A review of current trends and advances in modern bio-analytical methods: chromatography and sample preparation.

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

This article discusses dried blood spot (DBS) sampling in therapeutic drug monitoring (TDM). The most important advantages of DBS sampling in TDM are the minimally invasive procedure of a finger prick (home sampling), the small volume (children), and the stability of the analyte. Many assays in DBS have been reported in the literature over the previous 5 years. These assays and their analytical techniques are reviewed here. Factors that may influence the accuracy and reproducibility of DBS methods are also discussed. Important issues are the correlation with plasma/serum concentrations and the influence of hematocrit on spot size and recovery. The different substrate materials are considered. DBS sampling can be a valid alternative to conventional venous sampling. However, patient correlation studies are indispensable to prove this. Promising developments are dried plasma spots using membrane and hematocrit correction using the potassium concentration.

https://link.springer.com/content/pdf/10.1007%2Fs40262-014-0177-7.pdf

Therapeutic Drug Monitoring by Dried Blood Spot: Progress to Date and Future Directions

Liquid chromatography-mass spectrometry analysis of small molecules from biofluids requires sensitive and robust assays. Because of the very complex nature of many biological samples, efficient sample preparation protocols to remove unwanted components and to selectively extract the compounds of interest are an essential part of almost every bioanalytical workflow. This review describes the most common problems encountered during sample preparation, ways to optimize established sample preparation techniques and important recent developments to reduce or eliminate major interferents from biofluids.

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Recent advances in sample preparation techniques to overcome difficulties encountered during quantitative analysis of small molecules from biofluids using LC-MS/MS

Dried blood spot (DBS) sampling for quantitative determination of drugs in blood has entered the bioanalytical arena at a fast pace during the last decade, primarily owing to progress in analytical instrumentation. Despite the many advantages associated with this new sampling strategy, several issues remain, of which the hematocrit issue is undoubtedly the most widely discussed challenge, since strongly deviating hematocrit values may significantly impact DBS-based quantitation. In this review, an overview is given of the different aspects of the 'hematocrit problem' in quantitative DBS analysis. The different strategies that try to cope with this problem are discussed, along with their potential and limitations. Implementation of some of these strategies in practice may help to overcome this important hurdle in DBS assays, further allowing DBS to become an established part of routine quantitative bioanalysis.

Hemato-critical issues in quantitative analysis of dried blood spots: challenges and solutions

Acetonitrile, an organic solvent miscible with aqueous phase, has seen thousands of publications in the literature as an efficient deproteinization reagent. The use of acetonitrile for liquid-liquid extraction (LLE), however, has seen very limited application due to its miscibility with aqueous phase. The interest in LLE with acetonitrile has been pursued and reported in the literature by significantly lowering the temperature of the mixture or increasing the salt concentration in the mixture of acetonitrile and aqueous phase, resulting in the separation of the acetonitrile phase from aqueous phase, as observed in conventional LLE. However, very limited application of these methods has been reported. The throughput was limited. In this report, we report a new sample preparation technique, salting-out assisted liquid-liquid extraction with acetonitrile, for high-throughput good laboratory practice sample analysis using LCMS, Two compounds from an approved drug, Kaletra, were used to demonstrate the extractability of drugs from human plasma matrix. Magnesium sulfate was used as the salting-out reagent. Extracts were diluted and then injected into a reversed phase LC-MS/MS system directly. One 96-well plate was extracted with this new approach to evaluate multiple parameters of a good laboratory practice analytical method. Results indicate that the method is rapid, reliable and suitable for regulated bioanalysis. With minimal modification, this approach has been used for high-throughput good laboratory practice analysis of a number of compounds under development at Abbott.

Salting‐out assisted liquid/liquid extraction with acetonitrile: a new high throughput sample preparation technique for good laboratory practice bioanalysis using liquid chromatography–mass spectrometry

Simultaneous determination of a hydrophobic drug candidate and its metabolite in human plasma with salting-out assisted liquid/liquid extraction using a mass spectrometry friendly salt.

High-throughput salting-out assisted liquid/liquid extraction with acetonitrile for the simultaneous determination of simvastatin and simvastatin acid in human plasma with liquid chromatography.

Rapid, simultaneous determination of lopinavir and ritonavir in human plasma by stacking protein precipitations and salting-out assisted liquid/liquid extraction, and ultrafast LC–MS/MS

Huaiqin Wu

Papers

Salting‐out assisted liquid/liquid extraction with acetonitrile: a new high throughput sample preparation technique for good laboratory practice bioanalysis using liquid chromatography–mass spectrometry

Simultaneous determination of a hydrophobic drug candidate and its metabolite in human plasma with salting-out assisted liquid/liquid extraction using a mass spectrometry friendly salt.

High-throughput salting-out assisted liquid/liquid extraction with acetonitrile for the simultaneous determination of simvastatin and simvastatin acid in human plasma with liquid chromatography.

Rapid, simultaneous determination of lopinavir and ritonavir in human plasma by stacking protein precipitations and salting-out assisted liquid/liquid extraction, and ultrafast LC–MS/MS

Sugaring-out assisted liquid/liquid extraction with acetonitrile for bioanalysis using liquid chromatography–mass spectrometry