4-(1,4-dioxa-spiro[4.5]dec-8-yl)-cyclohexanone

Hydrolysis

Link to relevant study record(s)

Reference

Endpoint:

hydrolysis

Type of information:

experimental study

Adequacy of study:

key study

Study period:

1997-12-18 till 1998-01-16

Reliability:

1 (reliable without restriction)

Rationale for reliability incl. deficiencies:

guideline study

Qualifier:

according to guideline

Guideline:

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

Version / remarks:

December 1992

Deviations:

no

GLP compliance:

yes

Radiolabelling:

no

Analytical monitoring:

yes

Details on sampling:

Pretest: The concentration of the test substance was determined immediately after preparation (t=0), after 2.4 h and after 5 days.
Test 1: The concentration of the test substance was determined immediately after preparation (t=0) and 1, 2, 3, 3.5, 4, 4.5, 5, 5.5 and 6 hours after t=0.
Test 3: The concentration of the test substance was determined immediately after preparation (t=0), after 16 and 19 h after t=0.

Buffers:

- pH: 4
sterile 0.05 M acetat buffer (sodium acetate/ acetatic acid/ Milli-Q water)

- pH: 7
sterile 0.05 M phosphate buffer (potassium dihydrogenphosphate/ sodium hydroxide/ Milli-Q water)

- pH: 9
sterile 0.05 M borate buffer (boric acid/ potassium chloride/ sodium hydroxide/ Milli-Q water)

Details on test conditions:

Preparation of the test solutions
Prior to the performance of each test, the test solutions were freshly prepared. Therefore, an accurately weighed amount of approximately 23-32 mg test substance was added to 50.0 mL buffer solution pH 4, pH 7 or pH 9. After sonication for approximately 15 minutes, the resultant solutions were filter-sterilized through a 0.2 µm membrane filter (FP 030/3, Schleicher & Schuell, 's-Hertogenbosch, The Netherlands) and transferred into sterile glass vessels. To exclude oxygen, nitrogen gas was bubbled through each solution for approximately 5 minutes. Thereafter, each vessel was tightly sealed with a septum crimp cap.

Preliminary test
After preparation, the test solutions at pH 4, pH 7 and pH 9 were placed in a thermostatically controlled water bath at 50.0 ± 0.6°C in the dark. The concentration of the test substance was determined immediately after preparation (t=0), after 2.4 hours and after 5 days.

Testing of pseudo-first order behaviour (Test 1 according to EEC guideline C.7)
During the preliminary test, significant hydrolysis (i.e. a decrease in test substance concentration at 50°C < 50% after 2.4 hours but > 10% after 5 days) was observed at pH 4. Therefore, Test 1 was carried out at this pH value. After preparation, the test solution at pH 4 was placed in a thermostatically controlled water bath at 40.7 ± 0 1°C in the dark. The concentration of the test substance at pH 4 was determined immediately after preparation (t=0) and 1, 2, 3, 3.5, 4, 4.5, 5, 5.5 and 6 hours after t=0. For each test solution, the logarithms of the relative concentrations between 20% and 70% hydrolysis were plotted against time. When this is a straight line, the reaction is considered to be (pseudo)-first order and Test 3 according to EEC guideline C.7 must be performed. The results from Test 1 for pH 4 were also used for the calculation of k(25°C) in Test 3. For that purpose, the logarithms of all relative concentrations were plotted against time.

Determination of the half life time (t1/2) at 25°C (Test 3 according to EEC guideline C.7)
Because the resultant line from Test 1 was a straight line for pH 4 Test 3 was carried out. This test was performed at 30.5°C. After preparation, the test solution was placed in a thermostatically controlled water bath at the specified temperature, and in the dark. For pH 4 and 30.5°C, the concentration of the test substance was determined immediately after preparation (t=0) and 16 and 19 hours after t=0. The logarithms of all relative concentrations were plotted against time for these pH values. The results from Test 1 for pH 4 at 40.7°C were also used.

pH measurements
For each test solution the pH value at room temperature was determined at the beginning and at the end of each test.

Duration:

5 d

pH:

4

Temp.:

50 °C

Initial conc. measured:

0.359 g/L

Duration:

5 d

pH:

7

Temp.:

50 °C

Initial conc. measured:

0.479 g/L

Duration:

5 d

pH:

9

Temp.:

50 °C

Initial conc. measured:

0.365 g/L

Number of replicates:

Preliminary test, Test 1 and Test 3: duplicate analysis

Positive controls:

not specified

Negative controls:

not specified

Preliminary study:

At pH 7 and pH 9, a decrease in concentration < 10% was observed after 5 days (half-life time at 25°C > 1 year). Therefore, no further testing was necessary at pH 7 and pH 9. At pH 4, a decrease in concentration < 50% after 2.4 hours but > 10% after 5 days was observed. In order to determine if hydrolysis of the test item is a pseudo-first order reaction, a subsequent test was performed at pH 4.

Test performance:

Test 1:
For the test solution pH 4, the plot of the logarithms of the relative concentrations between 20% and 70% hydrolysis against time is a straight line. Therefore, the reaction at pH 4 is considered to be (pseudo) first order. Hence, the half-life time at 25 °C was determined at this pH value in a subsequent test. The results of Test 1 are also used.
Test 3:
From each regression line, the rate constants and the corresponding half-life time at 25 °C were calculated.

Transformation products:

not specified

% Recovery:

0

pH:

4

Temp.:

50 °C

Duration:

5 d

% Recovery:

104

pH:

7

Temp.:

50 °C

Duration:

5 d

% Recovery:

109

pH:

9

Temp.:

50 °C

Duration:

5 d

Key result

pH:

4

Temp.:

25 °C

DT50:

35 h

Key result

pH:

7

Temp.:

25 °C

DT50:

> 1 yr

Key result

pH:

9

Temp.:

25 °C

DT50:

> 1 yr

Details on results:

Please see Table 1 and 2 below for details on Test 1 and Test 3.

Table 1: Results of Test 1 at 40.7 °C ± 0.1°C

code pH	Measured pH value	Time (hours)	Concentration test substance [g/L]*	Relative concentration [%]	logarithm rel. conc.
4	4.0	0	0.512	100	2.00
	4.0	1	0.467	91.3	1.96
	4.0	2	0.409	79.9	1.90^1
	4.0	3	0.368	71.9	1.86^1
	4.0	3.5	0.352	68.7	1.84^1
	4.0	4	0.323	63.1	1.80^1
	4.0	4.5	0.318	62.1	1.79^1
	4.0	5	0.283	55.3	1.74^1
	4.0	5.5	0.27	52.8	1.72^1
	4.0	6	0.254	49.6	1.70^1

*Mean value of duplicate analysis. The maximum deviation between the responses was < 10%.

¹Value between 20% and 70% hydrolysis

 

Table 2: Results of Test 3 at 30.5 ± 0.1°C 

Code pH	Measured pH value	Time [hours]	Concentration test substance^*[g/L]	Relative concentration [%]	logarithm rel. conc.
4	4.0	0	0.411	100	2.00
	4.0	16	0.232	56.6	1.75
	4.0	19	0.198	48.1	1.68

 *Mean value of duplicate analysis. The maximum deviation between the responses was < 10%.

 

Validity criteria fulfilled:

yes

Conclusions:

The test item is hydrolytically stable (t1/2 > 1 year) at pH 7 and 9. At pH 4, the half-life time (t1/2) is 35 h at 25 °C.

Executive summary:

The determination of the hydrolysis as a function of pH was based on the EEC-Directive 92/69 EEC, Part C, Methods for the determination of Ecotoxicity, C.7: “Abiotic degradation: Hydrolysis as a function of pH”. The test item was determined to be hydrolytically stable (half-life time at 25 °C > 1 year) at pH 7 and pH 9 under the conditions of the test. At pH 4, a decrease in concentration < 50 % after 2.4 hours but > 10% after 5 days was observed. In order to determine if hydrolysis of the test item is a pseudo-first order reaction, a subsequent test was performed. The plot of the logarithms of the relative concentrations between 20 % and 70 % hydrolysis against time was a straight line. Therefore, the reaction at pH4 is considered to be (pseudo)-first order. Hence the half-life time at 25 °C was determined in a subsequent test. At pH 4, the half-life times were 18 h (30.5°C) and 5.9 h (40.7°C). From these values the half-life time at pH 4 and 25 °C was calculated to be 35 h.

Description of key information

The test item is hydrolytically stable (t1/2 > 1 year) at pH 7 and 9. At pH 4, the half-life time (t1/2) is 35 h at 25 °C (reference 5.1.2 -1).

Additional information

The determination of the hydrolysis as a function of pH was based on the EEC-Directive 92/69 EEC, Part C, Methods for the determination of Ecotoxicity, C.7: “Abiotic degradation: Hydrolysis as a function of pH”. The test item was determined to be hydrolytically stable (half-life time at 25 °C > 1 year) at pH 7 and pH 9 under the conditions of the test. At pH 4, a decrease in concentration < 50 % after 2.4 hours but > 10% after 5 days was observed. In order to determine if hydrolysis of the test item is a pseudo-first order reaction, a subsequent test was performed. The plot of the logarithms of the relative concentrations between 20 % and 70 % hydrolysis against time was a straight line. Therefore, the reaction at pH4 is considered to be (pseudo)-first order. Hence the half-life time at 25 °C was determined in a subsequent test. At pH 4, the half-life times were 18 h (30.5°C) and 5.9 h (40.7°C). From these values the half-life time at pH 4 and 25 °C was calculated to be 35 h (reference 5.1.2 -1).