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Evaluation AccuTOF DART for Postmortem Toxicology Screening Peter R. Stout

Evaluation AccuTOF DART for Postmortem Toxicology Screening Peter R. Stout. RTI International is a trade name of Research Triangle Institute. NIJ Project. Grant No. 2006-DN-BX-K014 Opinions are those of the authors and not necessarily the U.S. Department of Justice. Objective.

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Evaluation AccuTOF DART for Postmortem Toxicology Screening Peter R. Stout

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  1. Evaluation AccuTOFDART for Postmortem Toxicology ScreeningPeter R. Stout RTI International is a trade name of Research Triangle Institute

  2. NIJ Project • Grant No. 2006-DN-BX-K014 • Opinions are those of the authors and not necessarily the U.S. Department of Justice

  3. Objective • Evaluate the Jeol Ltd. AccuTOF DART system as a novel application for use in postmortem toxicology laboratories • Qualitative • Screening methodology • Specifically postmortem • Urine • Blood and tissue

  4. Presentation goals • Equipment set up • Discuss standards run • Urine based testing • Blood/tissue testing • Summary of experience with the instrument • Future work

  5. RTI set up • JEOL TOF • DART • LEAP Technologies CombiPal autosampler • Mass Center software • More recently trying the Schraeder software

  6. Snorkel LEAP Autosampler Ion Source DART AccuTOF DART

  7. How Autosampler Works

  8. Sample Introduction

  9. AccuTOF- DART Strengths • Minimal sample preparation • Broad range of sensitivity • Rapid analysis • Simultaneous determination of multi-drug analytes • Sufficient mass accuracy for formula determination

  10. Reference materials • Examined 112 compounds • drugs and metabolites • Methanolic standard materials • Directly introduced

  11. Isomers

  12. Near Isomers

  13. Urine Method • Drug standards diluted in blank human urine • Glass probe dipped in sample • Analyzed with DART in positive mode • DART temperature @ 300°C • Mass calibrated using polyethylene glycol • Orifice 1 voltage set at 20V • Archived previously confirmed postmortem samples analyzed

  14. Urine Components Urea+H Creatinine Dimer +H Creatinine+H

  15. Cocaine 100 µg/mL M + H = 304.1548

  16. Methamphetamine 100 µg/mL M+H = 150.1278

  17. Triazolam 100 µg/mL M + H = 343.0508

  18. LOD for Drug Analytes in Urine (µg/mL)

  19. Creatinine Issues Oxazepam (10 µg/mL) in DI water (M+H 287.0578) Oxazepam (10 µg/mL) with 10 µg/mL creatinine

  20. Creatinine Issues Oxazepam (10 µg/mL) with 100 µg/mL creatinine Oxazepam (10 µg/mL) with 200 µg/mL creatinine

  21. Archived Case Example EDDP=278.190 278.20765 +17mmu 310.21327 Methadone=310.216 -3mmu

  22. Evaluation of Postmortem Blood and Tissue Samples

  23. “Minimal” Blood/Tissue Method • 2 mL of ACN added to 1 mL of sample • Mixture was shaken and vortexed • Spun in centrifuge at 2500 rpm for 5 minutes • ~100uL of ACN saved for DART analyses • Remainder poured off and dried down at 45°C • Reconstituted in 100 uL of ACN and analyzed

  24. Cases used

  25. Blood method • Acetonitrile added to blood

  26. Blood Method • Vortexed

  27. Blood method • Centrifuged

  28. Blood method • Decanted off supernatant

  29. Blood method • Dry down under N2

  30. Blood method • Reconstitute

  31. SPE method

  32. Human Blood Cholestadiene Monosaccharide Urea Monosaccharide Plasticizer

  33. 310.21838 Methadone 100 ug/mL Methadone in Blank Blood Unextracted M + H =310.216 100 ug/mL Methadone in Blank Blood Extracted in 1:3 ACN

  34. Methadone 100 ug/mL Methadone in Blank Blood Extracted in 1:3 ACN/dried/reconstituted in 100 ug/mL 1 ug/mL Methadone in Blank Blood Extracted in 1:3 ACN/dried/reconstituted in 100 ug/mL ACN 310. 215

  35. Postmortem Aorta Blood Specimen Un-extracted Previously reported at 2.8 mg/mL Nondetected Extracted in 1:3 ACN

  36. Postmortem Aorta Blood Specimen Extracted in 1:3 ACN/dried/reconstituted in 100 ug/mL ACN

  37. Postmortem Liver Specimen Un-extracted Previously reported at 6.6 ug/mL Extracted in 1:3 ACN

  38. Postmortem Liver Specimen Extracted in 1:3 ACN/dried/reconstituted in 100 ug/mL

  39. 304.15962 Cocaine 1 ug/mL Cocaine in blank blood extracted in 1:3 ACN M+H=304.154 1 ug/mL Cocaine in Blank Blood Extracted in 1:3 ACN/dried/reconstituted in 100 ug/mL ACN

  40. Cocaine 0.1 ug/mL Cocaine in Blank Blood Extracted in 1:3 ACN/dried/reconstituted in 100 ug/mL ACN

  41. 304.16913 Postmortem Aorta Blood Specimen Extracted in 1:3 ACN Previously reported at 0.52 mg/mL Extracted in 1:3 ACN/dried/reconstituted in 100 ug/mL ACN

  42. ? BE=290.138 Norpropoxyphene=326.211 Propoxyphene=340.226 Postmortem Cocaine Case • All drugs not found • Could not see without concentrating SPE Extraction

  43. Postmortem case by LC/MS and SPE BE BE-d3 cocaine cocaine-d3 cocaethylene cocaethylene-d3 BE 1253 ng/mL, Cocaine 8.8 ng/mL, CE 2.7 ng/mL

  44. Original case reports

  45. Original case reports

  46. Comparison to “Traditional” MS platforms * No quantitation attempted on the trap ** Source was saturated, resulting in poor quantitation

  47. Table of Drugs Analyzed in PM Specimens a a a a a a a a a a a a a a a a a a

  48. Conclusions • Reasonable sensitivity for some drugs • Greater sensitivity for some drugs over others • Creatinine appears to interfere with ionization of some drugs • Requirement for some sample preparation to mitigate interference from matrix • Auto sampler helps with more consistent sample introduction

  49. Conclusions • At least minimal drug extraction appears necessary to achieve best sensitivity • Extensive (solid phase extraction), does not necessarily improve things

  50. Conclusions • Drug detected in spiked blood have better sensitivity than in postmortem specimens at the same or greater concentrations • Not ideal for detecting therapeutic drug levels • Caveat the post mortem samples used are old and compound degradationmay have occurred

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