Document La9K6zZm6YxMvJqr0MkvV4Vd

P H nu- aoif. 486 DuPont Haskell Laboratory for Health and Environmental Sciences M zu - i m TO: FROM: MRFIL] POCKET H-24921 J. C. Maslanka October 30, 2003 *-- Quality Assurance E. Mylchreest TEST SUBSTANCE STABILITY O F H-24921 Medical Research Project Number: Haskell Sample Number: Haskell Number (Analytical Reference): Analytical Test Code: Analytical Report Number: '24921 ^ 2492L (m m j DuPont-13707 Notebook References: 3 Attached is the analytical report to satisfy protocol requirements for studies with H-24921 but may also be used for other purposes. not c o n ta i TSC CBi Company Sanitized- 0es TEST SUBSTANCE STABILITY OF H-24921 Medical Research Project Number: Haskell Sample Number: Analytical Report Number: DuPont-13707 A sample of H-24921 was received July 11, 2003, and a i ^ y z e ^ n J ^ y 14, 2003. percentage of active ingredient (a.i.) was measured to bbi e f l ^ H H B w i t h a r a n g e o B H po I | M H R f nominal for replicate analyses (n=3). The sponsor reported a purity o\f l ^ j B wwhheen ^ tn ^ asammpplle was submitted. ACKNOWLEGMENTS Sample preparation by S. A. Riley (Chemistry Technician) and sample analysis by T. Ryan (Staff Chemist). SIGNATURES Report by: Janet C. Maslanka Senior Staff Chemist Date Date Issued: 3o,*. c4J<it--&aa3 ueffcontain TSCA CB1 DuPont-13707 page 2 of 3 METHODS Analysis o f H-24921 was by high-performance liquid chromatography/tandem mass spectrometry (IXj-MS/MS). Negative-ion electrospray is used to generate negatively charged ions o f H H S h e ions are selected with the first MS quadrupole, collisionally dissociated using argon, and a fragment ion is monitored.| H i s used as an internal standard. SAMPLE PREPARATION & ANALYSIS Aliquots (0.0300, 0.0301, and 0.0322 grams) of H-24921 were dissolved in Nanopure water (100 mL) to the nominal concentrations of 300, 301, and 322 ppm. These were further diluted to 0.00315,0.00316, and 0.00338 ppm with the appropriate amount of internal standard added and analyzed according to the following method. CHROMATOGRAPHIC CONDITIONS LC parameters Instrument: Column: Flow Rate: Injection Volume: Column Temperature: Column Switch: Mobile Phase: Gradient: Time (min.) 0.0 0.9 1.0 6.0 6.1 7.0 Hewlett Packard Model 1100 HPLC Zorbax RX-C8, 2.1 mm x 150 mm, 5/xm 0.4 mL/min 50 pi 30C 4.0 minutes 0.15% Acetic Acid/Acetonitrile % Acetonitrile 5 5 80 80 5 5 MS warameters Instrument: Micromass Quattro Micro Tandem MS Ionization mode: Electrospray (ESI), negative ion Capillary voltage: -2.7 kV Cone Voltage: 15 V Source Temperature: 120 Desolvation Temperature: ^ 350 ^ Scan function: 13 m/z (parent) to 369 m/z (daughter) T ] H 0 f l | 0 M Q 4 1 5 n^z (parent) to 369 m/z (d au g h te^ ^ ** Retention Time o1 injections) lam 'approximately 5.2 min (50 fiL Company Sanitized. Does not contain TSCA CB1 DuPont-13707 page 3 of 3 CALIBRATION & QUANTITATION A stock solution of the H-24921 (separate sample used as analytical reference) was made in Nanopure water. Appropriate aliquots of the stock were diluted with Nanopure water to make calibration standards that bracketed the target concentration of the diluted sample solutions^Before these aliquots were brought to volume, an appropriate amount of internal s t e n d a r d ^ l H H i H f l | H f l | | H f l f l f l ^ as added. Analysis of H-24921 was by high-performance liquid chromatography/tandem mass spectrojpetry^LC-MS/MS). Negative-ion electrospray is used to generate negatively charged ions o q P B 0 th e ions are selected with the fust MS quadrupole, collisionallyj^ssociated using argon, and a fragment ion is m o n ito m d .0 H H ||H H H B H M H R M H B s use^ as an internal standard. Triplicate injection of the sample solutions and calibration standards solutions were made and peak areas were calculated electronically. The calibration curve was generated by regression analysis using the peak area ratio from the H-24921 and the internal standard. Data for test solutions were compared to the calibration curves to evaluate the concentrations of the H-24921. RESULTS H-24921 eluted from the HPLC column as a resolved peak with a retention time approximately 5.2 minutes for the negative ion. The sponsor reported a purity the sample was submitted. Table 1. The percent of a.i. in the H-24921 sample analyzed July 14,2003. ppm H-24921 Percent Aliquot Targeted Measured Nominal 1 0.00315 0.00291 92.4 2 0.00316 0.00287 90.8 3 0.00338 0.00313 92.6 Average Percent Nominal Standard Deviation Coefficient of Variation 91.9 1.0 1% hen Company Sanitized. Does not contain TSCA CBS