17-acetoxy-1β,2β-methanopegna-4,6-diene-3,20-dione

Hydrolysis

Link to relevant study record(s)

Reference

Endpoint:

hydrolysis

Type of information:

experimental study

Adequacy of study:

key study

Study period:

5 Jul - 14 Oct 1994

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:

92/69/EEC

Deviations:

no

GLP compliance:

yes

Radiolabelling:

no

Analytical monitoring:

yes

Remarks:

HPLC UV

Buffers:

- pH: 4
- Type: acetate buffer, 0.05 M
- Composition of buffer:
0.67 g = 0.008 mole sodium acetate p.a Merck, Art. 6268 (M = 82.03 g/mol) were dissolved to 1000 ml with double distilled
water. The pH value was adjusted to 4 by adding 2.4 ml = 0.04 mole acetic acid 100 % p.a. (Merck, Art. 63 E).

- pH: 7
- Type: phosphate buffer, 0.05 M
- Composition of buffer:
4.34 g = 0.031 mole disodium hydrogen phosphate, Merck, Art. 6589 (M = 141.96 g/mol)
2.64 g = 0.019 mole potassium dihydrogen phosphate, Merck, Art. 4881 (M = 136.09 g/mol) were dissolved to 1000 ml with double distilled water.

- pH: 9
- Type: borate buffer, 0.05 M
- Composition of buffer:
4.76 g = 0.012 mole disodium tetraborate, Merck, Art. 6315 (M = 381.37 g/mol)
46 mL = 0.0046 mole 0.1 M hydrochloric acid Titrisol, Merck, Art. 9944 (M = 36.46 g/mol) were dissolved to 1000 ml with double distilled water.

Details on test conditions:

Preparation of the test solution:
- 3 test solutions, buffered to pH values of 4, 7 and 9
- each with a test substance concentration of approx. 1 mg/L am containing 1 % acetonitrile
- The buffer solution was equilibrated at 50 °C for one hour and then purged with nitrogen to remove dissolved oxygen.
- All experiments were performed using sterilized equipment, buffer solutions and double distilled water.

Duration:

7 h

pH:

9

Temp.:

80 °C

Initial conc. measured:

1 mg/L

Duration:

96.3 h

pH:

9

Temp.:

60 °C

Initial conc. measured:

1 mg/L

Remarks:

Solution II

Duration:

96.5 h

pH:

9

Temp.:

60 °C

Initial conc. measured:

1 mg/L

Remarks:

Solution I

Duration:

168 h

pH:

9

Temp.:

50 °C

Initial conc. measured:

1 mg/L

Duration:

168 h

pH:

7

Temp.:

50 °C

Initial conc. measured:

1 mg/L

Duration:

168 h

pH:

4

Temp.:

50 °C

Initial conc. measured:

1 mg/L

Positive controls:

no

Negative controls:

no

Preliminary study:

A preliminary study was performed and the results indicate that the test substance is hydrolytically stable at pH 4 and pH 7 therefore no further studies were performed for these pH values. At pH 9, more than 10 % decay of the test substance was measured within 5 days at 50 °C.

Transformation products:

not measured

% Recovery:

15.4

St. dev.:

1.379

pH:

9

Temp.:

80 °C

Duration:

7 h

Remarks on result:

other: Preliminary test: hydrolytically not stable

% Recovery:

88

St. dev.:

1.099

pH:

9

Temp.:

60 °C

Duration:

96.3 h

Remarks on result:

other: Preliminary test: hydrolytically not stable; Solution II

% Recovery:

8.5

St. dev.:

1.053

pH:

9

Temp.:

60 °C

Duration:

96.5 h

Remarks on result:

other: Preliminary test: hydrolytically not stable; Solution I

% Recovery:

29

St. dev.:

0.041

pH:

9

Temp.:

50 °C

Duration:

168 h

Remarks on result:

other: Preliminary test: hydrolytically not stable

% Recovery:

96

St. dev.:

0.201

pH:

7

Temp.:

50 °C

Duration:

168 h

Remarks on result:

hydrolytically stable based on preliminary test

% Recovery:

98.1

St. dev.:

0.079

pH:

4

Temp.:

50 °C

Duration:

168 h

Remarks on result:

hydrolytically stable based on preliminary test

Key result

pH:

9

Temp.:

25 °C

Hydrolysis rate constant:

0 h-1

DT50:

138 d

Type:

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

Key result

pH:

7

Temp.:

50 °C

Remarks on result:

hydrolytically stable based on preliminary test

Key result

pH:

4

Temp.:

50 °C

Remarks on result:

hydrolytically stable based on preliminary test

For further investigation of the hydrolysis at pH 9, hydrolysis experiments were conducted at temperatures of 60° and 80 °C with replication at 60 °C to extrapolate from the experimentally determined rate constants at the elevated temperatures to the rate constant at 25 °C.

The rate constant of the test substance at pH 9 and 25 °C was calculated by applying the Arrhenius equation. The extrapolated first-order rate constant k and half-life t_1/2 and confidence intervals for k and t_1/2 are shown in the table behind.

pH	k [h^-1]	95% confidence intervals for k [h^-1]	t 1/2 [h]	95% confidence intervals for t 1/2 [h]	t 1/2 [d]	95% confidence intervals for t 1/2 [d]
9	2.095*10^-4	2.029*10^-4 – 2.160*10^-4	3309.2	3209.4 -.3415.5	137.9	133.7 -142.3

Validity criteria for the hydrolysis study:

Target condition according to guideline:	Actual condition according to the study:	Validity criteria met:
The analytical method used will depend on the nature of the substance and must be sufficiently precise and sensitive to detect a reduction of 10 % of the initial concentration.	Analysis via HPLC-UV is senstive enough to detect a reduction of 10% of the initial concentration.	Yes
A sufficient number of samples (not less than four) should be chosen to cover the range 20 to 70 % of hydrolysis to test for pseudo-first order behaviour at the specified pH values.	pH 4 (50°C): 2 measurements pH 7 (50°C): 2 measurements pH 9 (50°C): 2 measurements pH 9 (60°C): 2 measurements pH 9 (80°C): 1 measurement	Yes
The method must be specific to allow determination of the test substance at the test solution concentrations and may well consist of some combination of suitable analytical techniques.	The test substance was determined by HPLC-UV detection at 290 nm. Chromatograms of the test and calibration solutions demonstrate the specificity of the method.	Yes

 

Validity criteria fulfilled:

yes

Remarks:

See 'Any other information on results incl. tables'

Conclusions:

The rate of hydrolysis of the test substance was determined at pH 4, 7 and 9. The test substance is hydrolytically stable at pH values 4 and 7 at 25 °C. The half-life at pH 9, 25°C was extrapolated from measurements at elevated temperatures using the Arrhenius equation and the experimental data to be 137.9 days.

Executive summary:

The study was conducted to assess the rate of hdrolysis. The study was conducted in accordance with EEC Method for "determination of the hydrolysis", Directive 92/69/EEC, method A.7. The rate of hydrolysis of the test substance was determined at pH 4, 7 and 9. The test substance is hydrolytically stable at pH values 4 and 7 at 25 °C. The half-life at pH 9 (25°C) was extrapolated from measurements at elevated temperatures using the Arrhenius equation to be 137.9 days.

Description of key information

The study was conducted to assess the rate of hydrolysis. The study was conducted in accordance with EEC Method for "determination of the hydrolysis", Directive 92/69/EEC, method A.7. The rate of hydrolysis of the test substance was determined at pH 4, 7 and 9. The test substance is hydrolytically stable at pH values 4 and 7 at 25 °C. The half-life at pH 9 (25 °C) was extrapolated from measurements at elevated temperatures using the Arrhenius equation to be 137.9 days.

Key value for chemical safety assessment

Half-life for hydrolysis:

138 d

at the temperature of:

25 °C