reaction mass of [(1R*,5S*)-3,3,5-trimethylcyclohexyl] acetate and [(1S*,5S*)-3,3,5-trimethylcyclohexyl] acetate

Hydrolysis

Link to relevant study record(s)

Reference

Endpoint:

hydrolysis

Type of information:

experimental study

Adequacy of study:

key study

Study period:

22 Jul - 02 Oct 2015

Reliability:

1 (reliable without restriction)

Rationale for reliability incl. deficiencies:

guideline study

Qualifier:

according to guideline

Guideline:

OECD Guideline 111 (Hydrolysis as a Function of pH)

Qualifier:

according to guideline

Guideline:

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

GLP compliance:

yes (incl. QA statement)

Radiolabelling:

no

Analytical monitoring:

yes

Details on sampling:

- Sampling intervals for the parent/transformation products: samples were taken at test start (0 h) and at 9 to 10 spaced points (between 10 and 90% of hydrolysis) until test end, at each test temperature
- Sampling intervals/times for temperature measurements: every hour
- Sampling intervals/times for sterility check: sterility was checked at test end by colony forming units determination with Water Plate Count Agar from additional samples of the advanced test by incubation at 36 ± 1 °C for 48 h and at 22 ± 1 °C for 72 h
- Sample storage conditions before analysis: all test item containing samples were prepared immediately (if possible less than 30 min until start of enrichment, but at least not more than 5% of the total study time)

Buffers:

- pH: 4, 7, 9
- Composition of buffer: according to guideline 111
- Buffer solution pH 4: NaOH and mono potassium citrate were dissolved in double distilled water.
- Buffer solution pH 7: NaOH and KH2PO4 were dissolved in double distilled water.
- Buffer solution pH 9: NaOH, KCI and H3BO3 were dissolved in double distilled water.

Details on test conditions:

TEST SYSTEM
- Type, material and volume of test flasks, other equipment used: sterile headspace flasks, volume: 119 mL, with aluminium tops with PTFE seals, test volume: 50 mL
- Sterilisation method: autoclaving
- Temperatures: 20, 30, 50 °C
- Measures taken to avoid photolytic effects: avoidance of direct sunlight and the use of opaque water baths
- Measures to exclude oxygen: buffers were purged with nitrogen for 5 min and subsequently pH was checked to a precision of 0.1 at the test temperature
- If no traps were used, is the test system closed/open: closed

TEST MEDIUM
- Kind and purity of water: double distilled water
- Identity and concentration of co-solvent: acetonitrile, =< 1% (v/v)

Duration:

30 d

pH:

4

Initial conc. measured:

>= 3.98 - <= 4.29 mg/L

Remarks:

Test was performed at 20, 30 and 50 °C.

Duration:

30 d

pH:

7

Initial conc. measured:

>= 3.98 - <= 4.29 mg/L

Remarks:

Test was performed at 20, 30 and 50 °C.

Duration:

30 d

pH:

9

Initial conc. measured:

>= 3.98 - <= 4.29 mg/L

Remarks:

Test was performed at 20, 30 and 50 °C.

Number of replicates:

Duplicates per pH value and sampling date.

Positive controls:

no

Negative controls:

yes

Remarks:

Buffer solutions (pH value 4, 7 and 9)

Preliminary study:

In the preliminary test, more than 10% of the test item was degraded after 120 h at pH 4, 7 and 9 (24, 11 and 78%, respectively).

Transformation products:

yes

No.:

#1

Details on hydrolysis and appearance of transformation product(s):

- Other: transformation product was quantified for all temperature conditions at pH 9

% Recovery:

90.6

pH:

9

Temp.:

20 °C

Duration:

28.5 h

% Recovery:

85

pH:

9

Temp.:

20 °C

Duration:

71.9 h

% Recovery:

72.5

pH:

9

Temp.:

20 °C

Duration:

723 h

% Recovery:

78.5

pH:

9

Temp.:

30 °C

Duration:

47.7 h

% Recovery:

52.8

pH:

9

Temp.:

30 °C

Duration:

338 h

% Recovery:

38.4

pH:

9

Temp.:

30 °C

Duration:

506 h

% Recovery:

28.1

pH:

9

Temp.:

30 °C

Duration:

723 h

% Recovery:

52.4

pH:

9

Temp.:

50 °C

Duration:

28.6 h

% Recovery:

36.3

pH:

9

Temp.:

50 °C

Duration:

53.4 h

% Recovery:

10.3

pH:

9

Temp.:

50 °C

Duration:

170 h

pH:

4

Temp.:

30 °C

Hydrolysis rate constant:

0 s-1

DT50:

12 132 d

Type:

(pseudo-)first order (= half-life)

Remarks on result:

other: Number of data points: 11, slope of regression graph: not significantly non zero

pH:

4

Temp.:

50 °C

Hydrolysis rate constant:

0 s-1

DT50:

63.9 d

Type:

(pseudo-)first order (= half-life)

Remarks on result:

other: Number of data points: 11, slope of regression graph: significantly non zero

pH:

7

Temp.:

20 °C

Hydrolysis rate constant:

0 s-1

DT50:

415 d

Type:

(pseudo-)first order (= half-life)

Remarks on result:

other: Number of data points: 11, slope of regression graph: not significantly non zero

pH:

7

Temp.:

30 °C

Hydrolysis rate constant:

0 s-1

DT50:

566 d

Type:

(pseudo-)first order (= half-life)

Remarks on result:

other: Number of data points: 11, slope of regression graph: not significantly non zero

pH:

7

Temp.:

50 °C

Hydrolysis rate constant:

0 s-1

DT50:

89 d

Type:

(pseudo-)first order (= half-life)

Remarks on result:

other: Number of data points: 11, slope of regression graph: significantly non zero

pH:

7

Temp.:

25 °C

Hydrolysis rate constant:

0 s-1

DT50:

451 d

Type:

(pseudo-)first order (= half-life)

Remarks on result:

other: values calculated via Arrhenius equation

pH:

9

Temp.:

20 °C

Hydrolysis rate constant:

0 s-1

DT50:

72.9 d

Type:

(pseudo-)first order (= half-life)

Remarks on result:

other: Number of data points: 11, slope of regression graph: significantly non zero

pH:

9

Temp.:

30 °C

Hydrolysis rate constant:

0 s-1

DT50:

17.3 d

Type:

(pseudo-)first order (= half-life)

Remarks on result:

other: Number of data points: 11, slope of regression graph: significantly non zero

pH:

9

Temp.:

50 °C

Hydrolysis rate constant:

0 s-1

DT50:

2.24 d

Type:

(pseudo-)first order (= half-life)

Remarks on result:

other: Number of data points: 10, slope of regression graph: significantly non zero

pH:

9

Temp.:

25 °C

Hydrolysis rate constant:

0 s-1

DT50:

29 d

Type:

(pseudo-)first order (= half-life)

Remarks on result:

other: values calculated via Arrhenius equation

Details on results:

TEST CONDITIONS
- pH, sterility, temperature, and other experimental conditions maintained throughout the study: no (for more information see any other information on results)

MAJOR TRANSFORMATION PRODUCTS

At pH 9 and 20 °C:
- Maximum concentrations in % of the parent substance and day(s) of incubation when observed: 100% at 0 h
- Maximum concentration in % of the parent substance at end of study period: 72.5% at 723 h

At pH 9 and 30 °C:
- Maximum concentrations in % of the parent substance and day(s) of incubation when observed: 100% at 0 h
- Maximum concentration in % of the parent substance at end of study period: 28.1% at 723 h

At pH 9 and 50 °C:
- Maximum concentrations in % of the parent substance and day(s) of incubation when observed: 100% at 0 h
- Maximum concentration in % of the parent substance at end of study period: 10.3% at 170 h

The test item showed a moderate hydrolysis rate (t1/2 =< 30 d) at pH 9 at 30 and 50 °C.

At pH 4 and 7, 50 °C and pH 9, 20 °C only a slow hydrolysis (t1/2 > 30 d) was observed.

At pH 4 and 7, 20 and 30 °C the half-lives were > 1 year indicating no significant hydrolysis of the test item.

Table 1: pH values in the test system measured before start of hydrolysis.

Test	Intended pH-value	Measured pH-value at 20 °C	Measure pH-value at 30 °C	Measured pH-value at 50 °C
Advanced	4.0 ± 0.1	4.02/4.03	4.03/4.02	4.03/4.02
	7.0 ±0.1	6.98 / 6.99	6.98/6.99	6.98/7.00
	9.0 ±0.1	9.03 / 9.03	9.03/9.03	9.03/9.02

Table 2: Mean, maximum and minimum values of temperature measured in the test system.

Test	pH value	Intended Temperature	Measured Temperature
			Mean ± SD	Min. / Max.
Preliminary	4, 7, 9	50.0 ± 0.5	50.0 ± 0.07	49.6 / 50.3
Advanced		20.0 ± 0.5	20.0 ±0.1	19.7/20.6*
		30.0 ± 0.5	30.0 ±0.1	29.3/30.4*
		50.0 ±0.5	50.0 ± 0.2	47.6 / 50.5**

* Temperature out of range for 0.1% of the total run time

** Temperature out of range for 1.4% of the total run time

No significant colony forming units were found during CFU-determination, confirming the sterility of the test solutions.

Validity criteria fulfilled:

yes

Conclusions:

The test item showed a moderate hydrolysis rate (t1/2 =< 30 d) at pH 9 at 30 and 50 °C.
At pH 4 and 7, 50 °C and pH 9, 20 °C only a slow hydrolysis (t1/2 > 30 d) was observed.
At pH 4 and 7, 20° and 30 °C the half-lives were > 1 year indicating no significant hydrolysis of the test item.
The transformation product was 3,3,5-Trimethylcyclohexanol which was identified via co-chromatography.

Description of key information

The test substance is hydrolytically stable under environmental conditions at pH 4 and pH 7. For pH 9 a DT50 of 29 d was calculated at 25 °C (Arrhenius equation).

Key value for chemical safety assessment

Half-life for hydrolysis:

1 yr

at the temperature of:

25 °C

Additional information

One experimental study is available investigating the hydrolytic properties of the substance. The study was performed according to OECD 111 (GLP). A preliminary test with a concentration of 5 mg/L was performed at three different pH values (pH 4, 7 and 9) and 50 °C. The test substance was found to be hydrolytically unstable (degradation > 10%) under the conditions of the preliminary test after 120 h. Thus, a main test with a maximum duration of 30 d was performed for pH 4, 7 and 9 at 20, 30 and 50 °C (initial test substance concentration: 3.98 - 4.29 mg/L). The results from GC/MS analysis indicated that the test substance is hydrolytically stable at pH 4 with a DT50 > 1 year at 30 °C and a DT50 of 63.9 d at 50 °C. A comparable result was observed at pH 7 with a DT50 of > 1 year at 20, 30 (regression analysis) and 25 °C (calculated using Arrhenius equation). The fastest hydrolysis was recorded at pH 9 with a DT50 of 72.9 d at 20 °C, 17.3 d at 30 °C and 2.24 d at 50 °C based on regression analysis. Applying the Arrhenius equation a DT50 of 29 d resulted at 25 °C. 3,3,5-trimethylcyclohexanol was identified by co-chromatography as the major transformation product during hydrolysis at pH 9. All validity criteria of the study were met.