Document Ran8KGg9RNZDqVnwmkB34oOOE

TRADE SECRET Study Title A flcL & - $ 3 & DuPont-13202 T e st G u id e l in e s : OECD Guideline 111 A u th o r : Bogdan Szostek, Ph.D. Study Completed on: November 24, 2003 Pe r f o r m in g L a b o r a t o r y : E.I. du Pont de Nemours and Company Haskell Laboratory for Health and Environmental Sciences Elkton Road, P.O. Box 50 Newark, Delaware 19714-0050 L aborato ry Project ID: DuPont-13202 soW o rk R eq uest N u m ber: Service C ode N um ber: Page 1 o f34 t s c &cbi DuPont-13202 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 consistent with the OECD Principles o f Good Laboratory Practice (as revised in 1997) published in ENV/MC/CHEM(98)17 and MAFF Japan Godd Laboratory Practice Standards (59 NohSan Number 3850. The test substance wa^xhamctgrized by the sponsor prior to the initiation of this study, except the percent content o | P H U The analysis o f percent content the test substance was completed after this study completion. 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 o f this study. Applicant / Sponsor EL du Pont de Nemours and Company Wilmington, Delaware 19898 U.S.A. Study Director Bogdan Szostek, PhD . Senior Research Chemist Date Applicant / Sponsor __________________________________________ __________ DuPont Representative Date Company Santtfeed. Does not contain TSCA CBt 2- - QUALITY ASSURANCE STATEMENT Haskell Sample Number(s): 25707 Dates of Inspections: Protocol: July 25, 2003 Conduct: August 1,2003 Records, Reports: September 30, 2003; October 1-3,6, 2003 Dates Findings Reported to: Study Director: October 15,2003; November 20, 2003 Management: October 15,2003; November 20, 2003 DuPont-13202 Reported by: K , l llL k i ........... Wonda K. iQslly Quality Assurance Auditor Date Ooupmy SanHizedl. / '"'n 3- - DuPont-13202 CERTIFICATION We, the undersigned, declare that this report provides an accurate evaluation o f data obtained from this study. Approved by: S. Mark KennadyAPh.D. Manager, Analytical Chemistry 2Y _ D ate Issued by Study D irector: -- ______________ j Bogdan Szostek, Ph.D. Senior Research Chemist - Aoqp, D ate Company Sanitized. D oes not contain TSCA CBI 4- - DuPont-13202 TABLE OF CONTENTS Page GOOD LABORATORY PR A C TIC E COMPLIANCE STA TEM EN T..................................... 2 QUALITY ASSURANCE STA TEM EN T..................................... 3 C ER TIFIC A TIO N ............................................................................. 4 LIST OF TA B LE S..................................................................................................................... .......... 6 LIST OF FIG U R E S ..............................................................................................................................6 STUDY IN FO R M A TIO N ................................................................................................................... 7 STUDY PERSO N NEL................................................................................... ...................................... 8 SUM M A RY .......................................................................... 9 INTRODUCTION.............................................................................................................................. .10 STUDY D ESIGN........................................... 10 M ATERIALS AND M E T H O D S.................................. 10 A. M aterials......................................... .10 1. Test Substance............................................................................................................................................. 10 2?f Preparation of buffer solutions.................................................................................................................... 11 3. Equipment..................................................................................................................................................... 12 B. Methods....................................................................................................................... 12 1. Preliminary test...................................................................................................................... 12 2. pH measurement and temperature................................................................................... 12 3. Analysis of test substance.................................................................................................. 13 4. Analysis of 8-2 TBA.................................................................................................................................... 13 5. Calculations..................................................................................................................................................14 RESULTS AND D ISCU SSIO N ........................................................ 16 CO NCLUSIO N S............................................................ 17 RECORDS AND SAMPLE STO R A G E......................................................................................... 17 R EFER EN C ES.....................................................................................................................................17 TABLES................................................................................................................................................ 18 FIGU RES.............................................................................................................................................. 29 -5 - LIST OF TABLES Page 1. Analytical results for aqueous stability 1.2, 37C, Days 0 (10-Sep-2003) and 5 (15-Sep-2003)............................... ZZ.... .............. .................................................................................... 19 2. Analytical results for aqueous stability 0f l |H H |H H p > o l i d s , pH 4.0, 50C, Days 0 (28-Aug-2003) and 5 (02-Sep-2003)............................... .................................................................................................................. 20 3. Analytical results for aqueous stability c ^ H ^ ^ H p i K o l i d s , pH 7.0, 50C, Days 0 (28-Aug-2003) and 5 (02-Sep-2003).............................. ...................... L .'T .................................................................................... 21 4. Analytical results for aqueous stability 9.0, S0C, Days 0 (03-Sep-2003) and 5 (08-Sep-2003)............................... ...................................7 ~ .........................................................................22 5. Measured concentrations o or pH 1.2, 37C, Days 0 (10-Sep-2003) and 5 (15-Sep-2003)................ 23 6. Measured concentrations o; or pH 4.0, 50C, Days 0 (28-Aug-2003) and 5 (02-Sep-2003)............... 24 7. Measured concentrations o. otpH 7.0, 50C, Days 0 (28-Aug-2003) and 5 (02-Sep-2003)...............25 8. Measured concentrations o or pH 9.0, 50C, Days 0 (03-Sep-2003) and 5 (08-Sep-2003)................26 9. Daily temperature readings of the shaking water bath............................................................................................. 27 10. pH measurements for Day 0 and Day 5.................................................................................................................... 28 LIST OF FIGURES Page 1. LC chromatograms of four Day 5, pH 1.2 implicate samples (A), 7500 mg/L (B), and 15000 mg/L (C) calibration standards 2. LC chromatograms of four pay 5, pH 4.0 ^ plicate samples (A), 7500 mg/L (B), and 15000 mg/L (C) calibration standards o f l H H f H ^ ^ B p o l i d s .................................................................................................30 3. LC chromatograms of four Bay 5, pH 7.0 ggplicate samples (A), 7500 mg/L (B), and 15000 mg/L (C) calibration standards o f l f l H B H U l S o l i d s ................................................................................................. 31 4. LC chromatograms of four Q jy ^ p H 9.0 replicate samples (A), 7500 mg/L (B), and 15000 mg/L (C) calibration standards 5. Representative LC chromatograms of blank samples for Day 5, pH 1.2,4.0, 7.0, and 9.0....................................32 6. Representative calibration curve 7. Representative chromatograms of 25.5 pg/l^H H j^pLalibration standard and 520 p g /l/f lH H jj|H A internal standard..........................................................mt...................................... ............... .................................... 33 8. Representative chromatograms for Day 0, pH 4.0 hydrolytic stability sample and 520 g g / L S ^ |M C T internal standard....................................................................................................................................... .22?!!:.......34 9. Representative chromatograms for Day 0, pH 4.0 blank sample............................................................................ 34 Company Sanitized, Does not contain TSCA CBI 6- - STUDY INFORMATION Substance Tested: H-25707 Submitter's Notebook Numbers'): Haskell Number: 25707 Purity: 100% Fluorinated polymer Known Impurities: Physical Characteristics: Off-white solid Stability: The test substance appeared to be stable under the conditions o f the study; no evidence o f instability was observed. Sponsor: E.I. du Pont de Nemours and Company Wilmington, Delaware 19898 U.S.A. Study Initiated/Completed: June 6, 2003 / (see report cover page) Experimental Start/Comnletion: June 9,2003 / September 15, 2003 Gompany Sanitized. D oes not contain TSCA CBt STUDY PERSONNEL Study Director: Bogdan Szostek, Ph.D. Primary Technician: Keith B. Prickett, B.S. Management: S. Mark Kennedy, Ph.D Analytical Associate: Keith B. Prickett, B.S. Analytical Chemist: Bogdan Szostek, Ph.D. Management: S. Mark Kennedy, Ph.D. Toxicology Report Preparation: Lisa G. Burchfield, A.A. Management: Nancy S. Selzer, M.S. Company Sanitized. D oes not contain TSCA CBI /" " "N DuPont-13202 SUMMARY The hydrolytic stability oij was testedji_bufiered aqueous solutions at pH 4.0, 7D, 9.0 at 50C and pH 1.2 at 37C for nve days. T h e fifl0 P H H H V ^ M M n was demonstrated to be hydrolytically stable under these test conditions. The stability was demonstrated by the comparison o f the percent recoveries of _______ tor Day 0 and Day 5 samples and analysis o f chromatographic peak shape for Day 5 samples. The average percent recoveries for the Day 0 and Day 5, pH 1.2 replicate samples are 101 1.2% and 101 1.0%, respectively. The average percent recoveries for the Day 0 and Day 5, pH 4.0 replicate samples are 100 0.5% and 97 2.1% , respectively. The average percent recoveries for the Day 0 and Day 5, pH 7.0 replicate samples are 101 1.3% and 100 0.8%, respectively. The average percent recoveries for the Day 0 and Day 5, pH 9.0 replicate samples are 100 1.0% and 104 2.5%, respectively. The difference in percent recoveries for Day 0 and Day 5 samples did not reach the critical level o f 10% that would indicate the hydrolysis of the test substance for any o f the investigated test conditions. The observed increase o ^ JP lB p c o n cg n tratio n is approximately two orders o f magnitude lower than the concentration o i y B B M skpected if 10% o f the test substance hydrolyzed. In addition, the measured concentrations o: indicated an increase of thq^B B (B jponcentration between Day 0 and Day 5. J^ieincsease o f ' * B oncentration can be accounted for by the presence o f the res id u a y H flM n 'a w material present in the test substance. Company Sanitized. D oes not contain TSCA CBI -9 - DuPont-13202 INTRODUCTION _The purpose o f the study was to investigate the hydrolytic stability W L. pH 1.2 ^ .0 , 7.0, and 9.0 aqueous buffer solutions. , ) [which was patterned after the the concentration o f th^ _ /as monitored to investigate i: Is released from the test substance during the test. STUDY DESIGN A preliminary test was performed on the test substance at 50 0 .1C at each o f the pH 4.0, 7.0, 9.0 and at 37 0.1C for pH 1.2. If less than 10% o f 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 The study design complies with the following test guidelines: OECD Guideline 1 1 1 ^ with the exception that the OECD Guideline 111 requires that the amount o f the test substance used is the smaller of 0.01 M or half the saturation concentration in water. The test s u b s ta n c ^ jjl^ ( p P H H H H I H i s insoluble in water and the analytical method does not allow detection o f the test substance at trace levels. Therefore, the study was conducted with an excess o f undissolved test substance. A. Materials MATERIALS AND METHODS The test substance was characterized to determine th characterization parameters: Mn (number average), PDI (polydispersity index) with the following results: content, % fluorine, and polymer t average), Mz (z average), and sanitized. D oes not contain C om p aq Sanitize* - 10- L -/o Fluorine (w/w): Mn: Mw: Mz: PDI: 38.8% 5660 23,400 43,900 4.13 2. Preparation o f buffer solutions The following buffer solutions were prepared as described in the Annex to OECD guideline 111(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 o f water. The 0.2 M hydrochloric acid (HC1) was prepared by diluting 1.83 g 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 161.3 mL of a 0.2 M HC1 solution into a 500-mL flask and bringing the flask to volume with water. The pH of the buffer was adjusted to 1.2 with hydrochloric acid. b. pH 4: The 0.1 M solution o f potassium hydrogen phthalate was prepared by dissolving 10.21 g of potassium hydrogen phthalate (purity 99+%, Sigma) in 500 mL o f water. The pH 4 buffer solution was prepared by placing 2 mL of a 0.1 M sodium hydroxide solution and 250 mL o f 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 o f potassium phosphate was prepared by dissolving 6.81 g o f potassium phosphate, monobasic (KH2PO 4, purity 99.5%, Sigma) in 500 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 of the buffer was adjusted to 7.0 with sodium hydroxide. d. pH 9: The 0.1 M boric acid (H3BO3) solution in 0.1 M potassium chloride (KC1) was prepared by dissolving 3.73 g o f KC1 (purity 99.0%, Sigma) and 3.09 g o f H3BO3 (purity 99.5%, Sigma) in 500 mL of water. ThqpH 9 buffer solution was prepared by placing 250 mL of 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 o f the buffer was adjusted to 9.0 with sodium hydroxide. Water used for buffer preparation was obtained from a Bamstead NANOpure Diamond water purification system. The water resistivity is >18.2 megaohm*cm . All buffer solutions were - II - /!***\ DuPont-13202 sterilized prior to use by filtering through Coming Sterilization Filter systems with 0.22 pm cellulose acetate filters. The buffer solutions were stored at room temperature. 3. Equipment p H Meier: Water Bath: Autoclave: Automatic Pipettes: Balance: Sonicator: Centrifuge: Vortexer: Glass Vials: Orion Model 250 A Water bath: Lab-Line Force Reciprocating Water Bath Shaker, Model 4682 AMSCO Model 3023 Research Pro 100, 1000, 5000; Eppendorff Mettler AE-100 Branson Model 1210 or Branson Model 5200 Sorvall RT7 Multi-tube vortexer, VWR 10 mL borosilicate crimp top glass vials (Supelco) with 20 mm T/butyl septa and aluminum foil. The vials, foil, and septa were autoclaved at 121 C for 30 min before use. B. Methods 1. Preliminary test The preliminary test was performed 50C at each o f the pH 4.0, 7.0, 9.0 and at 37C for pH 1.2. The same sample preparation procedure an^atudy design was followed for each o f the pH levels tested. Approximately 100 mg o f tl was weighed into each of sixteen 10-mL autoclaved glass vials and 3 mL o f appropriate buffer' solution was added. The vials were crimp capped with the bottom o f the septum covered with autoclaved aluminum foil. Eight o f 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 Dav 5 blanks. Additionally, six vials containing 3 mL of appropriate buffer were spiked w i t l ^ P H H u toc^ The vials were crimp capped with the bottom o f the septum covered with autocl ayed aluminum foil. Three o f the vials were used as the quality control (QC) samples fo i(J ^ P B | n a ly s is on Day 0. The other three vials were wrapped in aluminum foil and placed in the water bgth together with eight o f the test substance vials. These served a role of QC samples fo t^ B P ^ jY m a ly sis on Day 5. Four of the Day 0 or d two blank vials were processed for the pH measurement and thi etermination. The other fo)ujn^ppf the Qay i0 or Day 5 vials, two bl vials were processed for the determination. 2. pH measurement and temperature. The pH was measured on Day 0 and substance that were designated for th uncapped, the pH electrode wasjptroduced to vials were further processed fo: g the buffer and the test alysis. The vials were d the pH reading was recorded. These lysis. The temperature of the water Company Sanitized. Does not contain TSCA CBI -12- DuPont-13202 bath was monitored daily. The water bath temperature readings from the automatic temperature control were checked and recorded in the study records daily. 3. Analysis of test substance After the pH measurement, 0.3 mL of d i l u t e c j / f l | H | ^ J H p H H H H | H | p i w a s added to each vial, the vials were recapped, vortexe<mor ten minutes, and centrifuged for fifteen minutes ayS200 rpm. The addition of the phosphate fluorosurfactant is necessary for wetting of t h e j p f l ^ ^ l l l l f r a n d facilitates the pelleting o f the material at the bottom o f the vial after centrifugation. After centrifugation the vials were uncapped and the supernatant was discarded using a manual pipetter. The remaining solid material was dissolved by adding 10 mL of tetrahydrofuran (THF) and sonication for 30 minutes. Approximately, 1 mL of the THF solution was placed in a glass LC vial and subjected to LC-GPC analysis. The blank samples were subjected to the same procedure except the pH measurement. The stocktsolution of t h e n p 0 V M H B H H H f l^ ras made by dissolving 0.15 g o fl B H H f l p n 10 mL o f Th F! Thirty minutes of sopication were required to completely dissolve the test substance. Calibration standards for th fP B H H B H M M W p n a ly s is were made by a p p ropriate dilution o ^ h e stock. The calibration standards ranged from 3000 to 15000 mg/L Fresh calibration standards were prepared for each instance o f analysis. "Linear regression o f the peak height vs. standard concentration was used to establish the calibration curves. Instrumentation and conditions: LC System: Column: LC Solvent: Flow Rate: Column Temperature: RI Detector Temperature: Data Collection: Injection Volume: Agilent HP 1100 with Agilent Refractive Index Detector (RID) Styragel HR 4, 7.8 mm x 300 mm, Waters Tetrahydrofuran (THF) 1 mL/min 40 C 35 C 0-15 min 50 pL 4. Analysis o Samples designated f o tf l^ P H a n a ly s is were processed by injecting 6 mL acetonitrile through the vial septum using a glass syfmge. NeitljgrDayO nor Day 5 samples were uncapped before acetonitrile injection to avoid any losses v^ s containing injected acetonitrile were vortexed for 15 minutes on a multitube vortexer. Subsequently, the vials were uncapped and 1 mL o f the extract was transferred to a glass LC vial, spiked with internal standard, crimp capped, and subjected to LC/MS analysis foJ B W ts the analytical standard___ useffaslhe internal standard. Stock solutions (1000 mg/L) of the analytical standard anathe internal standard were prepared in methanol and were stored refrigerated. The calibration standards were prepared fresh for each calibration in 50/50% Company Sanitized. Does not cmtaim TSCACB1 - 13 - * t watg^iacetonitrile. The calibration standards were made in the range frorq_25 to 25^0 g A constant concentration o f internal standard was used: 520 pg/' The calibration curves were contracted using tljg ratio qfJb e peak area for ions m/z d peak area for ions monitored f o r H ^ ^ P ^ . n id the concentrations o 94* Instrumentation and conditions: HPLC Instrument: MS Instrument: Model 2795, Waters ZQ, Micromass LC Parameters: Column: Mobile Phase: Column Temperature: Injection Volume: Column Switch: Luna 3u C l8 (2), 20 mm x 2mm 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 Flow (mL/min) 70 0.25 70 0.25 100 0.25 100 0.25 70 0.25 70 0.15 MS Parameters: Ionization mode: Capillary voltage: Cone voltage: Source Temperature: Desolvation Temperature: Data Acquisition Function: Electrospray, negative ions 2.5 kV 10 V 120C 300C 8-2 TBA: SIR* of 463, 509,523; 0-6 min D- 8-2 TBA: SIR of 468, 514, 528 m/z; 0-6 min * These ions representJhe deprotonated ij ion o f o r m a t e adduct and acetate adduct o g H H H H P V f j IThe formate'ahd acetate adducfare formed becaus^TF residual presencwr of acetate and/or formate in the system I lginating from other methods that are run on the system and utilize these components in the mobile phase. 5. Calculations The follgying calculations were used to determine tbs nominal concentrations of jH H f lf lf T a b le s 1-4). For example, if 0.1009 g ofq M H ^ H H H H B H K v a s weighed into the vial andmibsequently dissolved in 10 mL of THF duflng sample processing, this would represent the nominal concentration o f 100.9 mg/0,01 L= 10,090 mg/L. The percent recovery is calculated by dividing the measured concentration by the nominal concentration, times 100%. The expected (nominal) C O ncentra^pnojB pJJJuin tihuec spiiktedu samples wwas ecailecuilaiteud as f* f o l i o s: the 4.5 pL of 1020 m g/I^p B jjjj^p ik e5 ep resen ts 4.5 uL x 1020 ng/pL= 4590 ngQ p} Total volume of extract is 9 mL: 3 fffL of buffer and 6 mL o f acetonitrile. The expected Company Sanitized. D oes not contain TSCA CBl - 14- DuPont-13202 nominal concentration o recovery is calculated by times 100%. the extract is: 4590 ng/9 mL=510 ng/mL. The percent ie measured concentration by the nominal concentration, The Dected concentration o l|jp f lf c f tin the acetonitrile extract if 10% o f t h ^ j l rydrol^zed can be estimated in the following way. Ten percent of 100 mg oj epresents 10 mg. Taking into account that the test sidgstancecontam 38.8% fluorine and assuming that all fluorine is present in the form of bouncflM H flH tie hydrolysis of the ^ mg o f the test substance would release: 10 mg x 0.38^/(324 i^ g T/mmol 4H b 164.1 mg/mmole)=5.56 This am oun^ovH H H H vould be contained in 9 mLs of extract. Therefore, expected concentration o l J J ^ ^ p p m u d ^ e: 5.56 mg/0.009 L= 618 mg/L. The; concentration o: extraction, is: lOi (1780 pg/L). rq^erial,^jP P B P expected, maximum Ifclpreseht in the extract, assuming complete ((I016%/100)/0.009 L= 1.78 mg/L Company Sanitized. Does not contain TSCACBf - 15- DuPont-13202 RESULTS AND DISCUSSION Tables 1-4 present the results for measured concentrations percent recoveries obtained for Day 0 and Day 5 replicate samples fo^>HL2^X)^H)^H)^respectively. The average percent recoveries for fne Day 0 and Day 5, pH 1.2 replicate samples are 101 1.2% and 101 1.0%, respectively. The average percent recoveries for the D ay 0 and Day 5, pH 4.0 replicate samples are 100 0.5% and 97 2.1%, respectively. The average percent recoveries for the Day 0 and Day 5, pH 7.0 replicate samples are 101 1.3% and 100 0.8%, respectively. The average percent recoveries for the Day 0 and Day 5, pH 9.0 replicate samples are 100 1.0% and 104 2.5%, respectively. The relative difference between the Day 5 and Day 0 concentrations o f th&,test substance was Ift&s than 10% for any o f the investigated pHs. This indicates that t h ^ ^ H H H H H H K s hydrolytically stable at investigated pHs. Figures 1 to 4 compare the LC chromatograrffs of four replicates o f Day 5 samples with the chromatograms o f two calibration standards for pH 1.2,4.0, 7.0, 9.0, respectively. The shape of the chromatographic peak, eluting fro g 7 to 11 minutes, in d i c a t i v e o f the molecular weight distribution o f th M p B f lH B M H S M B The hydrolysis o j U f J / f * /ould result in change o f me obsejy ed peak shape. Corggarison o f thepeak ~ shape of the fifthly made calibration standards the Day 5 samples indicates that no evident change of the peak shapefor Da)fc5 samples wits observed for any o f the investigated pHs. This further supports the conclusion o i p P |P p p M |p H B M i y d r o l y t i c stability. Figure 5 presents the LC chromatograms o f the Day 5 blank samples ior pH 1.2,4.0, 7.0, and 9.0. Additional evidence supporting the hydrolytic stability o: an be.derived from the measured concentration ThehydroJygus o: ould result in the release o n B H H B a n d the increase o f lH f M c o n c e n tra tio n in tb est system. Based on the charcterization data, the tesi substance contains 0.016% o f residu: Assuming that the buffer extracts all tb ing the time o f hydrolytic stability experiment, the expected concentration o: the processed extract (3 mL buffer and 6mL acetonitrileyvouM be 1.78 mg/L (seeT3.5 Ca illations). Similarly,J|ie expected concentration o iM p B 0 t[n the extract, if 10% o as hydrobfzeu, is 618 mg/L (see B.5 Calculations). Tables 5-8 pres' it the measured concentrations in the extracts for Day 0 and Day 5 re samples for pH 1.2, 4.0, 7.0, 9.0, respectively. The" data shows a significant increase, oncentration in the Day 5 samples, when the average concentration o t ^ H ^ p f l f o r Day Onnd Day 5 samples are compared. j FIowever, the concentrations reach only some o f the expected level of fully e x t r a c t e w ( ^ n ( 1780 pg/L) and these are approximately two orders o f magnitude lower than the concentrations expected when 10% o f the test substance would hydrolyze. The increase of th c jM B H rf i^ c e n tmtion between th(_Dav (Land Day 5 samples is simply a result of extraction of t h J l H H K f r o n M | | | ^ p ( f f e H B ^ u r i n g the h y d ro p ic stability experiment at elevated temperature (^.0C or 3JC). The percent recovery results foiW M M reT ables 5-8) support the validity of thq measurement in the Day 0 arm Day 5 samples for all investigated pHs. The Day 5 saiiiple percent recovery results demonstrate that no significant loss o ^ p ( ( J ^ ^ c c u r r e d from the test system during the hydrolytic stability experiment. Figure 6 shows representative calibration gom paay Sanitized. Does ssotconteltt TSCA Cffl - 16- curve obtained fo ^ B H B p ^ F ig u re s 7-9 show chromatograms fo: Day 0, pH 4.0 test sample, and Day 0, pH 4.0 blank sample, respectively. alibration standard, Table 9 summarizes the water bath temperature data 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 0.1C. Table 10 summarizes results o f the pH measurements for Day 0 and Day 5 samples for pH 1.2, 4.0, 7.0, and 9.0. The measured pH for Day 0 and Day 5 replicate samples for all o f the investigated pHs was at the specified nominal level with the deviations not exceeding 0.1. CONCLUSIONS The hydrolytic stability of was tested, buffered aqueous sgjytions at pH 470", 717, 9.0 at 50C and pH 1.2 at 37C for trve days. T h q |6 p ^ j j ( ( J j ( # S w a ^ r a i o n s t r a t e d t o be hydrolytically stable under these test conditions, m e stabilityjaf demonstrated by the comparison of the percent recoveries o f j B t f f l H H f l H H E>ay 0 and Day 5 samples and analysis o f chromatographic peak shape for Day 5 samples. The difference in percent recoveries for Day 0 and Day 5 samples did not reach the critical level o f 10% that would indicate the hydrolysis o f the test substanceTor any of the investigated test conditions. In addition, the measured concentrations o f f iM jB R n d ^ te d an increase o f thaflMHHHponcentration between Day 0 and Day 5. _heincrc^se of thsjB JJB M poncentration canJjeaccomited for by the presence of the residuaV flflM B tn the test substance. The increase o f f l H t f b o a e entratifin is approximately two orders of magnitude lower than the concentration expected if 10% o f the test substance hydrolyzed. --' 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, QECD Guideline 9No. 111. - 17- Company Sanitized. D oes not contain TSCA CBt DuPont-13202 TABLES Company Sanitized. D oes not contain TSCA CBI - 18- Table 1: Analytical results for aqueous stability ofj Days 0 (10-Sep-2003) and 5 (15-Sep-2003). DuPont-13202 >H 1.2,37C, pH Day Rep 1.2 0 1 1.2 0 2 1.2 0 3 1.2 0 4 (mg/L) 10100 10100 10000 10200 1.2 5 1 1.2 5 2 1.2 5 3 12 5 4 _________ 10200 10000 10200 10300 (mglL) 10100 10300 10200 10200 Average Standard Deviation 10100 10100 10300 10400 Average Standard Deviation % Recovery 100 102 102 100 101 1.2 99 101 101 101 101 1.0 Company Sanitized. D oes not contain TSCA CB1 - 19- ! ^ mmN Table 2: Analytical results for aqueous stability o j Days 0 (28-Aug-2003) and 5 (02-Sep-200T^ H 4 .0 , 50C, pH Day Rep 4.0 0 1 4.0 0 2 4.0 0 3 4.0 0 4 4.0 5 1 4.0 5 2 4.0 5 3 4.0 5 4 (mg/L) 10100 10100 10000 10200 10200 10100 10100 10100 ^G!suSS!SnS5iOTi (mg/L) 9960 10100 10000 10200 Average Standard Deviation 9660 9810 10100 9810 Average Standard Deviation % Recovery 99 100 100 100 100 0.5 95 97 100 97 97 2.1 - 20- Company Sanitized. D oes not contain TSCA CB1 DuPont-13202 Table 3: Analytical results for aqueous stabilliitty 0 o0n y f 8300-82 SolidsipH 7.0, S0C, Days 0 (28-Aug-2003) and 5 (02-Sep-200j}T `3 " S kH pH Day Rep jvominai Concentration (mg/L) 7.0 0 1 7.0 0 2 7.0 0 3 7.0 0 4 10200 10100 10100 10000 (mg/L) 10100 10200 10300 10100 Average Standard Deviation 7# 5 1 7.0 5 2 7.0 5 3 7.0 5 4 10200 10200 10100 10000 10200 10200 10200 9910 Average Standard Deviation % Recovery 99 101 102 101 101 1.3 100 100 101 99 100 0.8 s***'. Company Sanitized. D oes not contain TSCA CSi -21 - Table 4 Analytical results for aqueous stability o Days 0 (03-Sep-2003) and 5 (08-Sep-2003 pH Day Rep 9.0 0 1 9.0 0 2 9.0 0 3 9.0 0 4 9.0 5 1 9.0 5 2 9.0 5 3 9.0 5 4 (mg/L) 10100 10000 10000 10100 10000 10100 10000 10300 Measured Concentration (mg/L) 9950 10100 10000 10000 Average Standard Deviation 10200 10300 10500 11000 Average Standard Deviation % Recovery 99 101 100 99 100 1.0 102 102 105 107 104 2.5 r,*v Company Sanitized. Does not contain TSCA GBI - 22- Table 5: Measured concentrations o and 5 (15-Sep-2003). DuPont-13202 or pH 1.2,37C, Days 0 (10-Sep-2003) NominaHPSBHn pH Day Rep * Concentratn (pg/L) M easureqPBM BtJ Concentration (p g / ^ % Recovery 1.2 0 1 1.2 0 2 1.2 0 3 1.2 0 4 - 727 774 762 895 - Average Standard Deviation 790 73 1.2 0 QCl 1.2 0 QC2 1.2 0 QC3 510 510 510 595 117 594 116 633 124 Average Standard Deviation 119 4.4 1.2 5 1 1.2 5 2 1.2 5 3 1.2 5 4 - 493 688 658 490 - Average Standard Deviation 582 110 1.2 5 QCl 1.2 5 QC2 1-2 5 QC3 510 510 510 440 86 471 92 476 93 Average Standard Deviation s QCl, QC2 and QCJrepn^Hjt threJMBBBBfccoverv samples for each test day. b Concentrations o M fliH B jn ft16extract (3 mL buffer and 6 mL acetonitrile). 90 3.8 Company Sanitized. Does not contain TSCA CB) -23 - Table 6 Measured concentrations o and 5 (02-Sep-2003). DuPont-13202 for pH 4 .0 ,50C, Days 0 (28-Aug-2003) m nhm 3 C m m m m m J ti % Recovery pH Day R e p ' Concentration (jig/L) Concentration (pg/L c 4.0 0 1 4.0 0 2 4.0 0 3 4.0 0 4 - 778 699 795 740 - Average Standard Deviation 753 43 4.0 0 QC1 4.0 0 QC2 4.0 0 QC3 510 510 510 495 97 508 100 487 95 Average Standard Deviation 97 2.5 4.0 5 1 4.0 5 2 4.0 5 3 4.0 5 4 - 1560 1680 1680 1430 - Average Standard Deviation 1590 120 4.0 5 QC1 4.0 5 QC2 4.0 5 QC3 510 510 .510 464 91 471 92 447 88 Average Standard Deviation s QC1, QC2 and QC^^epr^git thre^JB B Pyecovery samples for each test day. b Concentrations ofa^jN p S In the extract (3 mL buffer and 6 mL acetonitrile). 90 2.1 Company Sanitteed. D oes not contain TSCA CB1 - 24- DuPont-13202 Table 7: Measured concentrations ofi nr pH 7.0, 50C, Days 0 (28-Aug-2003) and 5 (02-Sep-2003). ' pH Day Rep" 7.0 0 1 7.0 0 2 7.0 0 3 7.0 0 4 - - 796 875 841 801 Average Standard Deviation 828 37 7.0 0 QC1 7.0 0 QC2 7.0 0 QC3 510 510 510 519 494 499 Average Standard Deviation 7.0 5 1 7.0 5 2 7.0 5 3 7.0 5 4 - 1670 1300 1800 1380 Average Standard Deviation 1540 240 7.0 5 QC1 7.0 5 QC2 7.0 5 QC3 510 510 510 565 411 462 * QC1, QC2 and Qi b Concentrations o: Average Standard Deviation ecovery samples for each test day. mL buffer and 6 mL acetonitrile). % Recovery - 102 97 98 99 2.7 - Ill 81 91 94 15 Company Sanitized. D oes not contain TSC CB) - 25- DuPont-13202 Table 8: Measured concentrations o t f l H H p f o r pH 9 .0 ,50C, Days 0 (03-Sep-2003) and 5 (08-Sep-2003). pH Day R ep ' 9.0 0 1 9.0 0 2 9.0 0 3 9.0 0 4 - ^S ncentration (pg/L*J^ % Recovery 796 519 864 704 - Average Standard Deviation 721 150 9.0 0 QCl 9.0 0 QC2 9.0 0 QC3 510 510 510 483 95 492 96 -- Average Standard Deviation 96 0.7 9.0 5 1 9.0 5 2 9.0 5 3 9.0 5 4 - 800 757 868 1010 - Average Standard Deviation 859 110 9.0 5 QCl 9.0 5 QC2 9.0 5 QC3 510 510 510 437 86 472 93 417 82 Average Standard Deviation ' QCl, QC2 and Qgjreprgjfyt thre^PHPW(feeovery samples for each test day. b Concentrations o f lB f lp u in ^ie extract (3 InL buffer and 6 mL acetonitrile). 87 5.6 Company Sanitized. D oes not contain TSCA CBl - 26- Table 9: Daily temperature readings of the shaking water bath. DuPont-13202 Time (Day) pH Temperature Date (C) 0 1.2 10-Sep-2003 37.1 1 1.2 11-Sep-2003 37.0 2 1.2 12-Sep-2003 37.0 3 1.2 13-Sep-2003 37.0 4 1.2 14-Sep-2003 37.1 5 1.2 15-Sep-2003 37.0 0 4.0 28-Aug-2003 50.0 1 4.0 29-Aug-2003 50.1 2 4.0 30-Aug-2003 50.0 3 4.0 31-Aug-2003 49.9 4 4.0 01-Sep-2003 50.0 5 4.0 02-Sep-2003 50.0 0 7.0 28-Aug-2003 50.0 I 7.0 29-Aug-2003 50.1 2 7.0 30-Aug-2003 50.0 3 7.0 31-Aug-2003 49.9 4 7.0 01-Sep-2003 50.0 5 7.0 02-Sep-2003 50.0 0 9.0 03-Sep-2003 50.0 I 9.0 04-Sep-2003 50.0 2 9.0 05-Sep-2003 49.9 3 9.0 06-Sep-2003 50.0 4 9.0 07-Sep-2003 50.0 5 9.0 08-Sep-2003 50.0 TM a n v Sanitized. Does - 27- SCACBI Table 10: pH measurements for Day 0 and Day 5. Day 0 0 0 0 5 5 5 5 0 0 0 0 5 5 5 5 0 0 0 0 5 5 5 5 0 0 0 0 5 5 5 5 Rep Nominal pH Measured pH 1 1.2 1.25 2 1.2 1.25 3 U 1.26 4 1.2 1.25 1 1.2 1.29 2 1.2 1.30 3 1.2 1.28 4 1.2 1.29 1 4.0 4.10 2 4.0 4.10 3 4.0 4.10 4 4.0 4.10 1 4.0 4.10 2 4.0 4.10 3 4.0 4.10 4 4.0 4.10 1 7.0 7.05 2 7.0 7.06 3 7.0 7.05 4 7.0 7.05 1 7.0 7.10 2 7.0 7.10 3 7.0 7.10 4 7.0 7.10 1 9.0 9.05 2 9.0 9.06 3 9.0 9.05 4 9.0 9.05 1 9.0 9.10 2 9.0 9.08 3 9.0 9.09 4 9.0 9.08 Company Sanitized. D oes not contain TSCACB1 / '"'S DuPont-13202 FIGURES /-N Company Sanitized. Does not contain TSCA CSf - 29- DuPont-13202 /" v Figure 1: LC chromatograms of four Day 5, pH 1.2 ref gate samples (A), 7500 mg/L (B), and 15000 mg/L (C) calibration standards ofi Figure 2: LC chromatograms o f four Day 5, pH 4.0 replicate s pies (A), 75Q0 mg/L (B), and 15000 mg/L (C) calibration standards o Company Samlfcad. D o t. not m a m TSCA CB| - 30- DuPont-13202 Figure 3: LC chromatograms of four Day 5, pH 7.0 replicate^aipples (A), 7500 mg/L (B), and 15000 mg/L (C) calibration standards of r ^ - Figure AT LC chromatograms of four Day 5, pH 9.0 rep, patejamples (A), 7500 mg/L (B), and 15000 mg/L (C) calibration standards ofi 0 Company Sanitized. D oes not contain TSCA CB1 -31 - Figure 5 DuPont-13202 Representative LC chromatograms of blank samples for Day 5, pH 1.2,4. 0, 7.0, and 9.0. Company Sanitized. D oes not contain TSCA Cfil - 32- Figure 6: Representative calibration curve for. DuPont-13202 Figure GmnpangSanitized. Does not contain t&CAcm -33 - DuPont-13202 Figure 8: Representativ^hromatograms for Day 0, pH 4.0 hydrolytic stability sample and 520 pg/LA H H H BJinternal standard. Figure 9: Representative chromatograms for Day 0, pH 4.0 blank sample. Gompany Sanitized. Does noi coniai?! TSCA CB - 34-