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Hydrolysis

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Reference
Endpoint:
hydrolysis
Type of information:
experimental study
Adequacy of study:
key study
Reliability:
2 (reliable with restrictions)
Rationale for reliability incl. deficiencies:
other: According to OECD 111 however only preliminary test is performed
Qualifier:
according to
Guideline:
OECD Guideline 111 (Hydrolysis as a Function of pH)
Version / remarks:
Version 13 April 2004, Preliminary test (TIER 1)
Deviations:
yes
Remarks:
Temperature ranged from 49.30 to 50.67°C with an average of 49.79°C and not at 50°C +/- 0.5°C. It was not considered to have adverse effect to the study.
GLP compliance:
yes
Specific details on test material used for the study:
Product Name: A017 (Naphthol)
Chemical Name: 1-Naphthol
CAS No. : 90-15-3
Descrition : Light cream flakes
Purity: 99.1% ± 0.1%
Lot Number: 7215604684
Date of analysis : 09 September 2016
Method of analysis : HPLC-UV (purity) and LC/MS (identification)
Storage : room temperature

Analytical monitoring:
yes
Details on sampling:
- The Day 0 samples were prepared for HPLC analysis by adding 900 µL of the test solution with 100 µL of acetonitrile. Buffers containing no test material were prepared in the same way as the samples to check for background contamination or buffer interference.
- After five days of incubation at 50C, the Day 5 samples were removed from incubation and prepared for analysis by HPLC.
- Quality Control samples were prepared for HPLC analysis in the same way as the Day 0 samples.

Buffers:
- pH 4 Buffer: Each liter of 0.05 M pH 4 buffer solution was prepared by adding 4 mL of 0.1 M NaOH (aq) and 500 mL of 0.1 M potassium hydrogen phthalate (aq) to a total volume of 1 L with reagent water. A 0.01 M pH 4 buffer solution was prepared by adding 200 mL of the 0.05 M pH 4 buffer to 800 mL of reagent water. The final pH of the solution was 4.02.
- pH 7 Buffer: Each liter of 0.05 M pH 7 buffer solution was prepared by adding 296 mL of 0.1 M NaOH (aq) and 500 mL of 0.1 M KH2PO4 (aq) to a total volume of 1 L with reagent water. A 0.01 M pH 7 buffer solution was prepared by adding 200 mL of the 0.05 M pH 7 buffer to 800 mL of reagent water. The final pH of the solution was 7.07.
- pH 9 Buffer: Each liter of 0.05 M pH 9 buffer solution was prepared by adding 213 mL of 0.1 M NaOH (aq) and 500 mL of 0.1 M H3BO3 (aq) in 0.1 M KCl to a total volume of 1 L with reagent water. A 0.01 M pH 9 buffer solution was prepared by adding 200 mL of the 0.05 M pH 9 buffer to 800 mL of reagent water. The final pH of the solution was 9.05.
All buffer solutions used for testing were sterilized prior to use by filtering through a 0.45µm or smaller filter, and were autoclaved (~121 °C at 15 psi for ~30 minutes). The buffer solutions were stored at room temperature.

Estimation method (if used):
Concentrations of the test substance, A017, were determined using the external standard analysis function of Empower 3 software. The concentrations of A017 in the samples prepared for HPLC-UV analysis were determined directly from the standard curve (Equation below) :
[Concentration from standard curve (mg/mL)] x [analysis volume (mL)] / [sample volume (mL)] = mg/mL
The standard curve equation is of the form: y = mx + b
Where
y = A017 peak area units
m = slope of the standard curve [X Coefficients(s)]
x = mg a.i./mL
b = y-intercept (Constant)

Details on test conditions:
TEST SYSTEM
- Type, material and volume of test flasks, other equipment used: After preparation (see paragraph "Test Medium" below) the test solutions were distributed into 2 vials.
- Sterilisation method: the buffer solutions were filtered through a 0.45µm or smaller filter, and autoclaved (~121 °C at 15 psi for ~30 minutes)
- Measures taken to avoid photolytic effects: Samples wrapped in foil
- Measures to exclude oxygen: vials filled with test solutions without headspace
- If no traps were used, is the test system closed/open : vials sealed with PTFE lined caps
- Is there any indication of the test material adsorbing to the walls of the test apparatus? no information
- The 5 day samples were placed in an oven at 50°C

TEST MEDIUM
- Volume used/treatment : 25 ml dosing solutions prepared in volumetric flask and then distributed into 2 vials placed in an oven set to 50C
- Kind and purity of water: Reagent water used in this study was demineralized and purified using a Millipore Milli-Q Purification Water System.
- Preparation of test medium:
1) For analytical standards : Stock solutions were prepared at approximately 2 mg/mL by weighing approximately 20 mg of the test substance into a 10-mL volumetric flask, correcting for purity (99.1%), and bringing the flask to volume with 90:10 water/acetonitrile (v:v). Dilutions of these solutions, prepared in 90:10 water/acetonitrile (v:v), were used as the analytical standards.
2) For dosing solutions at pH 4 and 7 : A 99.2-mg/mL test substance stock solution was prepared by weighing 100.08 mg of the test substance into a 1-mL volumetric flask, correcting for purity (99.1%), and bringing the flask to volume with acetonitrile. To obtain a test solution at approximately 0.05 mg/mL, 12.6 µL of this stock solution was added into 25-mL volumetric flasks, and bringing to volume with the buffer pH4 and the buffer pH7 respectively. The flasks were mixed by inversion and vortexing and then distributed into the testing vials..
3) For dosing solutions at pH 9 : A 25.1-mg/mL test substance stock solution was prepared by weighing 25.28 mg of the test substance into a 1-mL volumetric flask, correcting for purity (99.1%), and bringing the flask to volume with acetonitrile. To obtain a test solution at approximately 0.025 mg/mL, 25 µL of this stock solution was added into 25-mL volumetric flasks, and bringing to volume with the buffer pH 9. The flasks were mixed by inversion and vortexing and then distributed into the testing vials
4) Day 0 and Day 5, Quality Control (QC) samples were prepared in pH 4 and 7 buffer at approximately 0.05 mg/mL by weighing approximately 0.0025 g of the test substance into a 50mL volumetric flask, correcting for purity (99.1%), and bringing the flask to volume with the respective buffer. Day 0 and Day 5 Quality Control (QC) samples were prepared in pH 9 buffer at approximately 0.025 mg/mL by weighing approximately 0.0025 g of the test substance into a 100-mL volumetric flask, correcting for purity (99.1%), and bringing the flask to volume with the respective buffer.



Duration:
5 d
pH:
4
Temp.:
50 °C
Initial conc. measured:
0.048 mg/L
Remarks:
Mean concentration on two vials
Duration:
5 d
pH:
7
Temp.:
50 °C
Initial conc. measured:
0.049 mg/L
Remarks:
Mean concentration on two vials
Duration:
5 d
pH:
9
Temp.:
50 °C
Initial conc. measured:
0.026 mg/L
Remarks:
Mean concentration on two vials
Number of replicates:
2 replicates
Positive controls:
no
Negative controls:
no
Statistical methods:
Calculations, including linear regression, were performed using Microsoft Excel (version 2010). Values in the Excel spreadsheets were not rounded during the calculations (i.e. calculations performed using Excel in full precision mode)
Preliminary study:
The stability of the test item was evaluated via the recovery rate. The recovery rate was calculated by comparing the values of "day 0" with the values of "day 5".
Transformation products:
not measured
% Recovery:
91.15
St. dev.:
0.8
pH:
4
Temp.:
50 °C
Duration:
5 d
Remarks on result:
hydrolytically stable based on preliminary test
% Recovery:
70.3
St. dev.:
0.6
pH:
7
Temp.:
50 °C
Duration:
5 d
Remarks on result:
other: not stable in the test conditions
% Recovery:
18.1
St. dev.:
0.4
pH:
9
Temp.:
50 °C
Duration:
5 d
Remarks on result:
other: not stable in the test conditions
Key result
pH:
4
Temp.:
50 °C
DT50:
37.2 d
Type:
other: value based on only 2 data points
Remarks on result:
hydrolytically stable based on preliminary test
Key result
pH:
7
Temp.:
50 °C
DT50:
9.74 d
Type:
other: value based on only 2 data points
Key result
pH:
9
Temp.:
50 °C
DT50:
2.02 d
Type:
other: value based on only 2 data points
Details on results:
TEST CONDITIONS
- Temperature : The temperature ranged from 49.3 to 50.67°C with an average from 49.79°C

Hydrolysis of A017, pH 4

Sample

Concentration of Sample (mg/mL)

Percent of Initial

C0/Ct

ln (C0/Ct)

Replicate 1

0.0474

na

na

na

Replicate 2

0.0494

na

na

na

Mean Initial Concentration

0.0484

100%

1

0

Replicate 1

0.0444

91.7%

1.09

0.0869

Replicate 2

0.0439

90.6%

1.10

0.0993

 

Hydrolysis of A017, pH 7

Sample

Concentration of Sample (mg/mL

Percent of Initial

C0/Ct

ln (C0/Ct)

Replicate 1

0.0497

na

na

na

Replicate 2

0.0491

na

na

na

Mean Initial Concentration

0.0494

100%

1

0

Replicate 1

0.0345

69.9%

1.43

0.359

Replicate 2

0.0349

70.7%

1.42

0.347

 

Hydrolysis of A017, pH 9

Sample

Concentration of Sample (mg/mL

Percent of Initial

C0/Ct

ln (C0/Ct)

Replicate 1

0.0259

na

na

na

Replicate 2

0.0258

na

na

na

Mean Initial Concentration

0.0258

100

1

0

Replicate 1

0.00477

18.4%

5.42

1.69

Replicate 2

0.00461

17.8%

5.6

1.72

 

 

Validity criteria fulfilled:
yes
Conclusions:
Following the preliminary test OECD 111, the test item is considered to be stable in sterile aqueous solution buffered at pH 4 and unstable at pH 7 and 9, after incubation for five days at 49.8°C.
After five days at pH 4, the test substance was >90% of the initial concentration and had a half-life of 37.2 days. The test substance had degraded by Day 5 at pH 7 with approximately 70% of the initial concentration, and at pH 9 with approximately 18% of the initial concentration, and had a half-life of 9.74 and 2.02 days, respectively.
Executive summary:

The test item was tested for hydrolysis according to the OECD Guideline 111 and GLP. The preliminary test was performed at 50°C and pH 4, 7 and 9.

Since less than 10% of hydrolysis was observed after 5 days at pH 4 and 50°C, the test item is considered stable at pH 4 and no additionnal test is required.

The test substance had degraded at pH 7 with approximately 70% of the initial concentration after 5 days at 50°C, and at pH 9 the concentration measured after 5 days at 50°C is approximately 18% of the initial concentration. At ph 7 and 9 the test item is not stable and had a half-life of 9.74 and 2.02 days, respectively.

Description of key information

Following the preliminary test OECD 111, the test item is considered to be stable in sterile aqueous solution buffered at pH 4 and unstable at pH 7 and 9, after incubation for five days at 49.8°C.

After five days at pH 4, the test substance was >90% of the initial concentration and had a half-life of 37.2 days.  The test substance had degraded by Day 5 at pH 7 with approximately 70% of the initial concentration, and at pH 9 with approximately 18% of the initial concentration, and had a half-life of 9.74 and 2.02 days, respectively.

Key value for chemical safety assessment

Half-life for hydrolysis:
9.74 d
at the temperature of:
50 °C

Additional information

The test item was tested for hydrolysis according to the OECD Guideline 111 and GLP. The preliminary test was performed at 50°C and pH 4, 7 and 9.

Since less than 10% of hydrolysis was observed after 5 days at pH 4 and 50°C, the test item is considered stable at pH 4 and no additionnal test is required.

The test substance had degraded at pH 7 with approximately 70% of the initial concentration after 5 days at 50°C, and at pH 9 the concentration measured after 5 days at 50°C is approximately 18% of the initial concentration. At ph 7 and 9 the test item is not stable and had a half-life of 9.74 and 2.02 days, respectively.