Sample Preparation Techniques Research Articles

Tethered Particle Motion Technique in Crowded Media: Compaction of DNA by Globular Macromolecules.

Tethered Particle Motion (TPM) is a single molecule technique, which consists in tracking the motion of a nanoparticle (NP) immersed in a fluid and tethered to a glass surface by a DNA molecule. The present work addresses the question of the applicability of TPM to fluids which contain crowders at volume fractions ranging from that of the nucleoid of living bacteria (around 30%) up to the jamming threshold (around 66%). In particular, we were interested in determining whether TPM can be used to characterize the compaction of DNA by globular crowders. To this end, extensive Brownian Dynamics simulations were performed with a specifically built coarse-grained model. Analysis of the simulations reveals several effects not observed in dilute media, which impose constraints on the TPM setup. In particular, the Tethered Fluorophore Motion (TFM) technique, which consists in replacing the NP by a much smaller fluorophore, is probably better suited than standard TPM. Moreover, a sample preparation technique which does not involve hydrophilic patches may be required. Finally, the use of a DNA brush may be needed to achieve DNA concentrations close to in vivo ones.

Advances in targeted liquid chromatography-tandem mass spectrometry methods for endocannabinoid and N-acylethanolamine quantification in biological matrices: A systematic review.

Liquid chromatography paired with tandem mass spectrometry (LC-MS/MS) is the gold standard in measurement of endocannabinoid concentrations in biomatrices. We conducted a systematic review of literature to identify advances in targeted LC-MS/MS methods in the period 2017-2024. We found that LC-MS/MS methods for endocannabinoid quantification are relatively consistent both across time and across biomatrices. Recent advances have primarily been in three areas: (1) sample preparation techniques, specific to the chosen biomatrix; (2) the range of biomatrices tested, recently favoring blood matrices; and (3) the breadth of endocannabinoid and endocannabinoid-like analytes incorporated into assays. This review provides a summary of the recent literature and a guide for researchers looking to establish the best methods for quantifying endocannabinoids in a range of biomatrices.

PANAMA-enabled high-sensitivity dual nanoflow LC-MS metabolomics and proteomics analysis.

High-sensitivity nanoflow liquid chromatography (nLC) is seldom employed in untargeted metabolomics because current sample preparation techniques are inefficient at preventing nanocapillary column performance degradation. Here, we describe an nLC-based tandem mass spectrometry workflow that enables seamless joint analysis and integration of metabolomics (including lipidomics) and proteomics from the same samples without instrument duplication. This workflow is based on a robust solid-phase micro-extraction step for routine sample cleanup and bioactive molecule enrichment. Our method, termed proteomic and nanoflow metabolomic analysis (PANAMA), improves compound resolution and detection sensitivity without compromising the depth of coverage as compared with existing widely used analytical procedures. Notably, PANAMA can be applied to a broad array of specimens, including biofluids, cell lines, and tissue samples. It generates high-quality, information-rich metabolite-protein datasets while bypassing the need for specialized instrumentation.

Optimization of sample preparation, fluorescence- and Raman techniques for environmental microplastics

Microspectroscopic imaging techniques based on spontaneous Raman scattering, Stimulated Raman Scattering (SRS), or fluorescence (with a selective dye) can be used to detect environmental microplastics (MPs) and determine their chemical as well as physical properties. The present study first focuses on optimizing the sample preparation, including a new design for a density separation apparatus and optimization of the Nile Red staining procedure. Tests were carried out with both white and colored reference materials, as well as environmental MPs in a suspended matter sample from the Rhine river. The new ‘MESSY’ system has a mean recovery of 95 ± 5.5 % (three polymer materials, in duplicate). The optimized Nile Red staining allows coarse categorization of MPs into “polar” vs. “non-polar” materials based on their Fluorescence Index (emission wavelength), but fluorescent additives in the polymer can cause misclassification. For unambiguous identification of the polymer type, Raman spectroscopy can be used. Even colored polymers, with or without Nile Red staining, were readily identified by Raman spectroscopy using a red laser (785 nm), except for particles containing carbon black. A Deep-UV Raman microscope (ex = 248.6 nm) was constructed, which allowed identification of all colored plastics, even those pigmented with carbon black. Since unsupervised mapping with spontaneous Raman is very slow, point measurements are preferably used after preselection of particles of interest based on fluorescence imaging. SRS is several orders of magnitude faster than spontaneous Raman mapping but requires multiple scans at different z-heights and at multiple wavenumber settings to detect and identify all particles. The results are expected to contribute to the development of suitable methodologies for the detection and identification of environmental microplastics.

Field isolation and capture of fluoroquinolones in waters with porous monolith-based multiple channels in-tip microextraction technique prior to HPLC quantification

Field isolation and capture is significant in the accuracy and reliable monitoring of fluoroquinolones (FQs) contents in environmental waters. For this purpose, a new porous monolith-based adsorbent (PMPA) was in-situ fabricated in pipette tip using 1-octyl-3-vinylimidazolium tetrafluoroborate and 1H,1H,2H,2H-heptadecafluorodecyl acrylate as mixed functional monomers. The PMPA prepared under the optimized parameters was characterized by various techniques. In addition, the prepared PMPA was employed as the adsorbent of home-developed multiple channels in-tip microextraction technique (MCIM) for the field sample preparation of FQs. Results revealed that the obtained PMPA exhibited satisfactory capture capability towards studied FQs, and the enrichment factors ranged from 128 to 203. Under the most beneficial extraction conditions, the developed PMPA/MCIM was employed to on-site extract trace FQs in a variety of aqueous samples and followed by quantification with HPLC equipped with fluorescence detector. The achieved limits of detection, precision and spiked recoveries were in the ranges of 0.22–0.78 ng/L, 1.4–9.9 % and 80.0–118 %, respectively. Furthermore, confirmation experiments with reported approach were utilized to evaluate method accuracy. In comparison with documented approaches, the current study exhibits some features such as high sensitivity, convenience, low cost and environmental friendliness. At the same time, the established method overcomes the dilemmas encountered in traditional lab-based sample preparation procedure.

A comparative evaluation of sample preparation techniques for the therapeutic monitoring of carbamazepine in human plasma by RPLC/DAD

A comparative evaluation of sample preparation techniques for the therapeutic monitoring of carbamazepine in human plasma by RPLC/DAD

Innovative Solid-Phase Extraction Strategies for Improving the Advanced Chromatographic Determination of Drugs in Challenging Biological Samples.

In the past few decades, considerable scientific strides have been made in the subject of drug analysis in human biological samples. However, the risk caused by incorrect drug plasma levels in patients still remains an important concern. This review paper attempts to investigate the advances made over the last ten years in common sample preparation techniques (SPT) for biological samples based on solid sorbents, including solid-phase extraction (SPE) and solid-phase micro-extraction (SPME), and in particular in the field of molecularly imprinted polymers (MIPs), including non-stimuli-responsive and stimuli-responsive adsorbents. This class of materials is known as 'smart adsorbents', exhibiting tailored responses to various stimuli such as magnetic fields, pH, temperature, and light. Details are provided on how these advanced SPT are changing the landscape of modern drug analysis in their coupling with liquid chromatography-mass spectrometry (LC-MS) analytical techniques, a general term that includes high-performance liquid chromatography (HPLC) and ultra-high performance liquid chromatography (UHPLC), as well as any variation of MS, such as tandem (MS/MS), multiple-stage (MSn), and high-resolution (HRMS) mass spectrometry. Some notes are also provided on coupling with less-performing techniques, such as high-performance liquid chromatography with ultraviolet (HPLC-UV) and diode array detection (HPLC-DAD) detection. Finally, we provide a general review of the difficulties and benefits of the proposed approaches and the future prospects of this research area.

Liquid-phase microextraction approach using supramolecular solvent formed in aqueous systems based on di(2-ethylhexyl)phosphoric acid salt: Spectrophotometric determination of antipyrine in saliva

In this study, di(2-ethylhexyl)phosphoric acid salt was investigated as a double-chain anionic surfactant for supramolecular solvent formation in a liquid-phase microextraction procedure for the first time. Inorganic electrolytes (sodium salts) were shown to be coacervation agents, due to their salting-out property in solution of sodium di(2-ethylhexyl)phosphate, obtained by deprotonation of the acid. The physico-chemical properties of the newly synthesised solvents were studied. Sodium carbonate and sodium phosphate were found to be the most suitable coacervation agents, due to their alkaline nature and low viscosity of the supramolecular solvent phases they produce. The extraction ability of the novel solvent was demonstrated in liquid-phase microextraction of a colored derivative of antipyrine from saliva for further spectrophotometric determination. The extraction recovery was found to be 70 %. The proposed green sample preparation technique enabled the exclusion of traditional toxic solvents and reduction of reagent consumption. The satisfactory analytical performance of the procedure was validated with real sample analysis. The calibration graph was linear over the concentration range of 0.03 to 0.5 mmol/L. The limit of detection (3σ) and limit of quantification (10σ), calculated from blank tests, were 0.01 and 0.03 mmol/L, respectively.

Exploring advanced strategies in SPME-HPLC-DAD: Enhancing analytical precision and diverse applications in modern era

In analytical chemistry, the synergy between innovative sample preparation techniques and advanced chromatographic methods has paved the way for unprecedented advancements in the analysis of complex samples. Solid Phase Microextraction (SPME) stands out as a powerful tool for extraction, and preconcentration. When coupled with High-Performance Liquid Chromatography (HPLC) equipped with a Diode Array Detector (DAD), the integrated SPME-HPLC-DAD system emerges as a sophisticated analytical platform, offering enhanced sensitivity, selectivity, and versatility for various applications. Desorption is accomplished by either depositing organic solvent in the desorption chamber or else by flowing the mobile phase through the desorption chamber specificity. An ordinary description, along with a visual illustration, has been thoroughly provided for concepts of developed well-known enhanced thin film SPME cartridges (Figure 2). The tabular representation is also available with proper description like technique name, targeted analyte, column used, mobile phase used, %RSD, extraction recovery, LOD, and data acquired R2 also considered Based on the information available in the present study. Discussed SPME with HPLC-DAD brings a great turning point by successfully utilization in biomedical sciences, environmental monitors, food safety, and clinical analysis This cost-effective, readily available technique is both environmentally friendly and stable, allowing for rapid, reliable, and accurate measurements across a wide range of analyte concentrations, thanks to the utilization of hyphenated techniques. The different strategies of SPME with HPLC -DAD and possible future apllications have been incorporated in the review. The understanding of these techniques is still limited and challenging.

Thermoset droplet curing performance in the microbond test

ABSTRACT Users of the microbond test assume that a microbond resin droplet’s properties are equivalent to a macroscale specimen. However, there is currently no standardised methodology for determining the cure state of droplet specimens used in the microbond test. In this paper, we present a technique for microbond test users to better understand the properties of thermoset droplet specimens. Utilising a conventional benchtop spectrometer, a novel sample preparation technique involving curing epoxy droplets on thin-steel filaments allowed for high-throughput determination of the microbond droplet cure state. The parity between steel filament and glass fibre microbond samples was confirmed by infrared microspectroscopy. It is shown that cure schedules used in manufacturing composite parts produced microbond droplets with degrees of cure lower than that of bulk matrix specimens subjected to an identical thermal history. Testable microbond droplets could only be prepared for commercial resin systems when introducing a room temperature pre-curing time of at least 2 h. It is concluded that microbond testing should be supported by some droplet cure state characterisation methods to ensure that interfacial effects are not artefacts of droplet sample preparation.

Increasing protein identifications in bottom-up proteomics of T. castaneum − Exploiting synergies of protein biochemistry and bioinformatics

Depending on the respective research question, LC-MS/MS based bottom-up proteomics poses challenges from the initial biological sample all the way to data evaluation. The focus of this study was to investigate the influence of sample preparation techniques and data analysis parameters on protein identification in Tribolium castaneum by applying free software proteomics platform Max Quant.Multidimensional protein extraction strategies in combination with electrophoretic or chromatographic off-line protein pre-fractionation were applied to enhance the spectrum of isolated proteins from T. castaneum and reduce the effect of co-elution and ion suppression effects during nano-LC-MS/MS measurements of peptides. For comprehensive data analysis, MaxQuant was used for protein identification and R for data evaluation. A wide range of parameters were evaluated to gain reproducible, reliable, and significant protein identifications.A simple phosphate buffer, pH 8, containing protease and phosphatase inhibitor cocktail and application of gentle extraction conditions were used as a first extraction step for T.castaneum proteins. Furthermore, a two-dimensional extraction procedure in combination with electrophoretic pre-fractionation of extracted proteins and subsequent in-gel digest resulted in almost 100% increase of identified proteins when compared to chromatographic fractionation as well as one-pot-analysis. The additionally identified proteins could be assigned to new molecular functions or cell compartments, emphasizing the positive effect of extended sample preparation in bottom-up proteomics. Besides the number of peptides during post-processing, MaxQuant’s Match between Runs exhibited a crucial effect on the number of identified proteins. A maximum relative standard deviation of 2% must be considered for the data analysis.Our work with Tribolium castaneum larvae demonstrates that sometimes – depending on matrix and research question − more complex and time-consuming sample preparation can be advantageous for isolation and identification of additional proteins in bottom-up proteomics.

Enhancing Odor Analysis with Gas Chromatography-Olfactometry (GC-O): Recent Breakthroughs and Challenges.

Gas chromatography-olfactometry (GC-O) has made significant advancements in recent years, with breakthroughs in its applications and the identification of its limitations. This technology is widely used for analyzing complex odor patterns. The review begins by explaining the principles of GC-O, including sample preparation, separation methods, and olfactory evaluation techniques. It then explores the diverse range of applications where GC-O has found success, such as food and beverage industries, environmental monitoring, perfume and aroma development, and forensic analysis. One of the major breakthroughs in GC-O analysis is the improvement in separation power and resolution of odorants. Techniques like rapid GC, comprehensive two-dimensional GC, and multidimensional GC have enhanced the identification and quantification of odor-active chemicals. However, GC-O also has limitations. These include the challenges in detecting and quantifying trace odorants, dealing with matrix effects, and ensuring the repeatability and consistency of results across laboratories. The review examines these limitations closely and discusses potential solutions and future directions for improvement in GC-O analysis. Overall, this review presents a comprehensive overview of the recent advances in GC-O, covering breakthroughs, applications, and limitations. It aims to promote the wider usage of GC-O analysis in odor analysis and related industries. Researchers, practitioners, and anyone interested in leveraging the capabilities of GC-O in analyzing complex odor patterns will find this review a valuable resource. The article highlights the potential of GC-O and encourages further research and development in the field.

Reflecting on the Influence of the Current State of Sample Preparation on GC, Part 2: Techniques

Recently, we examined major trends in sample preparation and how they relate to gas chromatography (GC), based on a 2023 user survey by LCGC International. In Part 2 of LCGC’s survey of sample preparation users, the prevalence of sample preparation techniques, from the most common, such as weighing, to the most complex, such as online extractions, was discussed. In this installment, we examine trends in the use of sample preparation techniques through the lens of instrumental analysis by gas chromatography (GC) and GC–mass spectrometry (GC–MS). We see that, although common and classical techniques, such as filtration and centrifugation, are still generally the most widely used, newer techniques that allow automation, simplification of analysis steps, and green analysis are gaining traction.

Unveiling Pharmacokinetics and Drug Interaction of Linagliptin and Pioglitazone HCl in Rat Plasma Using LC-MS/MS.

The linagliptin (LIN) and pioglitazone HCl (PIO) combination, currently undergoing phase III clinical trials for diabetes mellitus treatment, demonstrated significant improvements in glycemic control. However, the absence of an analytical method for simultaneous determination in biological fluids highlights a crucial gap. This underscores the pressing need for sensitive bioanalytical methods, emphasizing the paramount importance of developing such tools to advance diabetes management strategies and enhance patient care. Herein, a sensitive reverse-phase high-performance liquid chromatography-electrospray ionization-tandem mass spectrometry method was developed for simultaneous determination of LIN and PIO in rat plasma using alogliptin as an internal standard. Chromatographic separation was performed on an Agilent Eclipse Plus C18 (4.6 × 100 mm, 3.5 μm) using an isocratic mobile phase system consisting of ammonium formate (pH 4.5) and methanol using an acetonitrile-induced protein precipitation technique for sample preparation. Multiple reaction monitoring in positive ion mode was used for quantitation of the precursor to production at m/z 473.2 → 419.9 for LIN, 357.1 → 134.2 for PIO, and 340.3 → 116.1 for ALO. The linearity range was 0.5 to 100 and 1 to 2000 ng/mL for LIN and PIO, respectively. The developed method was validated as per US-FDA guidelines and successfully applied to clinical pharmacokinetic and drug-drug interaction studies with a single oral administration of LIN and PIO in rat plasma. Pharmacokinetic parameters of LIN were significantly influenced by the concomitant administration of PIO and vice versa. Molecular modeling revealed the significant interaction of LIN and PIO with P-glycoprotein. Therefore, the drug-drug interaction between LIN and PIO deserves further study to improve drug therapy and prevent dangerous adverse effects.

Isotope Distribution Analysis in H₂18O Pulse-Labeled Trees Frozen with Liquid Nitrogen.

Tracer injection has long been recognized as a valuable tool for delineating tree hydraulics and assessing water transport pathways. Recently, isotope tracers have emerged as innovative instruments for investigating tree hydraulics, providing new insights into tree water dynamics. Nevertheless, there is a critical need for further research to comprehensively grasp water movement and distribution within trees. A previously introduced technique for analyzing the isotopic ratio of water in wet tissues, offering millimeter-scale resolution for visualizing tracer movement, faces challenges due to its underdeveloped sample preparation techniques. In this study, we introduced an H2 18O tracer into S. gracilistyla samples, exclusively comprising indeterminate roots, stems, and leaves, cultivated through hydroponics and grown within the current year. Our objective was to assess the axial distribution of the tracer in the xylem. Additionally, we devised a novel method for preparing frozen wet tissue samples, enhancing the repeatability and success rate of experiments. The results demonstrated that all frozen wet tissue samples exhibited an average water loss rate of less than 0.6%. Isotopic analysis of these samples unveiled a consistent decline in tracer concentration with increasing height in all Salix specimens, with three out of five samples revealing a significant isotope gradient. Our findings affirm the efficacy and practicality of combining isotopic labeling with freezing, stabilization, and preparation techniques. Looking ahead, our isotopic labeling and analysis methods are poised to transcend woody plants, finding extensive applications in plant physiology and ecohydrology.

Aluminium Determination in Foodstuffs by ICP-MS: Influence of Microwave Digestion Parameters on the Recovery

Aluminium (Al) is the third most common element in the Earth’s crust and occurs naturally in drinking water and agricultural products, and humans are consequently exposed to the element from dietary sources. A tolerable weekly intake of 1 mg/kg has been established by the European Food Safety Authority (EFSA); however, no maximum levels for aluminium in foodstuffs have so far been established in the European Union (EU) legislation. Official food control requires validated methods for the determination of aluminium. Acid digestion assisted by microwaves is the main sample preparation technique used for the determination of aluminium, usually in combination with atomic spectrometry for quantification. In the present study, different parameters in the digestion step were investigated including test portion, digestion temperature, the reagent used and duration of the digestion to assess the aluminium extraction. The presented work is following up on an observation from a proficiency test (PT) on trace elements (including aluminium) in cocoa powder organised in 2020 by the European Union Reference Laboratory for metals and nitrogenous compounds in feed and food (EURL-MN), where the participant results for aluminium showed an unexpectedly large variation. In addition to the PT material, different certified reference materials were included in the present study, and the results highlighted that the temperature and reagent used are the most critical parameters to obtain a satisfactory sample digestion prior to aluminium determination. Based on the obtained results, it is recommended to digest food samples with a mix of ultrapure water and nitric acid for 25 min at a temperature of at least 240 °C with a mix of HNO3 and H2O to achieve satisfactory microwave-assisted digestion.

Natural kapok fiber as a green and efficient adsorbent for in-syringe solid-phase extraction in the determination of antidepressants in biofluids

Effective antidepressant monitoring is crucial for safe treatment, with liquid chromatography being the primary detection method. Due to the complexity of biofluids, meticulous sample preparation is vital to remove interferences and concentrate analytes. In this study, a novel solid-phase extraction method utilizing unprocessed kapok fiber (KF) as an eco-friendly and efficient sorbent, ingeniously incorporated within an in-syringe device for the extraction of antidepressants from biofluids was introduced. The extraction process displayed several advantages, including simplicity and rapidity (less than 3 min with minimal manual intervention), as well as the potential for automation, which represents significant advancements in sample preparation techniques. The method demonstrated excellent analytical performance when coupled with high-performance liquid chromatography-ultraviolet detection, achieving good linearity (R2 ≥ 0.991), excellent relative recovery (92.6–107.5 %), high precision in intraday and interday analyses (RSDs within 0.8–6.0 % and 1.2–7.9 %, respectively), and good sensitivity with limits of quantification between 5–10 ng/mL. Notably, given the use of a renewable and biodegradable material with KF, and a low-toxicity desorption solvent with ethanol, the developed method stands out for its environmental sustainability, aligning with green analytical chemistry principles. Its application to real biological samples validates its efficacy as a rapid and reliable tool for clinical and forensic monitoring of antidepressants. Moreover, the low cost of this method facilitated by the economical nature of KF and the reusability of syringes further enhances its practicability. This study presents a unique approach by employing natural and renewable KF as adsorbents, thereby eliminating the need for complex material synthesis and making a pioneering contribution to the field of analytical chemistry for drug analysis in complex matrices.

Rapid microcantilever preparation for conditional fracture toughness evaluation

The Ga+ focused Ion Beam (FIB) or Xe+ Plasma FIB (PFIB) procedure has enabled the ability to make specimens where mechanical tests in the microscale regime can be achieved with high precision. Nevertheless, milling times can be long and shape optimization can be arduous collectively limiting the ability to readily make a sufficient number of test samples. In this work, we present a sample preparation technique for microscale cantilever fabrication which streamlines the milling procedures by using combined electropolishing and FIB milling. Specifically, the procedure mitigates extensive undercutting requirements which reduces the fabrication time for the microcantilever to more than half that time as compared to direct milling methods from making similar geometries directly from the bulk material. This process is discussed along with specific aspects related to the sample alignment of the cantilever to specific fracture planes for single crystals. In addition, we comment on the applicability of this method to polycrystalline samples.

In vivo profiling of phytocannabinoids in Cannabis spp. varieties via SPME-LC-MS analysis

BackgroundIn vivo solid-phase microextraction (SPME) is a minimally invasive, non-exhaustive sample-preparation technique that facilitates the direct isolation of low molecular weight compounds from biological matrices in living systems. This technique is especially useful for the analysis of phytocannabinoids (PCs) in plant material, both for forensic purposes and for monitoring the PC content in growing Cannabis spp. plants. In contrast to traditional extraction techniques, in vivo SPME enables continuous tracking of the changes in the level of PCs during plant growth without the need for plant material collection. In this study, in vivo SPME utilizing biocompatible C18 probes and liquid-chromatography coupled to quadrupole time-of flight mass spectrometry (LC-Q-TOF-MS) is proposed as a novel strategy for the extraction and analysis of the acidic forms of five PCs in growing medicinal cannabis plants. ResultsThe SPME method was optimized by testing various parameters, including the extraction phase (coating), extraction and desorption times, and the extraction temperature. The proposed method was validated with satisfactory analytical performance regarding linearity (10–3000 ng/mL), limits of quantification, and precision (relative standard deviations below 5.5 %). The proposed method was then successfully applied for the isolation of five acidic forms of PCs, which are main components of growing medicinal cannabis plants. As a proof-of-concept, SPME probes were statically inserted into the inflorescences of two varieties of Cannabis spp. plants (i.e., CBD-dominant and Δ9-THC-dominant) cultivated under controlled conditions for 30 min extraction of tetrahydrocannabinolic acid (Δ9-THCA), cannabidiolic acid (CBDA), cannabigerolic acid (CBGA), cannabiviarinic acid (CBVA), and tetrahydrocannabivarinic acid (THCVA). Significance and noveltyThe results confirmed that the developed SPME-LC-Q-TOF-MS method is a precise and efficient tool that enables direct and rapid isolation and analysis of PCs under in vivo conditions. The proposed methodology is highly appealing option for monitoring the metabolic pathways and compositions of multiple PCs in medicinal cannabis at different stages of plant growth.

Fiber-in-tube SPME-CapLC-MS/MS method to determine Aβ peptides in cerebrospinal fluid obtained from Alzheimer's patients

Mass spectrometry is characterized by its high sensitivity, ability to measure very low analyte concentrations, specificity to distinguish between closely related compounds, availability to generate high-throughput methods for screening, and high multiplexing capacity. This technique has been used as a platform to analyze fluid biomarkers for Alzheimer's disease. However, more effective sample preparation procedures, preferably antibody-independent, and more automated mass spectrometry platforms with improved sensitivity, chromatographic separation, and high throughput are needed for this purpose. This short communication discusses the development of a fiber-in-tube SPME-CapLC-MS/MS method to determine Aβ peptides in cerebrospinal fluid obtained from Alzheimer's disease patients. To obtain the fiber-in-tube SPME capillary, we longitudinally packed 22 nitinol fibers coated with a zwitterionic polymeric ionic liquid into the same length of the PEEK tube. In addition, this communication compares this fiber-in-tube SPME method with the conventional HPLC scale (HPLC-MS/MS) and when directly coupled to CapESI-MS/MS without chromatographic separation, and, as a case study, discusses the benefits and challenges inherent in miniaturizing the flow scale of the sample preparation technique (fiber-in-tube SPME) to the CapLC-MS/MS system. Fiber-in-tube SPME-CapLC-MS/MS provided LLOQ ranging from 0.09 to 0.10 ng mL−1, accuracy ranging from 91 to 117 % (recovery), and reproducibility of less than 18 % (RSD). Analysis of the cerebrospinal fluid samples obtained from Alzheimer's disease patients evidenced that the method is robust. At the capillary scale (10 µL min−1), this innovative method presented higher analytical sensitivity than the conventional HPLC-MS/MS scale. Although fiber-in-tube SPME directly coupled to CapESI-MS/MS offers advantages in terms of high throughput, the sample was dispersed and non-quantitatively desorbed from the capillary at low flow rate. These results highlighted that chromatographic separation is important to decrease the matrix effect and to achieve higher detectability, which is indispensable for bioanalysis.