Short Courses

Monday October 17

• Chromatographic (HPLC and SFC) and Electrophoretic (CE) Method Development for
  LC/SFC/CE-MS
  
  

  This practical course on chromatographic and electrophoretic method development for LC/SFC/CE-MS covers the specifics of how separation methods are developed for interfacing to ESI- and APCI-MS. The course discusses the importance of separation methods when interfaced with mass spectrometry (MS) including the separation of undetected matrix components, isomers and labile metabolites. The development of separation methods coupled with MS for use in qualitative and quantitative analyses throughout drug discovery and development are presented. The most advantageous column chemistry for each application are thoroughly discussed including, reverse phase, normal phase, ion-exchange, HILIC, monolithic and polar embedded phases. This course also aims to broaden the analyst’s scope by offering introductions of other increasingly utilized separation techniques than HPLC, e.g. supercritical fluid chromatography (SFC) and capillary electrophoresis (CE), with emphasis on their hyphenation to MS and also their application in modern drug discovery and development. In addition to that, some recent advances in achieving higher selectivity, efficiency and speed by adopting unconventional methodologies such as ultrahigh pressure LC (UPLC), high temperature LC (HTLC), multidimensional separation and LC-on-chip, are elaborated as well. Step-wise method development tutorials of developing methods for specific compound classes based on the structure are included in the final summary.
  
  Instructors:  Shane Needham, Alturas Analytics Inc;  Yining Zhao, Pfizer Inc



• Peptide Sequencing & Protein Identification by µLC/MS/MS Ion Trap and Fourier Transform Mass Spectrometry
  
  

  This course is a distillation of experiences related to the use of ion trap mass spectrometers for the low level sequence analyses of peptides and proteins. It covers both fundamental principles and useful practices that will facilitate the use and application of µLC/MS/MS. Easy-to-follow mini-labs give attendees hands-on experience in µ-column construction and nano-spray source optimization. The analytical benefits of nanospray will be discussed, and keys to success for robust µLC and nanospray will be emphasized. Example applications direct participants through a step-by-step process of analyzing digested proteins, reviewing the data, and interpreting the results. The target audience includes chemical and biological scientists who want to learn how to begin identifying peptides and proteins using µLC/MS/MS ion-trap mass spectrometry. Some MS experience is recommended because basic LCMS and LCMSMS techniques are not directly covered. Examples will be based on the Thermo LCQ, LTQ, LTQ-FT ion traps.
  
  Outline:
  Ion Trap Mass Spectrometry: key concepts of operation
  Moving from microspray to nanospray on commercial instruments
  Analytical benefits of nanospray and µLC
  Micro-capillary HPLC: pumps, autosamplers, and column construction techniques (mini-lab)
  Nanospray source tuning (mini-lab)
  Performing µLC/MS/MS on the ThermoFinnigan LCQ (step-by-step example)
  Interpretation of LCMSMS data and sequence elucidation methods
  Making use of advanced instrumentation: LTQ and LTQ-FT
  
  Instructors:  Nathan A. Yates, Merck Research Laboratories;  Gary A. Valaskovic, New Objective



• Metabolic Profiling: Principles and Practice
  
  

  The profiles of biological fluids contain a vast array of endogenous low-molecular weight metabolites, the composition of which depends upon the sample type (plasma urine, bile etc) and factors such as the species, age, sex, diet of the organism from which the sample derives and indeed even the time of day at which the sample was taken. Disease, drugs (and other biologically active molecules) perturb concentrations and fluxes in intermediary metabolic pathways. The response to this perturbation involves adjustment of intracellular and extracellular environments in order to maintain homeostasis. Both perturbation and adjustment are expressed as changes in the normal patterns in biofluids or tissues that are characteristic (a ‘fingerprint’) of the nature or site of the disease process, toxic insult, pharmacological response or genetic modification. These patterns can be evaluated using analytical techniques combined with multivariate statistical methods to obtain insight into the response of a biological system to these perturbations in a time-related manner. To understand and interpret the data from these metabolic profiles it is vitally important to identify and characterize the metabolites both known and unknown when they are observed. This intent of this workshop is to describe methods for conducting metabolic profiling studies and to illustrate how the metabolites of both xenobiotic and various structurally endogenous metabolites can be identified using modern analytical techniques either alone or in tandem with hyphenated methods.
  
  Instructors:  John P. Shockcor, Waters Corporation;  Andrew Nicholls, GlaxoSmithKline