Direct analysis of dried blood spots coupled with mass spectrometry: concepts and biomedical applications

Because of the emergence of dried blood spots (DBS) as an attractive alternative to conventional venous plasma sampling in many pharmaceutical companies and clinical laboratories, different analytical approaches have been developed to enable automated handling of DBS samples without any pretreatment. Associated with selective and sensitive MS–MS detection, these procedures give good results in the rapid identification and quantification of drugs (generally less than 3 min total run time), which is desirable because of the high throughput requirements of analytical laboratories. The objective of this review is to describe the analytical concepts of current direct DBS techniques and to present their advantages and disadvantages, with particular focus on automation capacity and commercial availability. Finally, an overview of the different biomedical applications in which these concepts could be of major interest will be presented.     

Amantadine hydrochloride monitoring by dried plasma spot technique: High‐performance liquid chromatography–tandem mass spectrometry based clinical assay

Amantadine plasma concentrations correlate well with desired therapeutic effects and adverse outcomes; information on amantadine exposure could be useful when multiple amantadine clearance pathways are impaired or non‐compliance is suspected. Micro‐sampling strategies, like dried plasma spot, would be particularly useful because ambulatory patients that do not attend a clinic can easily sample a few drops of blood by themselves at the required time of the dosing interval. We developed and validated a dried‐plasma‐spot‐based high performance liquid chromatography–tandem mass spectrometry assay to quantify amantadine. This assay met relevant validation requirements within a hematocrit range of 20–50% and was linear from 100 to 2000 ng/mL. Amantadine was stable in dried plasma spots for up to 21 days at room temperature, regardless of whether the dried plasma spot was protected from light or not. The correlation between paired dried and wet plasma concentrations was assessed in 52 patients. Deming regression coefficients between wet plasma and simultaneously pipetted dried plasma spots were used to predict plasma concentrations. Bland–Altman plots revealed a strong agreement between dried and wet plasma concentrations, supporting the clinical usefulness of dried plasma spots for amantadine monitoring with a self‐sampling strategy at a convenient time and place for the patient.     

Dried blood spot analysis with liquid chromatography and mass spectrometry: Trends in clinical chemistry

Dried blood spot samples are simple to prepare and transport, enabling safe and accessible diagnostics, both locally and globally. We review dried blood spot samples for clinical analysis, focusing on liquid chromatography-mass spectrometry as a versatile measurement tool for these samples. Dried blood spot samples can provide information for, for example, metabolomics, xenobiotic analysis, and proteomics. Targeted analyses of small molecules are the main application of dried blood spot samples and liquid chromatography-mass spectrometry, but emerging applications include untargeted metabolomics and proteomics. Applications are highly varied, including analyses related to newborn screening, diagnostics and monitoring of disease progression and treatment effects of virtually any disease, as well as studies into the physiology and effects of diet, exercise, xenobiotics, and doping. A range of dried blood spot products and methods are available, and applied liquid chromatography-mass spectrometry instrumentation is varied with regard to liquid chromatography column formats and selectivity. In addition, novel approaches such as on-paper sample preparation (e.g., selective trapping of analytes with paper-immobilized antibodies) are described. We focus on research papers published in the last 5 years.     

Quantitative analysis of clofazimine (Lamprene®), an antileprosy agent,in human dried blood spots using liquid chromatography–tandem mass spectrometry

An LC–MS/MS method was developed and validated for bioanalysis of clofazimine in human dried blood spot (DBS) samples in support of a clinical study on multidrug‐resistant tuberculosis in developing countries. The validated assay dynamic range was from 10.0 to 2000 ng/mL using a 1/8 inch DBS punch. The accuracy and precision of the assay were ±11.0% (bias) and ≤13.5% (CV) for the LLOQs (10.0 ng/mL) and ±15% (bias) and ≤15% (CV) for all other QC levels. The assay was proved to be free from the possible impact owing to spot size and storage temperature (e.g. at 60°C, ≤ − 60°C). The validated assay is well suited for the intended clinical study where conventional pharmacokinetic sample collection is not feasible.     

The real-time analysis of gases by direct sampling–mass spectrometry: elemental and molecular applications

Real-time analysis of gases for volatile organic compounds or elements is required for a number of applications. Direct sampling-mass spectrometry (DS-MS) is one approach to solve these analytical problems. This article reviews various instrumental configurations and applications of DS-MS. Inlet systems employed for DS-MS include membranes, microtrap interfaces, atmospheric sampling glow discharge ionization, atmospheric pressure ionization, microwave plasma ionization, and capillary restrictors. The use of laser-based ionization methods for DS-MS is described, including resonance-enhanced multiphoton ionization and single photon ionization.     

Development and validation of dried blood spots technique for quantitative determination of topiramate using liquid chromatography–tandem mass spectrometry

An LC‐MS/MS method for determination of the anti‐epileptic drug topiramate (TPM) in dried blood spot (DBS) samples was developed and validated. DBS samples were prepared by spotting 30 μL of spiked whole blood onto FTA^TM DMPK‐C cards and drying for at least 3 h. Six‐millimetre punched spots were then extracted by using a mixture of methanol and water (90:10, v/v) with deuterated internal standard (topiramate‐d₁₂). The extracted samples were injected into a liquid chromatograph equipped with a tandem mass spectrometric detector. Negative ions were monitored in the selected reaction monitoring mode and transitions m/z 338.2 → 78.1 and m/z 350.3 → 78.1 were used for the quantitative evaluation of TPM and internal standard, respectively. The results obtained from validation were statistically evaluated according to the requirements of the European Medicines Agency and US Food and Drug Administration regulatory guidelines. The linearity of the method was checked within a concentration range from 10 to 2000 ng/mL. The validation results indicate that the method is accurate, precise, sensitive, selective and reproducible. Copyright © 2013 John Wiley & Sons, Ltd.     

Electrostatic field–induced tip‐electrospray ionization mass spectrometry for direct analysis of raw food materials

Rapid characterization of metabolites and risk compounds such as chemical residues and natural toxins in raw food materials such as vegetables, meats, and edible living plants and animals plays an important part in ensuing food quality and safety. To rapidly characterize the analytes in raw food materials, it is essential to develop in situ method for directly analyzing raw food materials. In this work, raw food materials including biological tissues and living samples were placed between an electrode and mass spectrometric (MS) inlet under a strong electrostatic field; analytes were rapidly induced to generate electrospray ionization (ESI) from the sample tip by adding a drop of solvent onto the sample. Therefore, the electrostatic field–induced tip‐ESI‐MS allows raw samples to avoid contacting high voltage, and thus this method has the advantage for in vivo analysis of food living plants and animals. Metabolite profiling, residues of pesticides and veterinary drugs, and natural toxins from raw food materials have been successfully detected. The analytical performances, including the linear ranges, sensitivity, and reproducibility, were investigated for direct sample analysis. The ionization mechanism of electrostatic field–induced tip‐ESI was also discussed in this work.     

Evaluation of matrix effects in analysis of estrogen using liquid chromatography–tandem mass spectrometry

Matrix effects of different biological samples, including phosphate‐buffered saline–bovine serum albumin (PBS‐BSA), gelded horse serum, mouse serum, and mouse brain, were investigated for the determination of 17α‐ and β‐estradiol using derivatization with dansyl chloride prior to LC‐MS/MS. Matrix effects were evaluated based on the slopes of regression lines plotted from results obtained in biological matrices versus pure standard solutions. Such plots indicate the enhancement or suppression of signal based on the presence of a particular biological fluid for a particular method. The matrix effects from PBS‐BSA were similar to those of mouse serum. In contrast, analyses performed from horse serum and mouse brain yielded significant ion suppression, especially for 17β‐estradiol. Precipitation during derivatization was observed when pre‐concentrated samples were processed with ethyl acetate as an extraction solvent. This was overcome with the use of methyl tert‐butyl ether; however, matrix effects from this preparation were still present, evidenced by signal suppression and poor linearity in the standard curve. This work affirms that caution should be taken in the transfer of methods for use with different biological matrices, especially in the case where surrogate matrices are necessary for calibration purposes.     

Early synthetic dyes – a challenge for tandem mass spectrometry

The present study concerns identification of early yellow synthetic dyes from silk fibers taken from the 1930 spring color palette of the Lyon Dyers’ Guild (La Chambre Syndicale des Teinturiers). The identification was based mainly on the electrospray ionization tandem mass spectrometry spectra obtained in the positive and negative ion modes. This technique was combined with high‐performance liquid chromatography, which enabled separation of the analyzed compounds. Spectra registered for each of the examined synthetic dye allowed identification of their lost fragments. Moreover, isotopic profiles and exact measurements of m/z by using time of flight analyzer made possible to evaluate their elemental composition. In consequence, all obtained data, including UV–vis spectra, allowed to reconstruct molecular structures of examined colorants. Due to the lack of standards, the identification of the dyes was based only on the registration of fragment and quasi‐molecular ions, what is rather uncommon in such analysis and means groping for the correct structure rather than proving signal identity by comparison with standards. Depending on substituents present in dye molecules, the lost fragments of the examined compounds involved SO₂, NO^?, NO₂^?, CH₄, C₂H₄, C₂H₅^?, C₂H₆, CH₂ = N–CH₃, (CH₃)₂NH, CH₂ = NH, CH₃–NH₂, as well as CO and CO₂. The performed study led to identification of various colorants: rhodamine 6 G, rhodamine B, malachite green, quinoline yellow, picric acid and acetoquinone yellow 5JZ. Copyright © 2013 John Wiley & Sons, Ltd.     

Determination of chlorine species by capillary electrophoresis – mass spectrometry

The applicability of CZE with mass spectrometric detection for the determination of four chlorine species, namely chloride and three stable chlorine oxyanions, was studied. The main aspects of the proper selection of BGE and sheath liquid for the CE‐MS determinations of anions with high mobility were demonstrated, pointing out the importance of pH and the mobility of the anion in the BGE. The possibility of using uncoated fused silica capillary and common electrolytes for the separation was shown and the advantage of using extra pressure at the inlet capillary end was also presented. The linear range was found to be 1–100 µg/mL for ClO₃^? and ClO₄^?, 5–500 µg/mL for ClO₂^?, and 25–500 µg/mL for Cl^?, but the sensitivity can be greatly improved if larger sample volume is injected and electrostacking effect is utilized. The LOD for ClO₃^? in drinking water was 6 ng/mL, when very large sample volume was injected (10 000 mbar·s was applied).