HPLC for Pharmaceutical Scientists 2007 (Part 13)

Recent advances in mass spectrometry have rendered it an attractive and versatile tool in industrial and academic research laboratories. As a part of this rapid growth, a considerable body of literature has been devoted to the application of mass spectrometry in clinical studies. In concert with separation techniques such as liquid chromatography, mass spectrometry allows the rapid characterization and quantitative determination of a large array of molecules in complex mixtures. Herein, we present an overview of the above techniques accompanied with several examples of the use of liquid chromatography– tandem mass spectrometry in pharmacokinetics/drug metabolism assessment during drug...
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HPLC for Pharmaceutical Scientists 2007 (Part 13)13THE ROLE OF LIQUIDCHROMATOGRAPHY–MASSSPECTROMETRY INPHARMACOKINETICS ANDDRUG METABOLISMRay Bakhtiar, Tapan K. Majumdar, and Francis L. S. Tse13.1 INTRODUCTIONRecent advances in mass spectrometry have rendered it an attractive and ver-satile tool in industrial and academic research laboratories. As a part of thisrapid growth, a considerable body of literature has been devoted to the appli-cation of mass spectrometry in clinical studies. In concert with separation tech-niques such as liquid chromatography, mass spectrometry allows the rapidcharacterization and quantitative determination of a large array of moleculesin complex mixtures. Herein, we present an overview of the above techniquesaccompanied with several examples of the use of liquid chromatography–tandem mass spectrometry in pharmacokinetics/drug metabolism assessmentduring drug development. Since the evolution of pharmaceutical research [1, 2], the stages of drug dis-covery and development have followed three predominant patterns: (i) thesystematic and methodical approach by chemists to rationally design and syn-thesize a molecule to target a specific molecular system (e.g., ion channels,receptors, enzymes, DNA); (ii) the isolation and purification of the activeingredients of medicinal plants or microorganisms to screen their spectrum ofHPLC for Pharmaceutical Scientists, Edited by Yuri Kazakevich and Rosario LoBruttoCopyright © 2007 by John Wiley & Sons, Inc. 605606 THE ROLE OF LC–MS IN PK AND DRUG METABOLISMactivity using in vitro models; or (iii) the serendipitous discovery of a com-pound with a novel pharmacological action (e.g., the accidental discovery ofantidepressants). Today, one of the increasingly popular and complementaryapproaches for drug discovery in the pharmaceutical industry is to performmassive parallel synthesis in solution or on a solid support. In addition, withthe advent of functional genomics and proteomics, cell-based assays, and mol-ecular biology, a multitude of therapeutic targets have been validated [3]. With an increasing number of potential molecular targets identified throughthe science of functional proteomics and genomics, diverse libraries of newchemical entities (NCEs) have to be generated and evaluated. Consequently,the rapid growth of combinatorial libraries has posed a need for faster,accurate, and sensitive analytical techniques capable of large-scale high-throughput screening (HTS). Although in vitro assays do not necessarilyreflect the complexity of the in vivo interactions, the speed and simplicity ofthe former have rendered them an integral part of the screening process. In recent years, the in silico and experimental modeling of pharmacokinetic/pharmacodynamic (PK/PD) relationship have become increasingly popular[4, 5]. The integration of PK (i.e., drug dose and biological fluid concentration)and PD (i.e., pharmacologic effect) provides a key determinant in under-standing the dosing regimen and therapeutic effect of a potential drug com-pound. To this end, analytical assays also play a pivotal role in defining thePK/PD relation of NCEs. In many cases, both the drug concentration and PDbiomarkers (vide infra) can be directly measured in peripheral fluids usingspecific analytical techniques. Furthermore, samples generated from large-scale clinical trials along withthe ambitious development timelines to get safe and efficacious drugs tomarket warrant the use of HT bioanalysis. Numerous improvements in speed,sensitivity, and accuracy, augmented with innovations in automation in con-junction with mass spectrometry (MS) detection, have allowed for versatileand multifaceted platforms [6–8].13.2 IONIZATION PROCESSESMass spectrometry (MS) is playing an increasingly visible role in the molecu-lar characterization of combinatorial libraries, natural products, drug meta-bolism and pharmacokinetics, toxicology and forensic investigations, andproteomics. Toward this end, electrospray ionization (ESI), atmospheric pres-sure chemical ionization (APCI), and atmospheric pressure photo-ionization(APPI) have proven valuable for both qualitative and quantitative screeningof small molecules (e.g., pharmaceutical products) [9–14]. The utility of ESI (Figure 13-1) lies in its ability to generate ions directlyfrom the solution phase into the gas phase. The ions are produced by appli-cation of a strong electric field to a very fine spray of the solution containingthe analyte. The electric field creates highly charged droplets whose subse-IONIZATION PROCESSES 607 Figure 13-1. A simplified schematic of the ESI process. (Courtesy of Dr. P. Tiller.)Figure 13-2. A simplified schematic of t ...