基礎講座 電気泳動(Electrophoresis)

in Mass Spectrometry-Based Metabolomics and Lipidomics Tomas Cajka† and Oliver Fiehn*,†,‡ †UC Davis Genome Center−Metabolomics, University of California Davis, 451 Health Sciences Drive, Davis, California 95616, United States ‡King Abdulaziz University, Faculty of Science, Biochemistry Department, P.O. Box 80203, Jeddah 21589, Saudi Arabia ■ CONTENTS Sample Extraction 525 Extraction of Polar Metabolites (Metabolomics) 525 Extraction of Lipids (Lipidomics) 527 Combined Extraction of Amphiphilic and Lipophilic Metabolites 527 Mass Spectrometry-Based Metabolomics and Lipidomics 528 Direct Infusion MS 528 Ion Mobility-Mass Spectrometry (IM-MS) 529 Liquid Chromatography−Mass Spectrometry (LC−MS) 533 Reversed-Phase Liquid Chromatography (RPLC) 533 Hydrophilic Interaction Chromatography (HILIC) 534 Normal-Phase Liquid Chromatography (NPLC) 535 Supercritical Fluid Chromatography (SFC) 535 Two-Dimensional Liquid Chromatography (2D-LC) 535 Mass Spectrometric Detection 536 Data Processing 540 Quality Control 541 Conclusions 541 Author Information 542 Corresponding Author 542 Notes 542 Biographies 542 Acknowledgments 542 References 542

Toward Merging Untargeted and Targeted Methods in Mass Spectrometry-Based Metabolomics and Lipidomics.

There has been a great interest in application of nanoparticles as biomaterials for delivery of therapeutic molecules such as drugs and genes, and for tissue engineering. In particular, biopolymers are suitable materials as nanoparticles for clinical application due to their versatile traits, including biocompatibility, biodegradability and low immunogenicity. Biopolymers are polymers that are produced from living organisms, which are classified in three groups: polysaccharides, proteins and nucleic acids. It is important to control particle size, charge, morphology of surface and release rate of loaded molecules to use biopolymer-based nanoparticles as drug/gene delivery carriers. To obtain a nano-carrier for therapeutic purposes, a variety of materials and preparation process has been attempted. This review focuses on fabrication of biocompatible nanoparticles consisting of biopolymers such as protein (silk, collagen, gelatin, β-casein, zein and albumin), protein-mimicked polypeptides and polysaccharides (chitosan, alginate, pullulan, starch and heparin). The effects of the nature of the materials and the fabrication process on the characteristics of the nanoparticles are described. In addition, their application as delivery carriers of therapeutic drugs and genes and biomaterials for tissue engineering are also reviewed.

https://www.mdpi.com/1422-0067/14/1/1629/pdf

Biopolymer-based nanoparticles for drug/gene delivery and tissue engineering.

Characterization of Therapeutic Antibodies and Related Products

Serum albumin is a multi-functional protein that is able to bind and transport numerous endogenous and exogenous compounds. The development of albumin drug carriers is gaining increasing importance in the targeted delivery of cancer therapy, particularly as a result of the market approval of the paclitaxel-loaded albumin nanoparticle, Abraxane®. Considering this, there is renewed interest in isolating and characterizing albumin-binding proteins or receptors on the plasma membrane that are responsible for albumin uptake. Initially, the cellular uptake and intracellular localization of albumin was unknown due to the large confinement of the protein within the vascular and interstitial compartment of the body. Studies have since assessed the intracellular localization of albumin in order to understand the mechanisms and pathways responsible for its uptake, distribution and catabolism in multiple tissues, and this is reviewed herein.

/pdf/unraveling-the-mysteries-of-serum-albumin-more-than-just-a-1gtrw4vbn9.pdf

Unraveling the mysteries of serum albumin—more than just a serum protein

The extent of drug binding to plasma proteins, determined by measuring the free active fraction, has a significant effect on the pharmacokinetics and pharmacodynamics of a drug. It is therefore highly important to estimate drug-binding ability to these macromolecules in the early stages of drug discovery and in clinical practice. Traditionally, equilibrium dialysis is used, and is presented as the reference method, but it suffers from many drawbacks. In an attempt to circumvent these, a vast array of different methods has been developed. This review focuses on the most important approaches used to characterize drug-protein binding. A description of the principle of each method with its inherent strengths and weaknesses is outlined. The binding affinity ranges, information accessibility, material consumption, and throughput are compared for each method. Finally, a discussion is included to help users choose the most suitable approach from among the wealth of methods presented.

/pdf/drug-protein-binding-a-critical-review-of-analytical-tools-2700fqrr8s.pdf

Drug-protein binding: a critical review of analytical tools.

In recent years, different approaches have been taken to improve chromatographic performance in terms of analysis time and/or resolution. The use of columns packed with sub-2μm particles in ultra-high-pressure liquid chromatography (UHPLC) has become a technique of choice in many laboratories. Furthermore, for the analysis of complex matrices (e.g., biological fluids, plant extracts, and food and environmental samples), coupling UHPLC with mass spectrometry (MS) or tandem MS (MS 2 ) provides a powerful analytical tool. This review describes major advances in the field of UHPLC-MS and UHPLC-MS 2 . We strongly emphasize the possibility of speeding up bioanalysis, drug metabolism, and multi-residue screening assays, while maintaining qualitative and quantitative performance equivalent to HPLC-MS and HPLC-MS 2 . We also report the possibility of gaining additional information in metabolomics, using high-resolution UHPLC with a time-of-flight analyzer. The studies summarized are discussed in this review in terms of throughput increases and resolution enhancements afforded by UHPLC. In addition, we highlight the impact of UHPLC conditions on MS detection capabilities (e.g., acquisition rate, limits of detection and matrix effects).

Coupling ultra-high-pressure liquid chromatography with mass spectrometry

Intact protein analysis in the biopharmaceutical field

Since its commercial introduction in 2004, there has been a considerable interest in ultra-high-performance (pressure) liquid chromatography (UHPLC), which dramatically increases the throughput of regular HPLC methods. Although the ability to achieve fast separations and good resolution are the main drivers for the increasing use of UHPLC, we describe a number of other trends in this review, such as: 1 use of UHPLC technology to perform high-resolution analysis of complex samples; 2 development of columns packed with sub-2-µm particles to achieve different chromatographic modes (i.e. chiral LC, SEC, IEX, HILIC, and SFC); 3 evaluation of higher pressure drop, higher temperature, smaller particle sizes and sub-2-µm core-shell particles, to further improve kinetic performance; 4 use of UHPLC for enhancing the characterization of biopharmaceuticals; and, 5 development of UHPLC-MS for applications, including bioanalysis, multi-residue screening, and metabolomics.

Current and future trends in UHPLC

Sample preparation is a crucial step in any bioanalytical workflow. Nowadays, microextractions are very popular because of the reduction of the organic solvent and sample volumes, the analysis time, and the operating costs. Electromembrane extraction (EME) is one of the most recently reported liquid phase microextraction (LPME) techniques and was first described in 2006. It is based on the application of an electric field between two aqueous compartments separated by a layer of water-immiscible organic solvent, reducing the time needed for the extraction process to a few minutes, enabling extraction selectivity towards a broad range of molecule polarity (−5

Julie Schappler

Papers

Drug-protein binding: a critical review of analytical tools.

Coupling ultra-high-pressure liquid chromatography with mass spectrometry

Intact protein analysis in the biopharmaceutical field

Current and future trends in UHPLC

Electromembrane extraction: Overview of the last decade