2-[(2S,3S)-3-[[(1,1dimethylethoxy)carbonyl]amino]-2-hydroxy-4-phenylbutyl]-2-[[4-(2pyridinyl)phenyl]methyl]-,1,1-dimethylethyl ester

Hydrolysis

Link to relevant study record(s)

Reference

Endpoint:

hydrolysis

Type of information:

experimental study

Adequacy of study:

key study

Study period:

from 06 Jun 2017 to 28 Aug 2017

Reliability:

1 (reliable without restriction)

Rationale for reliability incl. deficiencies:

guideline study

Reason / purpose for cross-reference:

reference to other study

Qualifier:

according to guideline

Guideline:

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

Version / remarks:

2008

Deviations:

no

Qualifier:

according to guideline

Guideline:

OECD Guideline 111 (Hydrolysis as a Function of pH)

Version / remarks:

2004

Deviations:

no

Qualifier:

according to guideline

Guideline:

EPA OPPTS 835.2120 (Hydrolysis of Parent and Degradates as a Function of pH at 25°C)

Version / remarks:

2008

Deviations:

no

GLP compliance:

yes

Radiolabelling:

no

Analytical monitoring:

yes

Details on sampling:

TESTS CONDUCTED
Tier 1 at 50°C (pH 4, 7 and 9) and Tier 2 at 3 different temperatures (pH 4, 7 and 9)

PREPARATION OF THE SAMPLES
- Test item was spiked to the buffers using a solution of the test item in acetonitrile.
- Target concentration: 0.2 mg/L
- For each sampling time, duplicate sterile vessels under vacuum were filled with 6 mL test solution.
- The vessels were placed in a temperature controlled environment in the dark.
- Blank buffers containing a similar content of co-solvent were treated similarly as the test samples (at t=0).

SAMPLING DETAILS
- Samples for analysis were taken immediately after preparation (t=0), at t= 5 days (Tier 1) and at several sampling points after t=0 (Tier 2).
- Samples taken from solutions with a temperature > 20°C were cooled to room temperature using running tap water.
- Samples not analysed on the sampling day were stored in the freezer. Stability has been demonstrated over a period of at least 17 days. On the day of analysis, the frozen samples were defrosted at room temperature, treated and analysed.


pH MEASUREMENTS
The pH of the test solutions (except for the blanks) was determined at each sampling point.

Buffers:

Buffer pH 4: 16.7% 0.01 M sodium acetate and 83.3% 0.01 M acetic acid in water.

Buffer pH 7: 0.01 M potassium di-hydrogenphosphate in water adjusted to pH 7 using 1 N sodium hydroxide.

Buffer pH 9: 0.01 M boric acid and 0.01 M potassium chloride in water adjusted to pH 9 using 1 N sodium hydroxide.

Details:
- Kind of water: tap water purified by a purification system (Milli-Q, Millipore)
- Sodium azide content: 0.0009% (w/v)

Details on test conditions:

TEST SYSTEM
- Sterilisation method: Each buffer was filter-sterilised through a 0.2 µm FP 30/0.2 CA-S filter (Whatman) and transferred into a sterile vessel.
- Measures to exclude oxygen: nitrogen gas was purged through the sterile buffers for 5 minutes prior to preparation of the blank and test solutions.
- Measures taken to avoid photolytic effects: the test vessels were kept in the dark.

TEST MEDIUM
- Identity and concentration of co-solvent: acetonitrile. The volume of the co-solvent was < 1% of the sample volume. Nominal concentrations were not corrected for the spiking volume.

DETAILS TIER 1
pH 4.0, pH 7.0 and pH 9.0 at 50.0 ± 0.2°C

DETAILS TIER 2
pH 4.0 - 4.1 at 19.3 ± 0.8°C, 49.8 ± 0.1°C and 59.0 ± 0.6°C
pH 6.9 - 7.1 at 19.3 ± 0.8°C, 49.6 ± 0.4°C and 59.1 ± 0.6°C
pH 8.9- 9.0 at 19.3 ± 0.8°C, 49.6 ± 0.3°C and 59.1 ± 0.6°C

Duration:

769.62 h

pH:

4

Temp.:

20 °C

Initial conc. measured:

>= 0.208 - <= 0.225 mg/L

Remarks:

Duplicate samples.To avoid an error in the Validation check, the' >=' and '<=' codes were used.

Duration:

266.08 h

pH:

4

Temp.:

50 °C

Initial conc. measured:

>= 0.161 - <= 0.203 mg/L

Duration:

75.77 h

pH:

4

Temp.:

60 °C

Initial conc. measured:

>= 0.167 - <= 0.176 mg/L

Duration:

769.4 h

pH:

7

Temp.:

20 °C

Initial conc. measured:

>= 0.196 - <= 0.201 mg/L

Duration:

745.93 h

pH:

7

Temp.:

50 °C

Initial conc. measured:

>= 0.187 - <= 0.187 mg/L

Duration:

98.07 h

pH:

7

Temp.:

60 °C

Initial conc. measured:

>= 0.171 - <= 0.187 mg/L

Duration:

768.98 h

pH:

9

Temp.:

20 °C

Initial conc. measured:

>= 0.191 - <= 0.207 mg/L

Duration:

576.85 h

pH:

9

Temp.:

50 °C

Initial conc. measured:

>= 0.18 - <= 0.194 mg/L

Duration:

75.02 h

pH:

9

Temp.:

60 °C

Initial conc. measured:

>= 0.175 - <= 0.186 mg/L

Number of replicates:

Two

Positive controls:

no

Negative controls:

no

Preliminary study:

At pH 4, pH 7 and pH 9, a degree of hydrolysis of > 10% was observed after 5 days at 50°C. The Tier 2 test is required.

Test performance:

RECOVERIES
- Recovery is the concentration analysed at t=0 relative to the nominal concentration.
- The mean (n=2) recovery at each pH and temperature was calculated (see table below).
- The mean recoveries obtained at pH 4 fell within the acceptable range for non-labelled chemicals of 70-110%.
- The mean recoveries obtained at pH 7 fand at pH 9 ell within the criterion range of 90-110%.
- The concentrations analysed in the test samples were not corrected for recovery.

Transformation products:

not specified

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

no data

% Recovery:

85

pH:

4

Temp.:

50 °C

Duration:

0 h

Remarks on result:

other: Tier 1

% Recovery:

78

pH:

7

Temp.:

50 °C

Duration:

0 h

Remarks on result:

other: Tier 1

% Recovery:

86

pH:

9

Temp.:

50 °C

Duration:

0 h

Remarks on result:

other: Tier 1

% Recovery:

> 100

pH:

4

Temp.:

20 °C

Duration:

0 h

Remarks on result:

other: Tier 2 (108%)

% Recovery:

91

pH:

4

Temp.:

50 °C

Duration:

0 h

Remarks on result:

other: Tier 2

% Recovery:

86

pH:

4

Temp.:

60 °C

Duration:

0 h

Remarks on result:

other: Tier 2

% Recovery:

99

pH:

7

Temp.:

20 °C

Duration:

0 h

Remarks on result:

other: Tier 2

% Recovery:

93

pH:

7

Temp.:

50 °C

Duration:

0 h

Remarks on result:

other: Tier 2

% Recovery:

90

pH:

7

Temp.:

60 °C

Duration:

0 d

Remarks on result:

other: Tier 2

% Recovery:

99

pH:

9

Temp.:

20 °C

Duration:

0 h

Remarks on result:

other: Tier 2

% Recovery:

94

pH:

9

Temp.:

50 °C

Duration:

0 h

Remarks on result:

other: Tier 2

% Recovery:

90

pH:

9

Temp.:

60 °C

Duration:

0 h

Remarks on result:

other: Tier 2

pH:

4

Temp.:

20 °C

Hydrolysis rate constant:

0 h-1

DT50:

1 601 h

Type:

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

Key result

pH:

4

Temp.:

25 °C

Hydrolysis rate constant:

0.001 h-1

DT50:

902 h

Type:

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

Remarks on result:

other: from the Arrhenius equation

pH:

4

Temp.:

50 °C

Hydrolysis rate constant:

0.01 h-1

DT50:

66 h

Type:

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

pH:

4

Temp.:

60 °C

Hydrolysis rate constant:

0.039 h-1

DT50:

18 h

Type:

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

pH:

7

Temp.:

20 °C

Hydrolysis rate constant:

0.002 h-1

DT50:

382 h

Type:

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

Key result

pH:

7

Temp.:

25 °C

Hydrolysis rate constant:

0.002 h-1

DT50:

313 h

Type:

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

Remarks on result:

other: from the Arrhenius equation

pH:

7

Temp.:

50 °C

Hydrolysis rate constant:

0.006 h-1

DT50:

124 h

Type:

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

pH:

7

Temp.:

60 °C

Hydrolysis rate constant:

0.028 h-1

DT50:

25 h

Type:

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

pH:

9

Temp.:

20 °C

Hydrolysis rate constant:

0.002 h-1

DT50:

365 h

Type:

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

Key result

pH:

9

Temp.:

25 °C

Hydrolysis rate constant:

0.002 h-1

DT50:

308 h

Type:

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

Remarks on result:

other: from the Arrhenius equation

pH:

9

Temp.:

50 °C

Hydrolysis rate constant:

0.005 h-1

DT50:

140 h

Type:

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

pH:

9

Temp.:

60 °C

Hydrolysis rate constant:

0.033 h-1

DT50:

21 h

Type:

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

pH 4: Statistical Parameters Pseudo-first order curve

Temperature (°C)	Slope	Intercept	Coefficient of correlation
20	-1.88E-04	2.00	0.852
50	-4.53E-03	1.98	0.996
60	-1.69E-02	1.91	0.986

pH 7: Statistical Parameters Pseudo-first order curve

Temperature (°C)	Slope	Intercept	Coefficient of correlation
20	-7.88E-04	1.90	0.828
50	-2.44E-03	1.91	0.936
60	-1.20E-02	1.82	0.967

pH 9: Statistical Parameters Pseudo-first order curve

Temperature (°C)	Slope	Intercept	Coefficient of correlation
20	-8.24E-04	1.99	0.946
50	-2.14E-03	1.86	0.985
60	-1.42E-02	1.89	0.983

Validity criteria fulfilled:

yes

Conclusions:

The half-life time of the substance at 25°C at pH 4, pH 7 and pH 9 was determined to be 902 hours, 313 hours and 308 hours, respectively.

Executive summary:

The rate of hydrolysis of the substance at pH values normally found in the environment (pH 4, 7 and 9) was determined in a GLP-compliant study according to EC C.7, OECD 111 and EPA OPPTS 835.2120.

A degree of hydrolysis of > 10% was determined at pH 4, pH 7 and pH 9 after 5 days and 50°C (Tier 1).

Hydrolysis rates at 20°C, 50°C and 60°C were determined in a Tier 2 study. The substance was shown to degrade by a pseudo-first order behaviour. Half-life times at 25°C were determined from the Arrhenius equation: 902 hours, 313 hours and 308 hours at pH 4, pH 7 and pH 9, resp.

Description of key information

The rate of hydrolysis of the substance at pH values normally found in the environment (pH 4, 7 and 9) was determined in a GLP-compliant study according to EC C.7, OECD 111 and EPA OPPTS 835.2120.

A degree of hydrolysis of > 10% was determined at pH 4, pH 7 and pH 9 after 5 days and 50°C (Tier 1).

Hydrolysis rates at 20°C, 50°C and 60°C were determined in a Tier 2 study. The substance was shown to degrade by a pseudo-first order behaviour. Half-life times at 25°C were determined from the Arrhenius equation: 902 hours, 313 hours and 308 hours at pH 4, pH 7 and pH 9, resp.

Key value for chemical safety assessment

Half-life for hydrolysis:

313 h

at the temperature of:

25 °C