytic degradation of 2-Ethylhexyl 4-methoxycinnamate was studied by irradiation of a test solution of 0.12 mg/L 2-Ethylhexyl 4-methoxycinnamate in milli-Q water and 20 % acetonitrile (to prevent hydrolysis) with a xenon lamp for 23 days, which was equivalent with 21 days in sunlight in a closed system .The test substance was radioactivily marked on its phenyl ring. TLC and a scintillation counter were used for seperation and identification of parent and metobalites.TLC plates were scanned and areas integrated for quantification. The mass balance decreased from 92 % at t = 0 to 52 % at t = 23 days, according to the authors due to adsorption to the test equipment. Comparison of the retention of reference substances and the metabolites on TLC was used for identification. pH temperature and sterility were monitored and kept stable. Light intensity was verified with an actinometre. See Conlusions for the final results.

Description of key information

Phototransformation in water: DT50 = 5 - 9 days (EPA Guideline Subdivision N 161-2 (Photodegradation Studies in Water))

Additional information

According to the classification scheme by Mensink, trans-2-Ethylhexyl 4-methoxycinnamate can be regarded as being very rapidly degradable when transition to cis-2-Ethylhexyl 4-methoxycinnamate is taken into account.

When trans-cis transition is left out, 2-Ethylhexyl 4-methoxycinnamate can be classified as fairly degradable with a DT50 in the range of ca. 5 - 9 days. The main photolytic degradation products are p-methoxycinnamate (34.8 % applied initial substance), unidentifiable substances M1 and M3 (according to TLC method I) or MA, MB and MD (according to TLC method II). The quantum yield for the trans-to-cis conversion was 0.0021. The quantum yield for the degradation into metabolites appeared to be in the range of 1*10^-5 to 2*10^-5.

It is concluded that the test substance is fairly degradable by phototransformation in water.