Separation of polar compounds on polar stationary phases with partly aqueous eluents is by no means a new separation mode in LC. The first HPLC applications were published more than 30 years ago, and were for a long time mostly confined to carbohydrate analysis. In the early 1990s new phases started to emerge, and the practice was given a name, hydrophilic interaction chromatography (HILIC). Although the use of this separation mode has been relatively limited, we have seen a sudden increase in popularity over the last few years, promoted by the need to analyze polar compounds in increasingly complex mixtures. Another reason for the increase in popularity is the widespread use of MS coupled to LC. The partly aqueous eluents high in ACN with a limited need of adding salt is almost ideal for ESI. The applications now encompass most categories of polar compounds, charged as well as uncharged, although HILIC is particularly well suited for solutes lacking charge where coulombic interactions cannot be used to mediate retention. The review attempts to summarize the ongoing discussion on the separation mechanism and gives an overview of the stationary phases used and the applications addressed with this separation mode in LC.

Hydrophilic interaction chromatography.

Recent advances in high-throughput quantitative bioanalysis by LC-MS/MS.

This paper reviews the recent developments in bioanalysis sample preparation techniques and gives an update on basic principles, theory, applications and possibilities for automation, and a comparative discussion on the advantages and limitation of each technique. Conventional liquid-liquid extraction (LLE), protein precipitation (PP) and solid-phase extraction (SPE) techniques are now been considered as methods of the past. The last decade has witnessed a rapid development of novel sample preparation techniques in bioanalysis. Developments in SPE techniques such as selective sorbents and in the overall approach to SPE, such as hybrid SPE and molecularly imprinted polymer SPE, have been addressed. Considerable literature has been published in the area of solid-phase micro-extraction and its different versions, e.g. stir bar sorptive extraction, and their application in the development of selective and sensitive bioanalytical methods. Techniques such as dispersive solid-phase extraction, disposable pipette extraction and micro-extraction by packed sorbent offer a variety of extraction phases and provide unique advantages to bioanalytical methods. On-line SPE utilizing column-switching techniques is rapidly gaining acceptance in bioanalytical applications. PP sample preparation techniques such as PP filter plates/tubes offer many advantages like removal of phospholipids and proteins in plasma/serum. Newer approaches to conventional LLE techniques (salting-out LLE) are also covered in this review article.

Recent advances in sample preparation techniques for effective bioanalytical methods

The analysis of highly hydrophilic, ionic, and polar compounds has been performed by HILIC-ESI-MS for the last few years. The use of low aqueous/high polar organic solvent content in HILIC separation mobile phase is almost ideal for ESI-MS detection in many cases, resulting in increased sensitivity. Although the addition of modifiers such as acids or salts is necessary in some circumstances for a good separation, the optimum concentrations used are still highly amenable for ESI-MS analysis, showing few deleterious effects. In this review, the mechanism of HILIC separation and ESI ion generation will be briefly discussed, followed by a summary of method development and applications in several fields of research including pharmaceutical, biomolecular, food, metabolic, and environmental analysis.

The advantages of ESI-MS detection in conjunction with HILIC mode separations: Fundamentals and applications

Liquid chromatography-mass spectrometry/mass spectrometry method development for drug metabolism studies: Examining lipid matrix ionization effects in plasma.

Swift growth in the use of LC-MS/MS for the analysis of drugs in biological matrices has been compelled by the need for timely and high-quality data at many stages in drug discovery and development process: from high throughput screening of drug candidates and rapid data generation for pre-clinical studies to almost 'real-time' analysis of clinical samples. Prompt and rational method development, validation, and transfer play a pivotal role in achieving the goals of "faster, better, and cheaper" for pharmacokinetic studies since this could easily account for more than 50% of the time and labor resources for a moderate-sized project. Strategy for rational method development, validation and transfer has been largely kept as institutional knowledge but rarely appeared in literature. In this review article, strategies for developing and validating robust high throughput LC-MS/MS methods will be critically reviewed and discussed. Automated sample preparation, fast chromatography, minimization of matrix effects, and strategy of narrowing the gap between validation and incurred sample analysis are just a few topics covered in this review. Other interesting approaches for improving method efficiency and ruggedness such as direct injection SPE and liquid/liquid extracts as well as multiplexing of LC columns will also be discussed. Potential pitfalls during method development and validation are pointed out. At the end, the question "how fast is fast enough and how fast is too fast?" will be answered after considering all aspects of the method development and validation.

Critical Review of Development, Validation, and Transfer for High Throughput Bioanalytical LC-MS/MS Methods

Automated 96-well solid phase extraction and hydrophilic interaction liquid chromatography-tandem mass spectrometric method for the analysis of cetirizine (ZYRTEC) in human plasma--with emphasis on method ruggedness.

Yong Tang

Papers

Critical Review of Development, Validation, and Transfer for High Throughput Bioanalytical LC-MS/MS Methods

Automated 96-well solid phase extraction and hydrophilic interaction liquid chromatography-tandem mass spectrometric method for the analysis of cetirizine (ZYRTEC) in human plasma--with emphasis on method ruggedness.