Document DD5mBdbznV7KbJG97ZDgBxGaO
LZ
JOLI
TRADE SECRET
Study Title Hydrolytic Stability of H-25435 as a Function of pH
D u P on t-14965
Test Guidelines: OECD Guideline 111
Author: Bogdan Szostek, Ph.D.
Study Completed on: October 15,2004
Performing Laboratory: E.I. du Pont de Nemours and Company HaskellSMLaboratory for Health and Environmental Sciences Elkton Road, P.O. Box 50 Newark, Delaware 19714-0050
Laboratory Project ID : DuPont-14965
Sponsor: E.I. du Pont de Nemours and Company Wilmington, Delaware 19898 U.S.A.
Work Request Number: 15028
Service Code Number: 392
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GOOD LABORATORY PRACTICE COMPLIANCE STATEMENT
This study was conducted in compliance with U.S. EPA TSCA (40 CFR part 792) Good Laboratory Practice Standards, which are compatible with the OECD Principles o f Good Laboratory Practice (as revised 1997), ENV/MC/CHEM(98)17, OECD, Paris, 1998, and MAFF Japan Good Laboratory Practice Standards (11 NohSan Number 6283), except for the item documented below. The item listed does not impact the validity of the study.
The test substance was characterized by the sponsor prior to the initiation of this study. Although the characterization was not performed under Good Laboratory Practice Standards, the characterization was done by an ISO 9000 certified laboratory, and the accuracy o f the data is considered sufficient for the purposes of this study.
Applicant / Sponsor: E.I. du Pont de Nemours and Company Wilmington, Delaware 19898 U.S.A.
Study Director:
y9
Bogdan Szostek, Ph.D.
Senior Research Chemist
1 9 - O C T '3 lqqI\ Date
Applicant / Sponsor:_________________________________________ __________
DuPont Representative
Date
2- -
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Hydrolytic Stability o f H -25435 as a Function o f pH
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QUALITY ASSURANCE STATEMENT
Work Request Number: 15028
Study Code Number:
392
The conduct of this study has been subjected to periodic Quality Assurance inspections. The dates of inspection are indicated below.
Dates Reported to Study Phase Audited______Audit Dates_______________________ Director and Management
Conduct:
06 May 2004
07 May 2004
Report/Records: 09-12 Aug 2004
24 Aug 2004
A T (g fc/^
Date
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CERTIFICATION
We, the undersigned, declare that this report provides an accurate evaluation of data obtained from this study.
Approved by: Issued by Study Director:
Mark Kennedy, Ph,? M anager
Bogdan Szostek, Ph.D. Senior Research Chemist
Date Date
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TABLE OF CONTENTS
Page
GOOD LABORATORY PRACTICE COMPLIANCE STATEMENT.....................................2
QUALITY ASSURANCE STATEMENT........................................................................................ 3
CERTIFICATION.................................................................................................................................4
LIST OF TABLES.................................................................................................................................6
LIST OF FIGURES.............................................................................................................................. 7
STUDY INFORMATION....................................................................................................................8
SUMMARY............................................................................................................................................ 9
INTRODUCTION................................................................................................................................10
STUDY DESIGN..................................................................................................................................10
MATERIALS AND M ETHODS...................................................................................................... 10 A. M aterials....................................................................................................................................10
1. Test Substance.....................................................................................................................................................10 2. Residual 8-2 TBA in H -25435......................................................................................................................... 10 3. Preparation o f buffer solutions......................................................................................................................... 11 4. Equipment........................................................................................................................................................... 12
B. Methods...................................................................................................................................... 12
1. Preliminary test...................................................................................................................................................12 2. pH measurement and temperature....................................................................................................................12 3. Analysis o f test substance................................................................................................................................. 12 4. Analysis o f 8-2 T B A .......................................................................................................................................... 18 5. Calculations.........................................................................................................................................................20
RESULTS AND DISCUSSION........................................................................................................21
CONCLUSIONS................................................................................................................................. 22
RECORDS AND SAMPLE STORAGE.........................................................................................23
REFERENCES.................................................................................................................................... 23
TABLES................................................................................................................................................ 24
FIGURES.............................................................................................................................................. 35
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LIST OF TABLES
Page
1. ANALYTICAL RESULTS FOR AQUEOUS STABILITY OF H-25435, pH 1.2, 37C, DAYS 0 (03-JUNE-2004) AND 5 (08-JUNE-2004).....................................................................................................................25
2. ANALYTICAL RESULTS FOR AQUEOUS STABILITY OF H-25435, pH 4.0, 50C, DAYS 0 (13-MAY-2004) AND 5 (18-MAY-2004)......................................................................................................................26
3. ANALYTICAL RESULTS FOR AQUEOUS STABILITY OF H-25435, pH 7.0, 50C, DAYS 0 (06-MAY-2004) AND 5 (ll-M A Y -2004)......................................................................................................................27
4. ANALYTICAL RESULTS FOR AQUEOUS STABILITY OF H-25435, pH 9.0, 50C, DAYS 0 (13-MAY-2004) AND 5 (18-MAY-2004)......................................................................................................................28
5. MEASURED CONCENTRATIONS OF 8-2 TBA FOR pH 1.2 ,37C, DAYS 0 (03-JUNE-2004) AND 5 (08-JUNE-2004)................................................................................................................................................................. 29
6. MEASURED CONCENTRATIONS OF 8-2 TBA FOR pH 4.0, 50C, DAYS 0 (13-MAY-2004) AND 5 (18-MAY-2004)................................................................................................................................................................. 30
7. MEASURED CONCENTRATIONS OF 8-2 TBA FOR pH 7.0, 50C, DAYS 0 (06-MAY-2004) AND 5 (ll-M A Y -2004)................................................................................................................................................................. 31
8. MEASURED CONCENTRATIONS OF 8-2 TBA FOR pH 9.0, 50C, DAYS 0 (13-MAY-2004) AND 5 (18-MAY-2004)................................................................................................................................................................. 32
9. DAILY TEMPERATURE READINGS OF THE SHAKING WATER BATH....................................................... 33
10. pH MEASUREMENTS FOR DAY 0 AND DAY 5..................................................................................................... 34
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LIST OF FIGURES
Page
1. EXAMPLE CALIBRATION CURVES OBTAINED FOR MONITORED [
]............. 36
2. EXAMPLE CHROMATOGRAMS OF MONITORED [
] OBTAINED FOR A
BLANK SAMPLE FOR pH 7.0 BUFFER ON DAY 0...............................................................................................37
3. EXAMPLE CHROMATOGRAMS OF MONITORED [
] OBTAINED FOR A
CALIBRATION STANDARD CONTAINING 5.3 mg/L H-25435..........................................................................38
4. EXAMPLE CHROMATOGRAMS OF MONITORED [
| OBTAINED FOR
DAY 0, pH 7.0 HYDROLYTIC STABILITY SAMPLE........................................................................................... 39
5. EXAMPLE CHROMATOGRAMS OF MONITORED [
] OBTAINED FOR
DAY 5, pH 7.0 HYDROLYTIC STABILITY SAMPLE........................................................................................... 40
6. REPRESENTATIVE CALIBRATION CURVE FOR 8-2 TBA................................................................................41
7. REPRESENTATIVE CHROMATOGRAMS OF 104 pg/L 8-2 TBA CALIBRATION STANDARD AND 515 pg/L D-8-2 TBA INTERNAL STANDARD............................................................................................. 42
8. REPRESENTATIVE CHROMATOGRAMS FOR DAY 5, pH 7.0 BUFFER BLANK SAMPLE......................43
9. REPRESENTATIVE CHROMATOGRAMS FOR DAY 5, pH 7.0 HYDROLYTIC STABILITY SAMPLE AND 515 pg/L D-8-2 TBA INTERNAL STANDARD............................................................................44
10. REPRESENTATIVE CHROMATOGRAMS FOR DAY 0, pH 7.0 HYDROLYTIC STABILITY SAMPLE AND 515 pg/L D-8-2 TBA INTERNAL STANDARD............................................................................ 45
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STUDY INFORMATION
Substance Tested: [
]
Svnonyms/Codes : [ . H-25435
[
] ]
Haskell Number: 25435
Composition: [ [ [ [ [
]
] ] ]
[] Known Impurities: None identified
Physical Characteristics: Amber brown liquid
Stability: The test substance appeared to be stable under the conditions o f the study; no evidence o f instability was observed.
Study Initiated/Completed: May 4, 2004 / (see report cover page)
In-Life Initiated/Completed: May 6, 2004 / July 6, 2004
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SUMMARY
The hydrolytic stability of H-25435 was tested in buffered aqueous solutions at pH 4.0, 7.0, 9.0
at 50C and pH 1.2 at 37C for five days. [
] representing
major chemical species present in H-25435 were monitored during the hydrolytic stability study.
Measured concentrations and recoveries of individual [
] reported as
average concentrations and recoveries for Day 0 and Day 5 replicate samples, were used to
evaluate the hydrolytic stability of H-25435. The average percent recoveries for the Day 0 and
Day 5, pH 1.2 replicate samples were 76 5.9% and 63 28%, respectively. The average
percent recoveries for the Day 0 and Day 5, pH 4.0 buffer replicate samples were 97 3.0% and
95 4.4%, respectively. The average percent recoveries for the Day 0 and Day 5, pH 7.0
replicate samples were 92 1.1% and 90 0.9%, respectively. The average percent recoveries
for the Day 0 and Day 5, pH 9.0 replicate samples were 96 0.6% and 95 9.3%, respectively.
The H-25435 was demonstrated to be hydrolytically stable at pH 9.0, pH 7.0, and pH 4.0. The
data obtained for hydrolytic stability o f H-25435 at pH 1.2 show the difference o f average
recoveries for monitored [
] between Day 0 and Day 5 samples is larger than
10 percent. However, a precipitate was observed after addition o f the test substance to the
pH 1.2 buffer. The precipitate was difficult to completely dissolve even after addition of
acetonitrile and extended sonication, especially for the Day 5 samples. This lowered the
observed recoveries, especially for the bis esters. Therefore, the observed change in the average
recoveries between the Day 0 and Day 5 samples is most likely because of observed solubility
problems rather than hydrolysis.
The measured concentration of 8-2 TBA did not indicate any hydrolytic degradation of H-25435 to 8-2 TBA for any of the investigated pHs. The observed concentrations of 8-2 TBA can be explained by the residual 8-2 TBA present in the test substance (H-25435). There was no significant increase of 8-2 TBA concentration observed between Day 0 and Day 5 samples for any of the investigated pHs.
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INTRODUCTION
The purpose of the study was to investigate the hydrolytic stability of H-25435 in pH 1.2, 4.0, 7.0, and 9.0 aqueous buffer solutions. The study was conducted according to the protocol: "Hydrolytic stability of H-25435 as a function of pH", which was patterned after the OECD Guideline 111.(,) Additionally, the concentration of the 8-2 Telomer B Alcohol (8-2 TBA) was monitored to investigate if 8-2 TBA is released from the test substance during the test.
STUDY DESIGN
A preliminary test was performed on the test substance at 50 0.1 C at each o f the pH 4.0, 7.0, 9.0 and at 37 0.1 C for pH 1.2. If less than 10% of the test substance hydrolysis is observed after 5 days, the test substance was considered hydrolytically stable and no additional testing was performed. In addition to the test substance determination during the preliminary test, the samples were analyzed for 8-2 TBA.
MATERIALS AND METHODS
A. Materials
1.
Name:
Test Substance
H-25435
Composition:
[
1 [ 1 [
Haskell number:
[
25435
] 1
1 1 ]
2. Residual 8-2 TBA in H-25435
Residual 8-2 TBA content in the test substance (H-25435) was determined by LC/MS using the same analytical method as described in Analysis of 8-2 TBA in Methods. Five replicate samples of H-25435 (approximately 45 mg) were introduced to glass vials, weighed, 10 mL of THF (tetrahydrofurane) was added, vials crimped, and sonicated for 1.5 hours. A 0.1 mL aliquot of dissolved sample was transferred to a glass LC vial and 0.9 mL acetonitrile and internal standard (D-8-2 TBA) was added. Calibration standards were made in 10/90% THF/acetonitrile in the range from 100 to 10200 ng/mL 8-2 TBA.
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3. Preparation of buffer solutions
The following buffer solutions were prepared as described in the Annex to OECD guideline 1 1 1 .(1)
a. pH 1.2:
The 0.2 M potassium chloride (KC1) was prepared by dissolving 3.73 g o f KC1 (purity 99.0%, Sigma) in 250 mL of water. The 0.2 M hydrochloric acid (HC1) was prepared by diluting 4.5 mL of hydrochloric acid (36.5-38%, Sigma) in 250 mL water. The pH 1.2 buffer solution was prepared by placing 125 mL o f 0.2 M KC1 and [ ] mL of a 0.2 M HC1 solution into a 500-mL flask and bringing the flask to volume with water. The pH o f the buffer was adjusted to 1.2 with sodium hydroxide.
b. pH 4:
The 0.1 M solution of potassium hydrogen phthalate was prepared by dissolving 5.1 g of potassium hydrogen phthalate (purity 99+%, Sigma) in 250 mL o f water. The pH 4.0 buffer solution was prepared by placing 2 mL of a 0.1 M sodium hydroxide solution and 250 mL of a 0.1 M potassium hydrogen phthalate solution into a 500-mL flask and bringing the flask to volume with water. The pH of the buffer was adjusted to 4.0 with hydrochloric acid.
c. pH 7:
The 0.1 M solution of potassium phosphate was prepared by dissolving 3.4 g of potassium phosphate, monobasic (KH2PO4, purity 99.5%, Sigma) in 250 mL o f water. The pH 7 buffer solution was prepared by placing 148 mL of a 0.1 M sodium hydroxide solution and 250 mL of the 0.1 M potassium phosphate solution into a 500-mL flask and bringing the flask to volume with water. The pH o f the buffer was adjusted to 7.0 with hydrochloric acid.
d. pH 9:
The 0.1 M boric acid (H3BO3) solution in 0.1 M potassium chloride (KC1) was prepared by dissolving 1.90 g of KC1 (purity 99.0%, Sigma) and 1.55 g o f H3BO3 (purity 99.5%, Sigma) in 250 mL of water. The pH 9.0 buffer solution was prepared by placing 250 mL o f a 0.1 M H3BO3 solution in 0.1 M KC1 and 106.5 mL o f a 0.1 M sodium hydroxide solution into a 500-mL flask
and bringing the flask to volume with water. The pH of the buffer was adjusted to 9.0 with
hydrochloric acid.
Water used for buffer preparation was obtained from a Bamstead NANOpure Diamond water purification system. The water resistivity was >18.2 megaohm*cm. All buffer solutions were sterilized prior to use by filtering through Coming Sterilization Filter systems with 0.22-pm cellulose acetate filter. The buffer solutions were stored at room temperature.
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4. Equipment
pH Meter: Water Bath: Autoclave: Automatic Pipettes: Balance: Centrifuge: Vortexer: Glass Vials:
Orion Model 250 A Precision Reciprocal Shaking Water Bath, Model 50, Precision AMSCO Model 3023 Research Pro 100,1000, 5000; Eppendorff Mettler AE-100 Sorval Legend RT Multi-tube vortexer, VWR 10 mL borosilicate Serum Type Reaction vials (Supelco) with 20 mm Barrier Septa (Foil/Silicone), 0.1" thick (Supelco). The vials and septa were autoclaved at 121C for 30 min before use.
B. Methods
1. Preliminary test
The preliminary test was performed on H-25435 at 50C at each of the pH 4.0, 7.0, 9.0 and at 37C for pH 1.2. The same sample preparation procedure and study design was followed for each of the pHs tested. Approximately 45 mg of the H-25435 was weighed into sixteen 10-mL autoclaved glass vials and 3 mL o f appropriate buffer solution was added. The vials were crimp capped with the aluminum foil covering the part o f the septa facing the inside o f the vial. Eight of the vials were processed for analysis as the Day 0 samples. The remaining eight samples were wrapped with the aluminum foil, submerged in the water bath, and shaken at 100 rpm in the water bath for five days at the appropriate temperature. Eight vials containing only 3 mL of appropriate buffer were prepared to serve as the blanks. Four o f them were used as the Day 0 blanks and the other four were wrapped in aluminum foil and placed in the water bath to be the Day 5 blanks. Additionally, six vials containing 3 mL o f appropriate buffer were spiked with 8-2 TBA stock solution (4.5 pL of 1040 mg/L 8-2 TBA). The vials were crimp capped with the aluminum foil covering the part o f the septa facing the inside o f the vial. Three of the vials were used as the QC samples for 8-2 TBA analysis on Day 0. The other three vials were wrapped in ahiminum foil and placed in the water bath together with eight of the test substance vials. These served a role of QC samples for 8-2 TBA analysis on Day 5. Four of the Day 0 or four of the Day 5 vials and two blank vials were processed for the H-25435 determination. The other four of the Day 0 or Day 5 vials, two blanks vials, and 8-2 TBA QC vials were processed for the 8-2 TBA determination.
2. pH measurement and temperature
The pH was measured on Day 0 and Day 5 in the vials containing the buffer and the test substance, designated for the H-25435 analysis. The vials were uncapped, the pH electrode was introduced to the vials and the pH reading was taken. These vials were further processed for H-25435 analysis. The temperature o f the water bath was monitored daily. The water bath has an automatic temperature control built in. The readings o f the temperature control were checked and recorded daily.
3. Analysis o f test substance
H-25435 is a mixture of [ present in H-25435 [
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] The [
]
][
Hydrolytic Stability o f H -25435 as a Function o f pH
[
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]
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[
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]
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The test substance H-25435 has not been characterized to determine the actual contents of the chemical species that were monitored during the study. The concentrations o f individual chemical species that were monitored during the study were represented as the total concentration of the H-25435.
Vials designated for H-25435 analysis were uncapped, 6 mL of acetonitrile was added, vials were recapped and vortexed for 30 minutes. The obtained extract was diluted 1:1000 with 2:1 acetonitrile:water before analysis. The blank samples were subjected to the same procedure, except the pH measurement.
The stock solution o f H-25435 was made by dissolving 0.01 g of H-25435 in 10 mL of 2 mL isopropanol: 8 mL water. Calibration standards for the H-25435 analysis were made by appropriate dilution of the stock. The calibration standards ranged from 1 to 10 mg/L of H-25435. Fresh calibration standards were prepared for each instance of analysis. Linear or quadratic regression of the peak area vs. standard concentration was used to establish the calibration curves.
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Instrumentation and conditions:
HPLC Instrument: MS Instrument:
LC Parameters: Column: Mobile Phase:
Model Agilent 1100, Agilent Quattro Micro, Waters
XterraMS C 18,30 m m x 2.1 mm, 2.5 pm
A- 0.15% (v/v) acetic acid/10 mM triethylamine in water
B- 0.15% (v/v) acetic acid/10 mM triethylamine in methanol
Gradient:
Time (min)
%B
Flow (mL/min)
0.0 50 0.25
2.0 100 0.25
8.0 100 0.25
8.1 100 0.50
10.1 100 0.50
10.2 50
0.25
15.0 50
0.25
Column Temperature: Injection Volume:
30C 10 pL
MS Parameters: Ionization mode: Capillary voltage: Cone voltage: Source Temperature: Desolvation Temperature: Data Acquisition Function:
Electrospray, negative ions 3.2 kV 20 V 120C
450C C6-P mono ester ion: SIR o f 443 m/z; 0-12 min C8-P mono ester ion: SIR o f 543 m/z; 0-12 min C10-P mono ester ion: SIR o f 643 m/z; 0-12 min C6-P-IPA mixed ester ion: SIR o f 485 m/z; 0-12 min C8-P-IPA mixed ester ion: SIR o f 585 m/z; 0-12 min C10-P-IPA mixed ester ion: SIR o f 685 m/z; 0-12 min C6-P-C6 bis ester ion: SIR o f 789 m/z; 0-12 min C6-P-C8 bis ester ion: SIR o f 889 m/z; 0-12 min C8-P-C8/C6-P-C10 bis ester ion: SIR o f 989 m/z; 0-12 min C10-P-C10 bis ester ion: SIR o f 1089 m/z; 0-12 min
4. Analysis of 8-2 TBA
Samples designated for 8-2 TBA analysis were processed by injecting 3 mL methyl-tertbutyl ether (MTBE) through the vial septum using a glass syringe. The pH 1.2 samples required neutralization before the extraction. Neutralization was done by injecting 0.27 mL o f IN sodium hydroxide to the sample vial before the MTBE addition. Neither Day 0 nor Day 5 samples were uncapped before MTBE injection to avoid any losses o f 8-2 TBA. The vials containing injected MTBE were vortexed for 20 or 30 minutes on a multitube vortexer, centrifuged for 20 minutes at 4150 rpm at room temperature. Occasionally, the MTBE and aqueous layers would not separate even after centrifugation. In these cases, samples were be placed in a freezer at -20PC for overnight storage, removed from the freezer, allowed to come to room temperature and centrifuged for 20 minutes. Subsequently, the sample vials were uncapped and 0.1 mL o f the
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MTBE extract was transferred to a glass LC vial, 0.9 mL acetonitrile added, the extract was spiked with internal standard, crimped capped, and subjected to LC/MS analysis for 8-2 TBA.
The 1H, 1H, 2H, 2H-perfluorodecan-l-ol (97.6%, Oakwood Products, West Columbia, SC) was used as the analytical standard of 8-2 TBA. The ID, ID, 2D, 2D, 3-13C-heptadecafluoro decanol (M+5) (abbreviated as D-8-2 TBA) was used as the internal standard. Stock solutions (1000 mg/L) of the analytical standard and the internal standard were prepared in methanol and were stored refrigerated. The calibration standards were prepared freshly for each calibration in 10/90% MTBE/acetonitrile. The calibration standards were made in the range from 100 to 10,000 pg/L 8-2 TBA. A constant level of internal standard was used: 515 pg/L D-8-2 TBA. The calibration curves were constructed using the ratio of the peak area for ions m/z monitored for 8-2 TBA and peak area for ions monitored for D-8-2 TBA and the concentrations of 8-2 TBA.
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Hydrolytic Stability o f H-25435 as a Function o f pH
Instrumentation and conditions:
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HPLC Instrument: MS Instrument:
Model 2795, Waters ZQ, Waters
LC Parameters: Column: Mobile Phase:
Column Temperature: Injection Volume: Column Switch:
Xterra MS C18, 30 mm x 2.1 mm, 2.5 pm
A - water
B - methanol
Gradient:
Time (min)
%A
0 30
1 30
20
60
6.1 30
8 30
30C
20 pL
2 and 6 min
%B 70 70 100 100 70 70
Flow (mL/min) 0.25 0.25 0.25 0.25 0.25 0.25
MS Parameters: Ionization mode: Capillary voltage: Cone voltage: Source Temperature: Desolvation Temperature: Data Acquisition Function:
Electrospray, negative ions 2.5 kV 10V 120C 300C 8-2 TBA: SIR o f 463, 523; 0-6 min D- 8-2 TBA: SIR o f 467, 528 m/z; 0-6 min
5. Calculations
The following calculations were used to determine the recoveries of individual chemical species monitored for H-25435 (Tables 1-4). For example, if 0.045 g of H-25435 was weighed into the vial and 3 mL of buffer was added to the vial the nominal concentration that was assigned to each of the chemical species was: (0.045 g *1000 mg/g) / 0.003 L = 15,000 mg/L. The nominal concentration used for recovery calculation was an average o f four replicates. The % recovery was calculated by dividing the measured concentration by the nominal concentration, times 100% .
The expected (nominal) concentration of 8-2 TBA in the spiked samples was calculated as follows: the 4.5 pL of 1040 mg/L 8-2 TBA spike represents 4.5 pL x 1040 ng/pL= 4680 ng 8-2 TBA. This amount was extracted with 3 mL o f MTBE. The expected nominal concentration of 8-2 TBA in the MTBE extract is: 4680 ng/3 mL=1560 ng/mL. The % recovery is calculated by dividing the measured concentration by the nominal concentration, times 100%.
The H-25435 contains residual 8-2 TBA. The expected, maximum concentration o f 8-2 TBA derived from residual 8-2 TBA in the MTBE extract (3 mL), assuming complete extraction for a 45 mg sample o f H-25435 is: 45 mg o f H-25435 x (0.129%/100)/0.003 L= 19.4 mg/L (19,400 pg/L).
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RESULTS AND DISCUSSION
Tables 1-4 present the results for measured concentrations and recoveries of individual [
] measured for H-25435 and average concentrations and recoveries for Day 0 and Day 5 replicate
samples for pH 1.2, 4.0, 7.0, 9.0, respectively. The average concentration and recoveries were
calculated as an average of concentrations and recoveries of [
]
monitored for H-25435. The average percent recoveries for the Day 0 and Day 5, pH 1.2
replicate samples were 76 5.9% and 63 28%, respectively. The average percent recoveries
for the Day 0 and Day 5, pH 4.0 buffer replicate samples were 97 3.0% and 95 4.4%,
respectively. The average percent recoveries for the Day 0 and Day 5, pH 7.0 replicate samples
were 92 1.1% and 90 0.9%, respectively. The average percent recoveries for the Day 0 and
Day 5, pH 9.0 replicate samples were 96 0.6% and 95 9.3%, respectively. The difference of
average recoveries between the Day 5 and Day 0 samples was less than 10% for pHs 4.0, pH 7.0,
and pH 9.0 buffers. This indicates that the H-25435 is hydrolytically stable at these pHs
(ti/2 >one year). Similarly, the difference in recoveries [
]
for Day 0 and Day 5 samples at pH 4.0, pH 7.0, and pH 9.0 buffers were less than 10%, except
the C10-P-C10 bis ester at pH 4.0 and pH 9.0 buffer. The average recoveries for Day 0 and
Day 5 samples of the pH 1.2 buffer differ by more than 10%. However, in case of pH 1.2 buffer
a precipitate was observed after addition of the test substance to the buffer. The precipitate was
difficult to completely dissolve even after addition o f acetonitrile and extended sonication,
especially for the Day 5 samples. This lowered the observed recoveries, especially for the bis
esters. Therefore, the observed change in the average recoveries between the Day 0 and Day 5
samples is most likely because o f observed solubility problems rather than hydrolysis.
Figure 1 shows example calibration curves for [
] monitored during the hydrolytic
stability study. Figures 2-5 show example chromatograms obtained for the monitored [
]
for a pH 7.0, Day 0 blank sample, calibration standard containing 5.3 mg/L H-25435, pH 7.0,
Day 0 test sample, and pH 7.0, Day 5 test sample.
The content of residual 8-2 TBA in H-25435 was measured using the procedure described in Section A.2. Based on the results of five replicates of the H-25435, the content of residual 8-2 TBA in H-25435 was calculated to be 0.1290.0048%. Assuming that the buffer extracts all the residual 8-2 TBA from the test substance during the time of hydrolytic stability experiment, the expected concentration o f 8-2 TBA in the test system would be 19.4 mg/L (19,400 pg/L). (see 5 Calculations). Tables 4-8 present the measured concentrations o f 8-2 TBA for Day 0 and Day 5 replicate samples for pH 1.2, 4.0, 7.0, 9.0, respectively. The concentrations o f 8-2 TBA measured for the Day 0 samples are slightly higher than the expected concentration of 8-2 TBA originating from residual 8-2 TBA. This may be due to different extraction methods used for determination o f residual 8-2 TBA in the test substance and the method used for analysis of hydrolytic stability samples. The Day 5 concentrations o f 8-2 TBA are comparable with these observed for Day 0 samples. The observed differences in concentration of 8-2 TBA between the Day 0 and Day 5 samples for pH 7.0, 9.0 buffers are in the range o f error for the analytical method used (15%). A decrease o f 8-2 TBA concentration between Day 0 and Day 5 may be explained by adsorption of 8-2 TBA on the precipitate observed for this pH. The difference of 8-2 TBA concentrations between Day 0 and Day 5 for pH 4.0 samples is larger than can be
-21Company Sanitized. Does Not Contain TSCA CBI.
AR-226
Hydrolytic Stability o f H -25435 as a Function o f pH
D u P on t-14965
explained by the analytical method error (Table 6). However, the observed QC sample recoveries for Day 5 samples indicate higher than expected bias of the analytical method for this set o f samples. Thus, the increase of 8-2 TBA concentration for Day 5 samples is most likely due to analytical method performance rather than the test substance hydrolysis.
Figure 6 shows a representative calibration curve obtained for 8-2 TBA. Figures 7-10 show chromatograms for 8-2 TBA calibration standard; Day 5, pH 7.0 blank sample; Day 5, pH 7.0 test sample; and Day 0, pH 7.0 test sample, respectively.
Table 9 summarizes the water bath temperature recording for pH 1.2, 4.0, 7.0, and 9.0. In all cases, the water bath temperature was maintained at the specified level with deviation not exceeding 0.1 C.
Table 10 summarizes results of the pH measurements for pH 1.2,4.0, 7.0, and 9.0. The measured pH of Day 0 and Day 5 replicate samples for all o f the investigated pHs was at the specified nominal level with the deviation not exceeding 0.1 .
CONCLUSIONS
The hydrolytic stability of H-25435 was tested in buffered aqueous solutions at pH 4.0, 7.0,
9.0 at 50C and pH 1.2 at 37C for five days. The H-25435 was demonstrated to be
hydrolytically stable at pH 9.0, pH 7.0, and pH 4.0 with ti/2 >one year. The data obtained for
hydrolytic stability of H-25435 at pH 1.2 show the difference o f average recoveries for
monitored [
] between Day 0 and Day 5 samples is larger than 10 percent.
However, in case o f pH 1.2 buffer a precipitate was observed after addition o f the test substance
to the buffer. The precipitate was difficult to completely dissolve even after addition of
acetonitrile and extended sonication, especially for the Day 5 samples. This lowered the
observed recoveries, especially for the [
] Therefore, the observed change in the
average recoveries between the Day 0 and Day 5 samples is most likely because of observed
solubility problems rather than hydrolysis.
The measured concentration o f 8-2 TBA did not indicate any hydrolytic degradation o f H-25435 to 8-2 TBA for any of the investigated pHs. The observed concentrations of 8-2 TBA can be explained by the residual 8-2 TBA present in the test substance (H-25435). There was no significant increase of 8-2 TBA concentration observed between Day 0 and Day 5 samples for any of the investigated pHs.
- 22-
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Hydrolytic Stability o f H -25435 as a Function o f pH
D u P on t-14965
RECORDS AND SAMPLE STORAGE
Specimens (if applicable), raw data, and the final report will be retained at Haskell Laboratory, Newark, Delaware, or at Iron Mountain Records Management, Wilmington, Delaware.
REFERENCES
1. Organisation for Economic Cooperation and Development (OECD). May 12,1981. OECD Guidelines for Testing of Chemicals, Hydrolysis as a Function of pH, OECD Guideline No. 111.
-23Company Sanitized. Does Not Contain TSCA CBI.
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Hydrolytic Stability o fH -2 5 4 3 5 as a Function o f pH
D u P on t-14965
TABLES
-24Company Sanitized. Does Not Contain TSCA CB1.
AR-226
Hydrolytic Stability o f H -25435 as a Function o f pH
DuPont-14965
Table 1: Analytical results for aqueous stability of H-25435, pH 1.2,37C, Days 0 (03-June-2004) and 5 (08-June-2004).
11 pH Day Rep (mg/L)
11 (mg/L)
1.2 0 1 1.2 0 2 1.2 0 3 1.2 0 4
17,800 16,400 17,100 18,300
17,200 16,400 17,100 18,200
11 (mg/L)
14,200 13,400 13,900 14,500
I1 (mg/L)
[ I[
1[
1( 1(
!1 !
(mg/L)
(mg/L)
(mg/L)
(mg/L)
(mg/L)
(mg/L)
Average (mg/L)
16,900 15,600 16,800 16,700
16,500 15,000 15,700 16,500
14,200 12,800 13,500 14,400
15,700 12,800 14,700 15,100
8,670 5,810 7,420 7,740
2,970 1,630 2,570 2,620
1,270 648 1,130 1,300
12,500 11,000 12,000 12,500
Average % RSD % Recovery
17,400 4.8 HO
17,200 4.3 109
14,000 3.4 89
16,500 3.7 104
15,900 4.5 101
13,700 5.3 87
14,600 8.6 92
7,410 16.1 47
2,450 23.4 16
1,090 27.7 7.0
12,000 5.9 76
1.2 5 1 14,700
1.2 5 2 1 3,260
1.2 5 3 15,000
1.2 5 4
19,800
14,900 3,300 9,610 19,200
14,200 4,000 6,420 17,000
10,400 1,630 9,910 12,700
10,400 1,530 7,990 12,700
11,000 952 7,160 12,800
10,000 411 4,240 11,300
7,980 640 2,910 8,500
6,890 2,870 4,400 7,160
3,230 768 1,850 3,830
10,400 1,940 6,950 12,500
Average % RSD % Recovery
16,500 17.3 109
14,600 33 97
12,500 44 83
11,000 13.5 73
10,400 22.7 69
10,300 27.9 68
8,510 44 56
6,460 47.8 43
6,150 24.7 41
2,970 34.2 20
9,950 28.2 66
1Replicate excluded from calculations o f Average and %RSD.
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Hydrolytic Stability o f H -25435 as a Function o f pH
D u P on t-14965
Table 2: Analytical results for aqueous stability of H-25435, pH 4 .0 ,50C, Days 0 (13-May-2004) and 5 (18-May-2004).
(1 pH Day Rep (mg/L)
4.0 0 1 15,300 4.0 0 2 16,100 4.0 0 3 16,200 4.0 0 4 16,600
I1 (mg/L)
14,600 15,300 16,700 16,500
11 (mg/L)
[i (mg/L)
14,900 15,700 16,400 15,400
14,900 15,600 16,000 15,800
(1 (mg/L)
( i [C6-P-C6]
(mg/L)
(mg/L)
11[
I1 I
(mg/L)
(mg/L)
(mg/L)
Average (mg/L)
15,200 15,600 15,700 15,800
15,300 15,800 16,300 15,400
15,100 15,500 16,200 15,800
15,200 15,400 16,100 15,500
14,400 15,600 15,900 15,400
10,600 11,300 11,300 10,800
14,600 15,200 15,700 15,300
Average % RSD % Recovery
16,100 3.4 103
15,800 6.3 101
15,600 4.0 100
15,600 3.1 100
15,600 1.7 100
15,700 2.9 100
15,700 3.0 100
15,600 2.5 100
15,300 4.2 98
11,000 3.2 70
15,200 3.0 97
4.0 5 1 15,500
4.0 5 2
15,400
4.0 5 3 14,400
4.0 5 4
16,200
15,300 16,600 15,500 16,700
14,300 15,300 14,400 15,600
15,100 14,900 13,800 15,300
14,700 14,600 13,400 15,100
15,000 15,100 13,600 15,200
15,000 15,000 14,100 15,400
15,000 15,100 14,400 15,900
14,300 14,800 13,800 15,300
8,760 8,800 8,220 9,890
14,300 14,600 13,600 15,100
Average % RSD % Recovery
15,400 4.8 101
16,000 4.5 105
14,900 4,3 98
14,800 4.5 97
14,500 5.1 95
14,700 5.1 97
14,900 3.7 98
15,100 4.1 99
14,600 4.4 96
8,920 7.8 59
14,400 4.4 95
-26Company Sanitized. Does Not Contain TSCA CBI.
AR-226
Hydrolytic Stability o f H -25435 as a Function o f pH
DuPont-14965
Table 3: Analytical results for aqueous stability of H-25435, pH 7 .0 ,50C, Days 0 (06-May-2004) and 5 (ll-M ay-2004).
ri pH Day Rep (mg/L)
ri (mg/L)
7.0 0 1 7.0 0 2 7.0 0 3 7.0 0 4
15,800 15,700 15,300 15,300
15,700 15,900 16,500 16,200
II (mg/L)
15,500 15,300 15,300 14,800
I1 (mg/L)
I1 (mg/L)
[I (mg/L)
15,400 15,000 15,400 15,000
15,400 15,200 15,300 15,100
14,800 15,100 15,300 15,200
[] (mg/L)
15,600 15,500 15,400 15,100
11 1
1I 1
(mg/L)
(mg/L)
(mg/L)
Average (mg/L)
15,800 14,900 14,800 14,300
15,000 14,700 14,300 14,000
9,890 9,720 9740 9570
14,900 14,700 14,700 14,500
Average %RSD % Recovery
15,500 1.7 97
16,100 2.2 101
15,200 2.0 95
15,00 1.5 95
15,300 0.8 96
15,100 1.4 95
15,400 1.4 97
15,000 4.2 94
14,500 3.0 91
9,730 1.3 61
14,700 1.1 92
7.0 5 1 15,000 7.0 5 2 14,400 7.0 5 3 15,300 7.0 5 4 14,700
14,300 14,900 15,500 15,400
13,400 13,600 15,000 14,100
14,500 14,500 14,600 14,600
14,300 14,300 14,300 14,400
14,500 15,100 14,000 14,700
14,600 14,300 14,700 14,300
14,400 14,500 14,600 14,500
13,500 14,000 13,800 13,800
9,330 9,410 9,580 9,660
13,800 13,900 14,100 14,000
Average % RSD % Recovery
14,900 2.6 96
15,000 3.7 96
14,000 5.1 90
14,600 0.4 94
14,300 0.3 92
14,600 3.1 94
14,500 1.4 93
14,500 0.6 93
13,800 1.5 89
9,500 1.6 61
1,4000 0.9 90
-27Company Sanitized. Does Not Contain TSCA CBI.
AR-226
Hydrolytic Stability o f H -25435 as a Function o f pH
DuPont-14965
Table 4: Analytical results for aqueous stability of H-25435, pH 9.0, 50C, Days 0 (13-May-2004) and 5 (18-May-2004).
11 PH Day Rep (mg/L)
11 (mg/L)
9.0 0 1 15,400 9.0 0 2 15,700 9.0 0 3 15,900 9.0 0 4 15,400
15,800 15,800 15,400 15,500
I1 (mg/L)
16,800 14,400 15,300 16,200
I1 (mg/L)
11 (mg/L)
I1 (mg/L)
15,300 15,600 15,300 15,600
15,300 15,600 15,100 15,500
15,300 15,500 15,300 15,400
11 (mg/L)
15,400 15,500 15,200 15,300
I1i
1[ 1
(mg/L)
(mg/L)
(mg/L)
Average (mg/L)
15,200 15,400 15,100 15,200
15,000 15,200 14,600 15,200
12,700 12,500 12,500 12,700
15,200 15,100 15,000 15,200
Average %RSD % Recovery
15,600 1.6 99
15,600 1.3 99
9.0 5 1 15,000
9.0 5 2 14,300
9.0 5 3 15,100
9.0 5 4
14,900
14,500 15,500 15,400 15,800
15,700 6.7 too
12,200 14,800 16,000 15,500
15,500 1.1 98
14,300 14,900 15,200 15,600
15,400 1.4 98
13,100 14,600 15,100 15,500
15,400 0.6 98
10,500 14,900 15,700 15,400
15,400 0.8 98
10,500 15,600 15,400 16,200
15,200 0.8 96
11,700 15,600 16,000 15,800
15,000 1.9 95
12,700 15,500 15,900 16,400
12,600 0.9 80
13,900 14,000 15,200 17,500
15,100 0.6 96
12,800 15,000 15,500 15,900
Average % RSD % Recovery
14,800 2.4 95
15,300 3.7 98
14,600 11.6 94
15,000 3.7 96
14,600 7.2 94
14,100 17.3 91
14,400 18.3 93
14,800 13.9 95
15,100 11.0 97
15,200 11.0 98
14,800 9.3 95
-28Company Sanitized. Does Not Contain TSCA CBI.
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Hydrolytic Stability o f H -25435 as a Function o f pH
D u P on t-14965
Table 5: Measured concentrations of 8-2 TBA for pH 1 .2 ,37C, Days 0 (03-June-2004) and 5 (08-June-2004).
Nominal 8-2 TBA PH Day Rep Concentration (gg/L)
1.2 0 1 1.2 0 2 1.2 0 3 1.2 0 4
-
Average Standard Deviation
1.2 0 QC1 1.2 0 QC2 1.2 0 QC3
1560 1560 1560
1.2 5 1 1.2 5 2 1.2 5 3 1.2 5 4
-
Average Standard Deviation
1.2 5 QC1 1.2 5 QC2 1.2 5 QC3
1560 1560 1560
Measured 8-2 TBA Concentration (pg/L)
25,300 25,700 25,600 24,800
25,400 404
1680 1610 1670
Average Standard Deviation
16,800 24,900 14,800 22,700
19,800 4,780
1470 1500 1450
Average Standard Deviation
% Recovery
-
108 103 107
106 2.7
-
94 96 93
94 1.5
- 29-
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Hydrolytic Stability o f H -25435 as a Function o f pH
D u P on t-14965
Table 6: Measured concentrations of 8-2 TBA for pH 4.0, 50C, Days 0 (13-May-2004) and 5 (18-May-2004).
Nominal 8-2 TBA PH Day Rep Concentration (pg/L)
4.0 0 1 4.0 0 2 4.0 0 3 4.0 0 4
-
Average Standard Deviation
4.0 0 QC1 4.0 0 QC2 4.0 0 QC3
1560 1560 1560
4.0 5 1 4.0 5 2 4.0 5 3 4.0 5 4
-
Average Standard Deviation
4.0 5 QC1 4.0 5 QC2 4.0 5 QC3
1560 1560 1560
Measured 8-2 TBA Concentration (pg/L)
25,400 24,300 21,900 23,200
23,700 1,500
1680 1660 1620
Average Standard Deviation
28,400 29,500 30,200 29,500
29,400 744
2010 2030 1630
Average Standard Deviation
% Recovery
-
108 106 104
106 2.0
-
129 130 104
121 15
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AR-226
Hydrolytic Stability o f H -25435 as a Function o f pH
D u P on t-14965
Table 7: Measured concentrations of 8-2 TBA for pH 7.0, 50C, Days 0 (06-May-2004) and 5 (ll-May-2004).
PH Day
7.0 0 7.0 0 7.0 0 7.0 0
Rep
Nominal 8-2 TBA Concentration
(pr/L)
Measured 8-2 TBA Concentration (pg/L)
1234-
22,300 22,300 22,800 23,900
Average
Standard Deviation
7.0 0 7.0 0 7.0 0
QC1 QC2 QC3
1560 1560 1560
22,800 750
1640 1630 1600
Average Standard Deviation
7.0 5 7.0 5 7.0 5 7.0 5
1 2 3 4
-
24,000 24,300 24,200 12,300 1
Average Standard Deviation
7.0 5 QC1 7.0 5 QC2 7.0 5 QC3
1560 1560 1560
24,200 150
1590 1540 1580
Average Standard Deviation
1Not used in the calculation of the average and standard deviation.
% Recovery
-
105 104 103
104 1.0
-
102 99 101
101 1.5
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AR-226
Hydrolytic Stability o f H -25435 as a Function o f pH
DuPont-14965
Table 8: Measured concentrations of 8-2 TBA for pH 9.0, 50C, Days 0 (13-May-2004) and 5 (18-May-2004).
PH Day
9.0 0 9.0 0 9.0 0 9.0 0
Nominal 8-2 TBA
Rep
Concentration (Pg/L)
1234-
Measured 8-2 TBA Concentration (ng/L)
25,000 25,300 25,300 25,700
Average Standard Deviation
9.0 0 QC1 9.0 0 QC2 9.0 0 QC3
1560 1560 1560
25,300 287
1590 1640 1680
Average Standard Deviation
9.0 5 9.0 5 9.0 5 9.0 5
1 2 3 4
Average Standard Deviation
9.0 5 QC1 9.0 5 QC2 9.0 5 QC3
-
1560 1560 1560
26,300 26,000 25,700 25,900
26,000 250
1550 1620 1510
Average Standard Deviation
% Recovery
-
102 105 108
105 3.0
-
99 104 97
100 3.6
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Hydrolytic Stability o f H-25435 as a Function o f pH
D u P on t-14965
Table 9: Daily temperature readings of the shaking water bath.
Time (Day)
pH
0 1.2 1 1.2 2 1.2 3 1.2 4 1.2 5 1.2
0 4.0 1 4.0 2 4.0 3 4.0 4 4.0 5 4.0
0 7.0 1 7.0 2 7.0 3 7.0 4 7.0 5 7.0
0 9.0 1 9.0 2 9.0 3 9.0 4 9.0 5 9.0
l
Temperature was not recorded.
Date Temperature (C)
03-June-2004 04-June-2004 05-June-2004 06-June-2004 07-June-2004 08-June-2004
37.0 37.0 37.0 37.0 37.0 37.0
13-May-2004 14-May-2004 15-May-2004 16-May-2004 17-May-2004 18-May-2004
50.0 50.0 50.0 50.0 50.0 50.0
06-May-2004 07-May-2004 08-May-2004 09-May-2004 10-May-2004 11-May-2004
50.0 50.0
X J
50.0 50.0
13-May-2004 14-May-2004 15-May-2004 16-May-2004 17-May-2004 18-May-2004
50.0 50.0 50.0 50.0 50.0 50.0
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AR-226
Hydrolytic Stability o f H -25435 as a Function o f pH
DuPont-14965
Table 10: pH measurements for Day 0 and Day 5.
Day
Rep
Nominal pH
Measured pH
0 1 1.2 1.27 0 2 1.2 1.25 0 3 1.2 1.27 0 4 1.2 1.27
5 1 1.2 1.24 5 2 1.2 1.23 5 3 1.2 1.24 5 4 1.2 1.24
0 1 4.0 4.10 0 2 4.0 4.11 0 3 4.0 4.09 0 4 4.0 4.10
5 1 4.0 4.07 5 2 4.0 4.07 5 3 4.0 4.08 5 4 4.0 4.07
0 1 7.0 7.10 0 2 7.0 7.09 0 3 7.0 7.10 0 4 7.0 7.10
5 1 7.0 7.06 5 2 7.0 7.07 5 3 7.0 7.07 5 4 7.0 7.05
0 1 9.0 9.10 0 2 9.0 9.10 0 3 9.0 9.08 0 4 9.0 9.09
5 1 9.0 9.09 5 2 9.0 9.09 5 3 9.0 9.08 5 4 9.0 9.07
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Hydrolytic Stability o f H -25435 as a Function o f pH
D u P on t-14965
FIGURES
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AR-226
Hydrolytic Stability o f H -25435 as a Function o f pH
Figure 1: Example calibration curves obtained for monitored[
DuPont-14965
].
Soeffidant of Determination: RA2 = 0.998822 Calibration curve: -3238.43 * x*2 + 140841 * x + Response type: External Std. Area Curve type: 2nd Order, Origin: Exdude, Weighting: 1/x, Axis trans:
Coefficient of Determination: RA2 * 0.998781 Calibration cu v e : -3018.23 x-2 + 122875 * x Response type: External Std, Area Curve type: 2nd Order, Origin: Exclude, Weighting: 1/x, Axis trams:
-36Company Sanitized. Does Not Contain TSCA CB1.
AR-226
Hydrolytic Stability o f H-25435 as a Function o f pH
Figure 2: Example chromatograms of monitored [ sample for pH 7.0 buffer on Day 0.
D u P on t-14965
]obtained for a blank
050704b06 pH 7, EUank 1. DayO
0.55
[1F3:SIR of 4 channels,ES7.402e+004
-37Company Sanitized. Does Not Contain TSCA CBI.
AR-226
Hydrolytic Stability o f H -25435 as a Function o f pH
D u P on t-14965
Figure 3: Example chromatograms of monitored [ calibration standard containing 5.3 mg/L H-25435,
] obtained for a
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AR-226
Hydrolytic Stability o f H -25435 as a Function o f pH
Figure 4: Example chromatograms of monitored [ hydrolytic stability sample.
D u P on t-14965
]obtained for Day 0, pH 7.0
050704b25 pH 7R ep1, DayO
1 0 0 -j
r sl)9 236259.2
%-
F2.S1R of 3 channels,ES0
0.51 0-
2.0
0507 )4b25 pH 7 *epl, DayO
1 0 0 -i
l 4.0 6.0
r J1
203 J4.1
8.0 10.0
F2:S!R of 3 channels,ESQ
2.240-005
%-
. It_______ ___ ________________ _ .. 2.0 4.0 6.0 8.0 10.0
0507 Mb25 pH 7 R ept, DayO 1 0 0 -j
%-
- ,3 5
56596.3
F3.SIR of 4 channels,ESQ
4.757e+005
2.0 4.0 6.0 8.0 10.0 12.0
-39Company Sanitized. Does Not Contain TSCA CBI.
AR-226
Hydrolytic Stability o f H -25435 as a Function o f pH
D u P on t-14965
Figure 5: Example chromatograms of monitored [ pH 7.0 hydrolytic stability sample.
051104b25 pH 7 R ept, Day 5
100
142012.2
FHSIR of 3 channels,ESn
6.244e+0Q5
]obtained for Day 5,
%-
^ 6 3 .5 .8 3 2.0 4.0 6.0 8.0 10.0
051104b25 pH 7 Rep1, Day 5
100-i
T& 236804.6
F2:S!R of 3 channels,ES-
a 2.676e+006
%-
0/49 2.0 4.0 6.0 8.0 10.0
051104b25 pH T R ept, Day 5
nF2:SlR of 3 channeis.ES2.106e+0G5
051104b25
Rep 1. Day 5
100-i
5.B5 537J 30.7
y v ) 'iii*v<"i 11 1>i<T~ min 10.0
F3:SIR of 4 channels,ESn
5.660e+006
2.6 4.0
0511 Mt>25 pH 7 Rep1, Day 5 1 0 0 -]
6.0 6 ' 10.6 ' 12.6
g. 5 525 57.9
F3.SIR of 4 channels, ES-
0 4.772e+005
2.0 4.0 6.0 8.0 10.0 12.0
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Hydrolytic Stability o f H -25435 as a Function o f pH
Figure 6: Representative calibration curve for 8-2 TBA.
D u P on t-14965
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AR-226
Hydrolytic Stability o f H -25435 as a Function o f pH
D u P on t-14965
Figure 7: Representative chromatograms of 104 pg/L 8-2 TBA calibration standard and 515 pg/L D-8-2 TBA internal standard.
051104b15 Smooth(Mn,1x3) std1
100-,
8-2TBA 3.50
16718.7
F1:SIR of 2 channels,ES463,523
7.490e+004
%-
n____, _, _'||.|_'|._||._I'_I'_I'_|_I_I_1_____I_I -)-r-T--T-T--JT--T 1.00 2.00
051104b15 Smooth(Mn,1x3) std1
100-,
IS-8-2TBA 3.48
78378.2
F2:SIR of 2 channels,ES467,528
3.339e+005
%-
'i"i1ri'i i1i'i. 4.00
min
- 42-
Company Sanitized. Does Not Contain TSCA CBI. AR-226
Hydrolytic Stability o f H -25435 as a Function o f pH
D u P on t-14965
Figure 8: Representative chromatograms for Day 5, pH 7.0 buffer blank sample.
- 43-
Company Sanitized. Does Not Contain TSCA CBI. AR-226
Hydrolytic Stability o f H-25435 as a Function o f pH
D u P on t-14965
Figure 9: Representative chromatograms for Day 5, pH 7.0 hydrolytic stability sample and 515 pg/L D-8-2 TBA internal standard.
051104b38 Smooth(Mn,1x3) rep 1; pH 7; Day 5
100-
8-2TBA 3.49
338073.C
F1 :SIR of 2 channels,ES463, 523
1.312e+006
%
i i i i | i i i i | i i ii | i i i"I""P i I i | I I I I 1.00 2.00 3.00
051104b38 Smooth(Mn,1x3) rep 1; pH 7; Day 5
100-
IS-8-2TBA 3.46
63591.5
| i I I I p f V r 'i' I I='"1'I- min
4.00
5.00
F2:SIR of 2 channels,ES467, 528
2.392e+005
1.00
2.00
3.00
4.00
5.00
t t t - min
-44Company Sanitized. Does Not Contain TSCA CBI.
AR-226
Hydrolytic Stability o f H -25435 as a Function o f pH
D u P on t-14965
Figure 10: Representative chromatograms for Day 0, pH 7.0 hydrolytic stability sample and 515 pg/L D-8-2 TBA internal standard.
012904b37 Smooth(Mn,1x3) rep 1; pH 7; DayO
100t
8-2TBA
F1 :SIR of 2 channels,ES463, 523
1 406e+006
%-
O' '"I.Illl1llll| 1 I rT"|'"T I 'l l | I 1 I ) | I i 1.00 2.00
3.00
4.00
5.00
- 45-
Company Sanitized. Does Not Contain TSCA CBI. AR-226
