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The introduction of high-performance liquid chromatography (HPLC) to the analysis of peptides and proteins some 25 years ago revolutionized the biological sciences by enabling the rapid and sensitive analysis of peptide and protein structure through the exquisite speed, sensitivity, and resolution that can be easily obtained. Today, HPLC in its various modes has become the pivotal technique in the characterization of peptides and proteins and currently plays a critical role in both our understanding of biological processes and in the development of peptide- and protein-based pharmaceuticals. The number of applications of HPLC in peptide and protein purification continues to expand at an extremely rapid rate. Solid-phase peptide synthesis and recombinant DNA techniques have allowed the production of large quantities of peptides and proteins that need to be highly purified. HPLC techniques are also used extensively in the isolation and characterization of novel proteins that will become increasingly important in the postgenomic age. The design of multidimensional purification schemes to achieve high levels of product purity further demonstrates the power of HPLC techniques not only in the characterization of cellular events, but also in the production of pepti- and protein-based therapeutics. HPLC continues to be at the heart of the analytical techniques with which scientists in both academia and in industry must arm themselves to be able to fully characterize the identity, purity, and potency of peptides and proteins.
Why a Second Edition?The Second Edition provides practical answers to the general question, "How can I obtain useful sequence information from my protein or peptide?" rather than the more specific question asked in the first edition, "How can I obtain the N-terminal sequence?" Important new methods include ways of dealing with blocked N termini, computer analysis of protein sequences, and the recent revolution in mass spectrometry. - Mass spectrophotometric characterization of proteins and peptides - N-terminal sequencing of proteins with blocked N termini - Internal amino acid sequence analysis after protease digestion in-gel and on-blot - Improved microscale peptide purification methods - Computer analysis of protein sequences - New protocols tested and refined through everyday use in authors' laboratories - Updated reference chapter covering all aspects of protein microsequencing
This book is dedicated to the characterization of peptides and their applications for the study of biochemical systems. The contributing authors are all leaders in the field of peptide research. Part I, Characterization, presents the most recent advances in select analytical techniques. Part II, Application, presents a variety of specific applications for synthetic peptides. This book is an indispensable aid in the pursuit of new directions in peptide research.
How to synthesize native and modified proteins in the test tube With contributions from a panel of experts representing a range of disciplines, Total Chemical Synthesis of Proteins presents a carefully curated collection of synthetic approaches and strategies for the total synthesis of native and modified proteins. Comprehensive in scope, this important reference explores the three main chemoselective ligation methods for assembling unprotected peptide segments, including native chemical ligation (NCL). It includes information on synthetic strategies for the complex polypeptides that constitute glycoproteins, sulfoproteins, and membrane proteins, as well as their characterization. In addition, important areas of application for total protein synthesis are detailed, such as protein crystallography, protein engineering, and biomedical research. The authors also discuss the synthetic challenges that remain to be addressed. This unmatched resource: Contains valuable insights from the pioneers in the field of chemical protein synthesis Presents proven synthetic approaches for a range of protein families Explores key applications of precisely controlled protein synthesis, including novel diagnostics and therapeutics Written for organic chemists, biochemists, biotechnologists, and molecular biologists, Total Chemical Synthesis of Proteins provides key knowledge for everyone venturing into the burgeoning field of protein design and synthetic biology.
Revised and Expanded Handbook Provides Comprehensive Introduction and Complete Instruction for Sample Preparation in Vital Category of Bioanalysis Following in the footsteps of the previously published Handbook of LC-MS Bioanalysis, this book is a thorough and timely guide to all important sample preparation techniques used for quantitative Liquid Chromatography–Mass Spectrometry (LC-MS) bioanalysis of small and large molecules. LC-MS bioanalysis is a key element of pharmaceutical research and development, post-approval therapeutic drug monitoring, and many other studies used in human healthcare. While advances are continually being made in key aspects of LC-MS bioanalysis such as sensitivity and throughput, the value of research/study mentioned above is still heavily dependent on the availability of high-quality data, for which sample preparation plays the critical role. Thus, this text provides researchers in industry, academia, and regulatory agencies with detailed sample preparation techniques and step-by-step protocols on proper extraction of various analyte(s) of interest from biological samples for LC-MS quantification, in accordance with current health authority regulations and industry best practices. The three sections of the book with a total of 26 chapters cover topics that include: Current basic sample preparation techniques (e.g., protein precipitation, liquid-liquid extraction, solid-phase extraction, salting-out assisted liquid-liquid extraction, ultracentrifugation and ultrafiltration, microsampling, sample extraction via electromembranes) Sample preparation techniques for uncommon biological matrices (e.g., tissues, hair, skin, nails, bones, mononuclear cells, cerebrospinal fluid, aqueous humor) Crucial aspects of LC-MS bioanalytical method development (e.g., pre-analytical considerations, derivation strategies, stability, non-specific binding) in addition to sample preparation techniques for challenging molecules (e.g., lipids, peptides, proteins, oligonucleotides, antibody-drug conjugates) Sample Preparation in LC-MS Bioanalysis will prove a practical and highly valuable addition to the reference shelves of scientists and related professionals in a variety of fields, including pharmaceutical and biomedical research, mass spectrometry, and analytical chemistry, as well as practitioners in clinical pharmacology, toxicology, and therapeutic drug monitoring.
Preparative Chromatography for Separation of Proteins addresses a wide range of modeling, techniques, strategies, and case studies of industrial separation of proteins and peptides. • Covers broad aspects of preparative chromatography with a unique combination of academic and industrial perspectives • Presents Combines modeling with compliantce useing of Quality-by-Design (QbD) approaches including modeling • Features a variety of chromatographic case studies not readily accessible to the general public • Represents an essential reference resource for academic, industrial, and pharmaceutical researchers
HPLC for Pharmaceutical Scientists is an excellent book for both novice and experienced pharmaceutical chemists who regularly use HPLC as an analytical tool to solve challenging problems in the pharmaceutical industry. It provides a unified approach to HPLC with an equal and balanced treatment of the theory and practice of HPLC in the pharmaceutical industry. In-depth discussion of retention processes, modern HPLC separation theory, properties of stationary phases and columns are well blended with the practical aspects of fast and effective method development and method validation. Practical and pragmatic approaches and actual examples of effective development of selective and rugged HPLC methods from a physico-chemical point of view are provided. This book elucidates the role of HPLC throughout the entire drug development process from drug candidate inception to marketed drug product and gives detailed specifics of HPLC application in each stage of drug development. The latest advancements and trends in hyphenated and specialized HPLC techniques (LC-MS, LC-NMR, Preparative HPLC, High temperature HPLC, high pressure liquid chromatography) are also discussed.
For food scientists, high-performance liquid chromatography (HPLC) is a powerful tool for product composition testing and assuring product quality. Since the last edition of this volume was published, great strides have been made in HPLC analysis techniques-with particular attention given to miniaturization, automatization, and green chemistry. Tho
The use of computers in numerical characterization of molecular structures has given chemists fundamentally new information on chemical structures, leading to major developments in physical, analytical, and medicinal chemistry. This book, written by a pioneer in the field, extends and updates research on quantitative structure retention relationships (QSRR) by consolidating and critically reviewing the extensive literature on the subject while providing basic theoretical and practical information required in all investigations involving chromatography, analytical chemistry, biochemistry, and pharmaceutical research. Coverage includes detailed discussions of the general theories and mechanisms of chromatographic separations, prediction of retention coefficients, statistical techniques and formal requirements of QSRR studies, specific applications of chromatographic data, and much more. Also provides several carefully selected figures and tables plus extensive bibliographies.
Leading practitioners authoritatively describe the newest and most effective spectrometric techniques for the analysis of proteins and peptides. The areas covered range from the elucidation of primary and secondary protein structure and the rapid identification of proteins using database techniques to methods for sequencing, as well as methods for the quantitative determination of peptides. Other chapters provide detailed information on the analysis of glycoproteins and glycopeptides and on the use of mass spectrometry to probe the interactions of proteins, both covalent and noncovalent.