Reaction mass of methyl 2-[[(E)-(2,4-dimethylcyclohex-3-en-1-ylidene)methyl]amino]benzoate and methyl 2-[[(Z)-(2,4-dimethylcyclohex-3-en-1-ylidene)methyl]amino]benzoate and oxydipropanol

Hydrolysis

Link to relevant study record(s)

Reference

Endpoint:

hydrolysis

Type of information:

experimental study

Adequacy of study:

supporting study

Study period:

2009

Reliability:

4 (not assignable)

Rationale for reliability incl. deficiencies:

other: non-GLP; well documented study report following a screening study and method equivalent or similar to guideline with acceptable deviations according to the regulatory conclusion that the substance is hydrolytically unstable in specific pH ranges.

Remarks:

The study would not fulfil the requirements of OECD TG 111 (2004) Tier 2 or Tier 3 requirements. The test was conducted at physiologically relevant temperature (40 °C) rather than the guideline preliminary test temperature 50 °C.

Reason / purpose for cross-reference:

other:

Qualifier:

equivalent or similar to guideline

Guideline:

OECD Guideline 111 (Hydrolysis as a Function of pH)

Deviations:

yes

Remarks:

Screening study examined: pH 2, 5, 7, 8.5 and 12 rather than guideline pH range 4, 7 and 9; duplicate tests were not performed; full analytical validation was not documented within the study; degradation products were not identified.

Qualifier:

equivalent or similar to guideline

Guideline:

EU Method C.7 (Degradation: Abiotic Degradation: Hydrolysis as a Function of pH)

Deviations:

yes

Remarks:

see above.

Principles of method if other than guideline:

The test followed a method equivalent or similar to OECD TG 111 (hydrolysis as a function of pH) - as a screening study for the hydrolysis properties of the test substance. The screening study examined: pH 2, 5, 7, 8.5 and 12 using suitable buffers; at 40 °C for 28d by analytically monitoring by GC-FID the peak that represented constituents 1 and 2. The degradation is representative of (i) worst case kinetics and (ii) represents a simple majority on %wt terms of the substance in aqueous environment.

GLP compliance:

no

Radiolabelling:

no

Analytical monitoring:

yes

Details on sampling:

- Sampling intervals for the parent/transformation products: on a regular basis throughout the test (typically at time = 0, 0.25, 1, 2, 4, 7, 15, 21 and 28 days
- Sampling method: Small aliquots of the test solution are extracted with an organic solvent (typically cyclohexane or ethyl acetate) containing a hydrocarbon standard (typically C12, C17 or C20). The extracts are analyzed by GC-FID and the results are plotted as (Area/Area Std) expressed in [%]. The measurement at time = 0 is set at 100% and the succeeding measurements are calculated relatively to the time = 0 measurement. Therefore the curves represent the percentage of product remaining in the test solution at the time of analysis.
- Other observation, if any (e.g.: precipitation, color change etc.): None reported.

Buffers:

- pH: test media are standard aqueous buffers at pH 2, pH 5, pH 7, pH 8.5 and pH 12 containing 1% of non ionic surfactant (Arkopal N 150).
- Type and final molarity of buffer: Molarity not reported.
• pH 2 (± 0.1) buffer : Reference Handbook of Chemistry and Physics buffer type A*
• pH 5 (± 0.1) buffer : Reference Handbook of Chemistry and Physics buffer type C*
• pH 7 (± 0.1) buffer : Reference Handbook of Chemistry and Physics buffer type D*
• pH 8.5 (± 0.1) buffer : Reference Handbook of Chemistry and Physics buffer type F*
• pH 12 (± 0.1) buffer : Reference Handbook of Chemistry and Physics buffer type I*

Details on test conditions:

TEST MEDIUM
- Volume used/treatment: Total volume not reported
- Kind and purity of water: Deionised water (no further details reported)
- Preparation of test medium: ; 200 – 300 ppm of raw material are dissolved in the pH buffer containing the surfactant (Arkopal N 150) and put into storage in an oven at 40°C.
- Renewal of test solution: None reported.
- Identity and concentration of co-solvent: non-ionioc surfactant (Arkopal N 150) was used at 1% concentration
OTHER TEST CONDITIONS
- Adjustment of pH: None reported.
- Dissolved oxygen: Not reported.

Number of replicates:

None (with occasional re-injection if required).

Positive controls:

no

Negative controls:

no

Preliminary study:

For constituent 1:
At pH 2: At 1 day there is >90% disappearance
At pH 5: At 1 day there is ca. 40% disappearance and 48 hours ca. 60% At 5 days there is 80% disappearance. At 7.5 days there is 90% disappearance and complete disappearance at 15 days.
At pH 7: The substance appears relatively stable (< 10% degradation up to 28 days).
At pH 8.5: The substance appears relatively stable (< 10% degradation up to 28 days).
At pH 12: At 1 day there is ca. 10% disappearance and 48 hours ca. 20% At 5 days there is2% disappearance. At 7.5 days there is 30% disappearance. At 15 days there is 50% disappearance. At day 28 there is 70% disappearance.

It can be concluded that under the conditions of the present test, test substance is not hydrolytically stable (as defined in the OECD TG 111 for hydrolysis as a function of pH) at acidic pH.

Test performance:

No unusual findings were reported in the study.

Transformation products:

not measured

pH:

5

Temp.:

40 °C

DT50:

ca. 48 h

Remarks on result:

other: constituent 1

pH:

7

Temp.:

40 °C

DT50:

> 28 d

Remarks on result:

other: constituent 1

pH:

8.5

Temp.:

40 °C

DT50:

> 28 d

Remarks on result:

other: constituent 1

Details on results:

TEST CONDITIONS
- pH, sterility, temperature, and other experimental conditions maintained throughout the study: Yes, by use of appropriate buffers however pH was not continuously monitored throughout the study, sterility was not examined.

Validity criteria fulfilled:

yes

Remarks:

The study meets the validity criteria and reliable with limitations. This is limited as detailed in 'Rationale for reliability incl. deficiencies'. Guideline Tier 2 and 3 requirements are not met.

Conclusions:

The test item was found to be unstable to hydrolysis in water at pH 5.0. Under the conditions of the study, the test item is hydrolytically stable at pH 7 and pH 8.5 up to 28 days.

Executive summary:

The hydrolytic stability of the test item was investigated using a method similar or equivalent to OECD TG 111 (hydrolysis as a function of pH) and EU Method A.7. The test media are standard aqueous buffers at pH 2, pH 5, pH 7, pH 8.5 and pH 12 containing 1% of non-ionic surfactant (Arkopal N 150). The tests are done in accelerated conditions at 40°C for approximately one month (28d). 200 – 300 ppm of test substance are dissolved in the pH buffer containing the surfactant and put into storage in an oven at 40°C. Commercial reference grades of buffer are utilised as listed in documented literature sources. Small aliquots of the test solution are extracted with an organic solvent (typically cyclohexane or ethyl acetate) containing a hydrocarbon standard (typically C12, C17 or C20) on a regular basis throughout the test (typically at time = 0, 0.25, 1, 2, 4, 7, 15, 21 and 28 days). The extracts are analysed by GC-FID. Under the analytical conditions employed constituent 1 isomer was eluted and the results were then plotted with time to show the degradation curves of the substance. For constituent 1 of the test item at pH 5 there is evidence of hydrolytic instability of the test item. The test item rapidly degrades to 90% degradation within 10 days and by day 15 there is < 1% test item remaining. The t1/2 appears to be 48 hours. At pH 7 and pH 8.5 the test item appears under the conditions of the test, relatively stable up to 28 days with less than 10% degradation. It can be concluded that under the conditions of the present test the test substance is not hydrolytically stable as defined in the OECD TG 111 for hydrolysis as a function of pH at acidic pH ranges. Applicant assessment of the data indicates unexpected stability at pH 7 which may be influenced by the presence of constituent 3 preventing degradation within the test system.

Description of key information

Substance half-life for hydrolysis: pH 7: t1/2: < 24 hours ; pH 8.5 : t1/2: < 24 hours, at 20 °C, 1 atm, OECD TG 105, 2016

Substance half-life for hydrolysis: pH 5: t1/2: ca. 48 hours ; pH 7: t1/2: > 28d; pH 8.5: t1/2: > 28d, at  40 °C, 1 atm, eq. or similar to OECD TG 111, 2009

 

Weight of evidence: the test item contains hydrolytically unstable constituents (pH 5 to 8.5, t1/2 < 24 hours) and stable constituent: oxydipropanol (pH 5 to 8.5, t1/2 > 28 days).

Additional information

Hydrolysis:

Eq. or similar to OECD TG 111 - preliminary screening study, 2009: The hydrolytic stability of the test item was investigated using a method similar or equivalent to OECD TG 111 (hydrolysis as a function of pH) and EU Method A.7. The test media are standard aqueous buffers at pH 2, pH 5, pH 7, pH 8.5 and pH 12 containing 1% of non-ionic surfactant (Arkopal N 150). The tests are done in accelerated conditions at 40°C for approximately one month (28d). 200 – 300 ppm of test substance are dissolved in the pH buffer containing the surfactant and put into storage in an oven at 40°C. Commercial reference grades of buffer are utilised as listed in documented literature sources. Small aliquots of the test solution are extracted with an organic solvent (typically cyclohexane or ethyl acetate) containing a hydrocarbon standard (typically C12, C17 or C20) on a regular basis throughout the test (typically at time = 0, 0.25, 1, 2, 4, 7, 15, 21 and 28 days). The extracts are analysed by GC-FID. Under the analytical conditions employed constituent 1 isomer was eluted and the results were then plotted with time to show the degradation curves of the substance. For constituent 1 of the test item at pH 5 there is evidence of hydrolytic instability of the test item. The test item rapidly degrades to 90% degradation within 10 days and by day 15 there is < 1% test item remaining. The t1/2 appears to be 48 hours. At pH 7 and pH 8.5 the test item appears under the conditions of the test, relatively stable up to 28 days with less than 10% degradation. It can be concluded that under the conditions of the present test the test substance is not hydrolytically stable as defined in the OECD TG 111 for hydrolysis as a function of pH at acidic pH ranges. Applicant assessment of the data indicates unexpected stability at pH 7 which may be influenced by the presence of constituent 3 preventing degradation within the test system.

Water Solubility:

Flask Method, OECD TG 105, 2016: The water solubility of the substance was determined using the slow stirring flask method according to OECD TG 105 and EU Method A.6 under GLP. The test item water solubility was determined to be between two loading rates. Complete disappearance of chromatograph peak 2 occurred at both loading rates within the period of the test (24 to 72 hours) under the conditions of the study. Therefore the water solubility of the test item at 20°C is estimated on the basis of chromatograph peak 1. It was estimated to be 16.4 mg/L at and initial loading rate 30 mg/L and 63 mg/L at an initial loading rate of 300 mg/L. The pH of the aqueous samples was 7.8 - 8.8. Applicant assessment indicates there was clear hydrolytic degradation at (pH 7 to pH 8) during the conduct of the study. With potentially faster degradation at lower concentration of test item. The degradation rate occurred within a period of 24 to 72 hours. Within the lower loading rate all water solubility concentrations were based upon chromatograph peak 1. Which was observed to remain during the study. Within the higher loading rate the water solubility was based upon chromatograph peak 1 and 2. The latter chromatograph peak 2 completely disappeared during the period of the test (< 72 hours.

 

Conclusion: There is clear evidence of test item hydrolytic instability of specific constituents of the test item within the pH range 5 to 8.5 within the two separate studies and separate conditions. This was also evidenced in available ecotoxiciology studies in Daphnia sp. and Algae inhibition tests that utilised WAF loading rates of up to 100 mg/L. For example, detected actual concentrations of 6.1 mg/L (WAF: 10 mg/L) and 59 mg/L (WAF: 100 mg/L) in Daphnia medium (analytically maintained within 80 to 110% nominal) under static conditions. This corresponded to one specific test item component (oxydipropanol). No ecotoxicological effects were observed within the WAF loading ranges of concentration and between 48 and 72 hours, respectively.