N-[[3,5-dichloro-2-fluoro-4-(1,1,2,3,3,3-hexafluoropropoxy)phenyl]carbamoyl]-2,6-difluorobenzamide

Hydrolysis

Link to relevant study record(s)

Reference

Endpoint:

hydrolysis

Type of information:

experimental study

Adequacy of study:

key study

Study period:

25 June 1999 to 03 November 1999

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)

Deviations:

no

Qualifier:

according to guideline

Guideline:

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

Deviations:

no

Qualifier:

according to guideline

Guideline:

EPA Guideline Subdivision N 161-1 (Hydrolysis)

Deviations:

no

GLP compliance:

yes

Radiolabelling:

yes

Analytical monitoring:

yes

Details on sampling:

- Sampling intervals for the parent/transformation products: Duplicate samples were taken at 0, 1, 3, 6, 13, 21 and 33 days after treatment for the 25 °C samples and on day 7 for the 50 °C samples; all aliquots were collected using a silanised glass syringe.
- Sampling intervals/times for pH measurements: 1-2 mL were sampled at 0, 1, 3, 6, 13, 21 and 33 days after treatment for the 25 °C samples and on day 7 for the 50 °C samples; all aliquots were collected using a silanised glass syringe.
- Sampling intervals/times for sterility check: Sterility checks were performed at study initiation and each sampling time: 0, 1, 3, 6, 13, 21 and 33 days after treatment for the 25 °C samples and on day 7 for the 50 °C samples; all aliquots were collected using a silanised glass syringe. 500 µL of the test solution or buffer was added to 10 mL of soy broth solution. The solutions were incubated for 3 days at 25 °C. Positive and negative controls were performed for each sterility check.

Buffers:

- pH: 5, 7 and 9
- Composition of buffer: Buffers were prepared using water purified by a Millipore purification unit. The pH 5 buffer was prepared by adjusting 0.01 M sodium acetate trihydrate with glacial acetic acid. The pH 7 buffer was prepared by adjusting 0.01 M 4-(2-hydroxyethyl)-1-piperazineethanesulfonic acid with 2.0 N NaOH. The pH 9 buffer was prepared by adjusting 0.01 M sodium borate with concentrated HCl. All buffers were sterilised by autoclaving.

Details on test conditions:

TEST SYSTEM
- Type, material and volume of test flasks, other equipment used: Silanised glass vials
- Sterilisation method: Autoclaving
- Is there any indication of the test material adsorbing to the walls of the test apparatus? Silanised glass vials were used to reduce sorption to the containers. The test vessels were selected based on preliminary suitability tests.

TEST MEDIUM
- Volume used/treatment 40 mL
- Kind and purity of water: Millipore purified water
- Preparation of test medium: 15 µL of a 5 ng/mL dosing solution containing the test material was added to 40 mL of buffer solution in individual silanised glass vials. The application rate was 2 ng/mL.

Duration:

33 d

pH:

5

Temp.:

25 °C

Initial conc. measured:

ca. 1.896 - 1.898 µg/L

Duration:

33 d

pH:

7

Temp.:

25 °C

Initial conc. measured:

ca. 1.874 µg/L

Duration:

33 d

pH:

9

Temp.:

25 °C

Initial conc. measured:

ca. 1.97 - 1.984 µg/L

Number of replicates:

2

Positive controls:

no

Remarks:

Controls were only included for buffer sterility checks

Negative controls:

no

Remarks:

Controls were only included for buffer sterility checks

Statistical methods:

Statistical methods included calculation of means, standard deviations, r², and slope. Data were entered into a Microsoft Excel spreadsheet for all calculations, including linear regression analysis of degradation kinetics. Within an Excel spreadsheet, intermediate values were not rounded prior to final calculation.
Plotting a regression analysis of In Parent concentration versus time gave a slope, which is equal to - k'b (the observed decay rate constant (days^-1)). For the pH 9 samples this regression analysis gives a k’b value of 0.036 days^-1. This was used to calculate half-life and DT90 value using the following:
t1/2 = 0.693 / 0.036 days^-1
DT90 = ln (10 %) / 0.036 days^-1

The second-order rate constant was calculated at pH 9 using the following:
The concentration of OH- is 10E-05 M, therefore: kb = k’b / [OH-] = 0.036 days^-1 / 10E-05 M

The second-order rate constant was used to predict the pseudo first-order rate constant (k'b) at various pH values using the following:
k'b = k’b[OH-]

Transformation products:

yes

No.:

#1

No.:

#2

No.:

#3

No.:

#4

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

- Pathways for transformation: At pH 5 and 7, the test material was found to be stable to hydrolysis (slightly positive slope). At pH 9, degradation was assumed to be due to base-catalysed hydrolysis.

% Recovery:

93.7

pH:

5

Temp.:

50 °C

Duration:

7 d

% Recovery:

78.6

pH:

7

Temp.:

50 °C

Duration:

7 d

% Recovery:

8.5

pH:

9

Temp.:

50 °C

Duration:

7 d

% Recovery:

100

pH:

5

Temp.:

25 °C

Duration:

33 d

% Recovery:

94.8 - 96.2

pH:

7

Temp.:

25 °C

Duration:

33 d

% Recovery:

28.9 - 30.5

pH:

9

Temp.:

25 °C

Duration:

33 d

pH:

5

Temp.:

25 °C

Hydrolysis rate constant:

-0.002 d-1

Remarks on result:

other: r² = 0.590. Half-life was not calculated due to no degradation being observed

Key result

pH:

7

Temp.:

25 °C

Hydrolysis rate constant:

-0.001 d-1

Remarks on result:

other: r² = 0.095. Half-life was not calculated due to no degradation being observed

pH:

9

Temp.:

25 °C

Hydrolysis rate constant:

0.036 d-1

DT50:

19 d

Type:

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

Remarks on result:

other: r² = 0.989, DT90 = 64

pH:

11

Temp.:

25 °C

Hydrolysis rate constant:

3.6 d-1

DT50:

0.19 d

Type:

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

Remarks on result:

other: Calculated using a second-order rate constant of 3600 days^-1 M^-1

pH:

7

Temp.:

25 °C

Hydrolysis rate constant:

0 d-1

DT50:

1 930 d

Type:

second order

Remarks on result:

other: Calculated using a second-order rate constant of 3600 days^-1 M^-1

Details on results:

TEST CONDITIONS
- pH, sterility, temperature, and other experimental conditions maintained throughout the study: Yes

PATHWAYS OF HYDROLYSIS
- Description of pathways: Degradation at pH 9 was assumed to be due to base-catalysed hydrolysis. As hydroxide ions were in large excess, it was assumed to be a constant; this forced the degradation into first order form. The degradation of the test material was therefore through pseudo-first-order. From the values calculated for pH 9 at 25 °C, a second-order rate constant was derived which could be used to calculate the half-lives at different pH values.
- Figures of chemical structures attached: Yes

Table 1: Hydrolysis of the test material after 7 days at 50 °C

pH	Replicate	Percent of Applied
		Parent		Urea metabolite	Amine metabolite
		Actual	Average
5	A	91.8	93.7	1.2	3.4
	B	95.6		0.9	1.3
7	A	77.6	78.6	15.1	5.6
	B	79.6		12.9	6.2
9	A	8.9	8.5	42.7	18.4
	B	8.1		59.7	23.2

 

Table 2: Hydrolysis of the test material at 25 °C

Sample Time (days)	Replicate	Percent of Applied
		Parent			Urea metabolite at pH 9	Amine metabolite at pH 9
		pH 5	pH 7	pH 9
0	A	94.9	-	98.5	0.7	0.0
	B	94.8	93.7	99.2	0.0	0.0
1	A	-	92.9	89.9	3.0	1.9
	B	95.0	99.3	87.3	3.6	1.5
3	A	96.1	97.2	84.9	8.2	3.6
	B	91.2	85.8	82.5	4.5	4.1
6	A	94.9	-	77.2	17.1	2.7
	B	91.6	92.3	75.3	18.1	3.1
13	A	99.4	100	-	-	-
	B	96.5	97.2	-	-	-
21	A	98.0	98.5	45.3	35.5	13.6
	B	96.8	99.1	39.4	37.5	10.0
33	A	100	96.2	28.9	40.9	8.3
	B	100	94.8	30.5	50.7	9.1

 - Not calculated due to low recovery

Validity criteria fulfilled:

yes

Remarks:

Recovery of the test material was acceptable; where recovery was too low at individual endpoints, these were excluded from analyses.

Conclusions:

Under the conditions of the test, the test material was stable to hydrolysis at pH 5 and 7. At pH 9, the test material degraded through base-catalysed hydrolysis following pseudo-first-order kinetics to form four metabolites (urea, amine, amide and difluorobenzoic acid forms of the parent material). At pH 9, the half-life was 19 days with a DT90 of 64 days.

Executive summary:

The hydrolysis of the test material was investigated in a study conducted in accordance with the standardised guidelines OECD 111, EU Method C.7 and US EPA 161-1 under GLP conditions.

The rate and route of hydrolysis was studied at pH 5, 7, and 9 using [14C]radiolabelled material. The pseudo first-order rate constant was determined and used to calculate the half-life and DT90 of hydrolysis at pH 9. The second-order rate constant at pH 9 was calculated and used to predict degradation rates at various pH values, and degradates approaching or exceeding 10 % of applied radiocarbon were identified.

Samples (2 ng/mL) were incubated at 25 ± 1 °C and sampling times were 0, 1,3,6, 13,21, and 33 days after treatment. Additional samples were incubated at 50 °C and collected at 1 day after treatment (pH 9 only) and 7 days after treatment (pH 5, 7, and 9). All samples were analysed by reverse phase HPLC.

The samples were sterile throughout the study and the pH for all samples was constant. Average material balance for kinetics samples ranged from 101.5 % at pH 9 to 103.3 % at pH 7.

The test material was degraded via base-catalysed hydrolysis. Following 7 days of incubation at 50 °C, it was stable to hydrolysis at pH 5 (93.7 % remaining), slightly hydrolysed at pH 7 (78.6 % remaining) and mostly hydrolysed at pH 9 (8.5 % remaining). At 25°C the test material was stable to hydrolysis at pH 5 and 7. Therefore, rate constants and observed half-life values were not calculated at these pH values. At pH 9, the pseudo-first order rate constant of the test material hydrolysis was 0.036 days^-1 giving half-life and DT90 values of 19 days and 64 days, respectively. The second-order rate constant of hydrolysis was 3600 days^-1 M^-1, giving predicted half-life values of 0.19 days at pH 11 to 1930 days at pH 7. At pH 9, the test material was hydrolysed to form urea, amine, amide, and difluorobenzoic acid forms of the parent material. The study met all required validity criteria.

Description of key information

Hydrolysis = pH 9, 19 days at 25 °C, OECD 111, EU Method C.7, US EPA 161-1, Cook (1999)

Key value for chemical safety assessment

Half-life for hydrolysis:

19 d

at the temperature of:

25 °C

Additional information

In the key study (Cook 1999), the hydrolysis of the test material was investigated in a study conducted in accordance with the standardised guidelines OECD 111, EU Method C.7 and US EPA 161-1 under GLP conditions. The study was assigned a reliability score of 1 in accordance with the principles for assessing data quality as defined in Klimisch et al. (1997).

The rate and route of hydrolysis was studied at pH 5, 7, and 9 using [14C]radiolabelled material. The pseudo first-order rate constant was determined and used to calculate the half-life and DT90 of hydrolysis at pH 9. The second-order rate constant at pH 9 was calculated and used to predict degradation rates at various pH values, and degradates approaching or exceeding 10 % of applied radiocarbon were identified.

Samples (2 ng/mL) were incubated at 25 ± 1 °C and sampling times were 0, 1,3,6, 13,21, and 33 days after treatment. Additional samples were incubated at 50 °C and collected at 1 day after treatment (pH 9 only) and 7 days after treatment (pH 5, 7, and 9). All samples were analysed by reverse phase HPLC.

The samples were sterile throughout the study and the pH for all samples was constant. Average material balance for kinetics samples ranged from 101.5 % at pH 9 to 103.3 % at pH 7.

The test material was degraded via base-catalysed hydrolysis. Following 7 days of incubation at 50 °C, it was stable to hydrolysis at pH 5 (93.7 % remaining), slightly hydrolysed at pH 7 (78.6 % remaining) and mostly hydrolysed at pH 9 (8.5 % remaining). At 25°C the test material was stable to hydrolysis at pH 5 and 7. Therefore, rate constants and observed half-life values were not calculated at these pH values. At pH 9, the pseudo-first order rate constant of the test material hydrolysis was 0.036 days^-1 giving half-life and DT90 values of 19 days and 64 days, respectively. The second-order rate constant of hydrolysis was 3600 days^-1 M^-1, giving predicted half-life values of 0.19 days at pH 11 to 1930 days at pH 7. At pH 9, the test material was hydrolysed to form urea, amine, amide, and difluorobenzoic acid forms of the parent material. The study met all required validity criteria.