Advantages and Disadvantages of High-end Mass Spectrometry in a Forensic Toxicology Lab

Transcription

1 Advantages and Disadvantages of High-end Mass Spectrometry in a Forensic Toxicology Lab Raymond Van Orden Forensic Scientist Supervisor Controlled Substances and Toxicology

2 Objectives 1) Discuss advantages/disadvantages of each mass spectrometry system GC/MS LC/MS Tandem MS 1) Best approach to implementing tandem MS systems

3 What kind of samples are you testing? Makes a difference to what kind of system you want to use 1) Blood 2) Urine 3) Post mortem or Anti mortem 4) Other matrices?

4 Goals of Mass Spec Quantitative or Qualitative Quantitative Calibration curve Confirmation Retention time Ion Ratio Quant/Qual ion coelution Qualitative Retention Time Spectral Match (ion trap or scan) Product ion ratios

5 Factors affecting Mass Spec Performance Sample concentration Matrix Analyte type Buffers and purity Purity of Organic solvents Purity of curtain gas and collision cell gas Run time Number of samples run

6 Single Quad GC/MS Higher concentration samples Usually involves derivitization depending on analyte SIM Electron impact ionization (EI) or chemical ionization (CI) Most common in labs Get fingerprint identification, good for library matches of unknowns when in scan mode

7 Single Quad LC/MS Tends to have better sensitivity than GC No derivitization needed Have to over come the fear of LC Uses collision induced dissociation softer technique not as many ions Have to be aware of adducts Not good for library matches, gives you the molecular ion in most cases.

8 Collision Induced Dissociation CID spectra affected most by: Type and pressure of collision gas Initial kinetic energy of the ions Structure of the ions Instrument configuration Charge site Factors that influence CID spectra: Collision energy Collision gas choice and pressure CID time

9 Collision Induced Dissociation Most Common Proton acceptors R NH 2 R NH R R 2 PH, R 2 PH 3 R OH, R - SH R OR, R C = O Hydrocarbons/Aromatics Better + Ion formation Worse + Ion formation

10 Collision Induced Dissociation Atmospheric Pressure ionization processes most often produce even electron ions The most favorable losses will be neutral fragments (stable molecules) and production of another even electron ion Water (H 2 O) 18 Daltons, is a common neutral loss Try to avoid using losses of 18 Uncommon to lose a mass of 4-14 Daltons and Daltons

11 Tandem MS Used with either GC or LC Removes matrix interferences Provides reliable confirmation Allows for the selective quantitation of target compounds in high background samples Better S/N in complex matrices than can be achieved by single quad scan and SIM approaches

12 Scan MRM

13 Who should use QQQ (MS/MS)? User doing Selected Ion Monitoring(SIM) for target compound analysis in laboratories, needing additional sensitivity and selectivity with less sample prep, to meet more demanding analytical requirements.

14 Why Tandem MS LC or GC-MS/MS: Routine Targeted trace analysis in complex matrix MRM Sensitivity unsurpassed (up to a few hundred compounds) Complex matrix with less clean-up (within reason)

15 Getting started Maximize separation (LC/GC first) Can use existing LC or GC methods Good chromatography assists with higher sensitivity and confidence

16 For LC beginners Getting started Choose a good buffer in appropriate ph range of your compounds Select an appropriate column Usually use a gradient technique Mixture of MEOH and buffer Make sure buffers are filtered!

17 Pheniramine Chromatograph of Analyzed Compounds Quetiapine CE Cocaine PCP TFMPP Methylphenidate Propylhexadrine Ketamine Norketamine MDEA BE DMS D8-BE Dehydro MDMA Methcathinone Pseudoephedrine Ephedrine Diphenhyd D3-Coc D10- Amp D5- MDMA Phentermine Methamp MDA D11-Meth Amphetamine BZP Cathinone Cathine PPA EME D3-EME x

18 LC can provide: Baseline separation of stereoisomer's PPA (Norephedrine) and Cathine (Norpsuedoephedrine) Ephedrine and Pseudoephedrine Which makes identification and quantification simple Pseudoephedrine Ephedrine Cathine PPA

19 Specific Hints Filter, Filter, Filter All buffers Good Sample prep SPE, filter samples Guard Column or Frits ( crud catchers ) Garbage in garbage out! Takes time, but optimize each analyte Monitor pressures Column, nebulizer, collision cell Monitor Vacuum Monitor Log book

20 Getting started Optimize Mass spec Tune (manual or auto?) Spray chamber Drying gas temp. and flow Nebulizer Capillary voltage Quad 1 Capillary voltage Fragmentor voltage Collision Cell Collision energy Quad 3 (grouped or dynamic) Maximize daughter ions EMV

21 Third Quad Optimization Group MRM

22 Third Quad Optimization Dynamic MRM

23 Third Quad Optimization Dynamic MRM

24 Noted Problems Changing of fragmentor from 80 (Meth) to 100 (Phentermine) half way through created a split in the peak Fragmentor values changed for Phentermine, MDA, MDMA, and D5-MDMA from 100 to 80 and changed the Meth and Phentermine RT window from 1 min to 2 min

25 Noted Problems Improper setting of retention time for methamphetamine and phentermine, daughter ion caused a jump in the 91 ion half way through the peak

26 Benefits of LC/GC tandem MS (LC) Eliminates the need for derivatization Can separate and ID similar compounds Superior sensitivity and selectivity of analytes When operating in MRM mode Quantification of low levels of compounds in biological matrices Wide dynamic range

27 Summary 1) Each system has advantages and disadvantages, but combining them makes the laboratory more efficient and productive

28 Questions?