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The integration of microfluidics with electric field control, commonly referred to as electrofluidics, has led to new opportunities for biomedical analysis. The requirement for closed microcapillary channels in microfluidics, typically formed via complex microlithographic fabrication approaches, limits the direct accessibility to the separation processes during conventional electrofluidic devices. Textile structures provide an alternative and low-cost approach to overcome these limitations via providing open and surface-accessible capillary channels. Herein, we investigate the potential of different 3D textile structures for electrofluidics. In this study, 12 polyester yarns were braided around nylon monofilament cores of different diameters to produce functional 3D core-shell textile structures. Capillary electrophoresis performances of these 3D core-shell textile structures both before and after removing the nylon core were evaluated in terms of mobility and bandwidth of a fluorescence marker compound. It was shown that the fibre arrangement and density govern the inherent capillary formation within these textile structures which also impacts upon the solute analyte mobility and separation bandwidth during electrophoretic studies. Core-shell textile structures with a 0.47 mm nylon core exhibited the highest fluorescein mobility and presented a narrower separation bandwidth. This optimal textile structure was readily converted to different geometries via a simple heat-setting of the central nylon core. This approach can be used to fabricate an array of miniaturized devices that possess many of the basic functionalities required in electrofluidics while maintaining open surface access that is otherwise impractical in classical approaches., (© 2024 The Author(s). ELECTROPHORESIS published by Wiley‐VCH GmbH.)

The coupling of capillary electrophoresis (CE) with capacitively coupled contactless conductivity detection (C 4 D) has become convenient analytical method for determination of small molecules that do not possess chromogenic or fluorogenic group. The implementations of CE with C 4 D in the determination of inorganic and organic ions and amino acids in biomedical field are demonstrated. Attention on background electrolyte composition, sample treatment procedures, and the utilize of multi-detection systems are described. A number of tables summarizing highly developed CE-C 4 D methods and the figures of merit attained are involved. Lastly, concluding remarks and perspectives are argued., (© 2023 Wiley-VCH GmbH.)

Topiramate (TPM) is the main antiepileptic drug used for the control of partial and generalized seizures in both adults and children. In association with clinical observations, the analysis of plasmatic concentration of TPM is of utmost importance for the individual adjustment of the administered dose to the patient. In the present work, a bioanalytical method was developed and validated for TPM analysis in plasma samples by capillary electrophoresis with capacitively-coupled contactless conductivity detection (CE-C 4 D). A simple background electrolyte composed of 15 mmol/L triethylamine, hydrodynamic injections (0.8 psi for 5 s) and a moderate separation voltage (20 kV) were used, rendering relatively short analysis times (<3 min). The sample pre-treatment was carried out by liquid-liquid extraction using methyl terc-butyl ether as solvent and 200 μL of plasma. The method was validated according to the official guidelines from the European Medicine Agency and showed linearity in plasmatic concentration range from 1 to 30 μg/mL, which covers the clinically-relevant interval. The lower limit of quantification of 1 μg/mL obtained also allows following patients with low dosage of the drug. The method was successfully applied to analysis of plasma samples and allowed the identification of 80% under-medicated patients in the analyzed patient pool., (© 2018 WILEY-VCH Verlag GmbH & Co. KGaA, Weinheim.)

This review is in support of the development of selective, precise, fast, and validated capillary electrophoresis (CE) methods. It follows up a similar article from 1998, Wätzig H, Degenhardt M, Kunkel A. "Strategies for capillary electrophoresis: method development and validation for pharmaceutical and biological applications," pointing out which fundamentals are still valid and at the same time showing the enormous achievements in the last 25 years. The structures of both reviews are widely similar, in order to facilitate their simultaneous use. Focusing on pharmaceutical and biological applications, the successful use of CE is now demonstrated by more than 600 carefully selected references. Many of those are recent reviews; therefore, a significant overview about the field is provided. There are extra sections about sample pretreatment related to CE and microchip CE, and a completely revised section about method development for protein analytes and biomolecules in general. The general strategies for method development are summed up with regard to selectivity, efficiency, precision, analysis time, limit of detection, sample pretreatment requirements, and validation., (© 2023 The Authors. Electrophoresis published by Wiley-VCH GmbH.)

Composting is a sustainable approach to manage animal and vegetal waste generated in the Fundação Parque Zoológico de São Paulo. The resulting compost is often used in ZOO's premises as an organic fertilizer for the production of vegetables, which is further used to feed the animals. The composting product provides many forms of mineral and also amino acids (AA) that are absorbed by plants as nutrients. Since most amino acids absorb only slightly or not at all in the UV wavelengths, we developed a method for the determination of AA of agricultural interest in the composting samples. Due to the complexity of samples, we used ion exchange chromatography for the purification of AA prior to analysis. The proposed CZE-C 4 D method allowed a separation of the AA in a short analysis time (less than 3.0 min), with great linearity (with R 2 ranging from 0.993 to 0.998). Using a BGE of 10 mmol/L TEA, reduction of high-frequency noise and lower baseline fluctuations were obtained. The LOQ for the five AA were around 35 μmol/L, and were adequate for our purpose. In addition, the method showed good precision (RSD of peak area and migration time less than 1.55 and 1.16%, respectively)., (© 2016 WILEY-VCH Verlag GmbH & Co. KGaA, Weinheim.)

Cancer has been a disease of great concern because it is the second main cause of death in the world. Cures for most cancer pathologies have not yet been found, and an accurate and early diagnosis is essential for successful treatment. Therefore, research on tumor biomarkers has noticeably increased in recent years. The determination of such biomolecules, together with the routinely used laboratory exams for cancer diagnosis, would constitute a more reliable approach, known as systems biology. The "omics" era has corroborated in such investigations since the development of new technologies has arisen along with it. One of the techniques applied to the investigation of tumor biomarkers is CE, and the increasing applications of CE-MS in this field are also observed. This review covers the published literature on tumor biomarker investigations by CE-MS and related techniques, mostly within the last decade, but not limited to it. For didactic reasons this review is divided into the tumor biomarkers chemical classes, namely, proteins and related molecules, DNA adducts and modified nucleosides.

Protein hydrolysates have been used as active principles in cosmetic products conferring different properties to the final formulations, which are mostly controlled by the peptide size and its amino acid sequence. In this work, capillary electrophoresis coupled to mass spectrometry analyses were carried out in order to investigate such characteristics of protein hydrolysates. Samples of different origins (milk, soy and rice) were obtained from a local company, and were analyzed without a previous preparation step. The background electrolyte (BGE) and sheath liquid compositions were optimized for each sample. The best BGE composition (860 mmol/L formic acid--pH 1.8--in 70:30 v/v water/methanol hydro-organic solvent) was chosen based on the overall peak resolution whereas the best sheath liquid was selected based on increased sensitivity and presented different compositions to each sample (10.9-217 mmol/L formic acid in 75:25-25:75 v/v water/methanol hydro-organic solvent). Most of the putative peptides in the hydrolysate samples under investigation presented molecular masses of 1000 Da or less. De novo sequencing was carried out for some of the analytes, revealing the hydrophobicity/polarity of the peptides. Hence, the technique has proved to be an advantageous tool for the quality control of industrial protein hydrolysates., (Copyright © 2011 WILEY-VCH Verlag GmbH & Co. KGaA, Weinheim.)

The possibility to compress analyte bands at the beginning of CE runs has many advantages. Analytes at low concentration can be analyzed with high signal-to-noise ratios by using the so-called sample stacking methods. Moreover, sample injections with very narrow initial band widths (small initial standard deviations) are sometimes useful, especially if high resolutions among the bands are required in the shortest run time. In the present work, a method of sample stacking is proposed and demonstrated. It is based on BGEs with high thermal sensitive pHs (high dpH/dT) and analytes with low dpK(a)/dT. High thermal sensitivity means that the working pK(a) of the BGE has a high dpK(a)/dT in modulus. For instance, Tris and Ethanolamine have dpH/dT=-0.028/ degrees C and -0.029/ degrees C, respectively, whereas carboxylic acids have low dpK(a)/dT values, i.e. in the -0.002/ degrees C to+0.002/ degrees C range. The action of cooling and heating sections along the capillary during the runs affects also the local viscosity, conductivity, and electric field strength. The effect of these variables on electrophoretic velocity and band compression is theoretically calculated using a simple model. Finally, this stacking method was demonstrated for amino acids derivatized with naphthalene-2,3-dicarboxaldehyde and fluorescamine using a temperature difference of 70 degrees C between two neighbor sections and Tris as separation buffer. In this case, the BGE has a high pH thermal coefficient whereas the carboxylic groups of the analytes have low pK(a) thermal coefficients. The application of these dynamic thermal gradients increased peak height by a factor of two (and decreased the standard deviations of peaks by a factor of two) of aspartic acid and glutamic acid derivatized with naphthalene-2,3-dicarboxaldehyde and serine derivatized with fluorescamine. The effect of thermal compression of bands was not observed when runs were accomplished using phosphate buffer at pH 7 (negative control). Phosphate has a low dpH/dT in this pH range, similar to the dK(a)/dT of analytes. It is shown that mid R:dK(a)/dT-dpH/dTmid R>>0 is one determinant factor to have significant stacking produced by dynamic thermal junctions.

In a previous work [M. Mandaji, et al., this issue] a sample stacking method was theoretically modeled and experimentally demonstrated for analytes with low dpK(a)/dT (analytes carrying carboxylic groups) and BGEs with high dpH/dT (high pH-temperature-coefficients). In that work, buffer pH was modulated with temperature, inducing electrophoretic mobility changes in the analytes. In the present work, the opposite conditions are studied and tested, i.e. analytes with high dpK(a)/dT and BGEs that exhibit low dpH/dT. It is well known that organic bases such as amines, imidazoles, and benzimidazoles exhibit high dpK(a)/dT. Temperature variations induce instantaneous changes on the basicity of these and other basic groups. Therefore, the electrophoretic velocity of some analytes changes abruptly when temperature variations are applied along the capillary. This is true only if BGE pH remains constant or if it changes in the opposite direction of pK(a) of the analyte. The presence of hot and cold sections along the capillary also affects local viscosity, conductivity, and electric field strength. The effect of these variables on electrophoretic velocity and band stacking efficacy was also taken into account in the theoretical model presented. Finally, this stacking method is demonstrated for lysine partially derivatized with naphthalene-2,3-dicarboxaldehyde. In this case, the amino group of the lateral chain was left underivatized and only the alpha amino group was derivatized. Therefore, the basicity of the lateral amino group, and consequently the electrophoretic mobility, was modulated with temperature while the pH of the buffer used remained unchanged.

In this work, a CE equipment, online hyphenated to an IT MS analyzer by a linear sheath liquid interface promoting ESI, was used to develop a method for quantitative determination of amino acids. Under appropriate conditions (BGE composition, 0.8% HCOOH, 20% CH3OH; sheath liquid composition, 0.8% HCOOH, 60% methanol; V ESI, +4.50 kV), analytical curves of all amino acids from 3 to 80 mg/L were recorded presenting acceptable linearity (r >0.99). LODs in the range of 16-172 micromol/L were obtained. BSA, a model protein, was submitted to different hydrolysis procedures (classical acid and basic, and catalyzed by the H+ form of a cation exchanger resin) and its amino acid profiles determined. In general, the resin-mediated hydrolysis yields were overall similar or better than those obtained by classical acid or basic hydrolysis. The resulting experimental-to-theoretical BSA concentration ratios served as correction factors for the quantitation of amino acids in Brazil nut resin generated hydrolysates.

CE-ESI-MS with a liquid sheath interface and IT mass analyzer was used for analysis of siderophores from different strains of Methylobacterium spp. citrus endophyte extracts. Three Methylobacterium strains were investigated according to positive bioassay tests. Bacteria cultures were grown under Fe(III) absence (siderophore producing cultures) and under Fe(III) presence (control cultures). Siderophores were extracted from culture supernatant with polystyrene resins. BGE and sheath-liquid composition were optimized, respectively, in order to assure both, best peak resolution and ESI-MS sensitivity. The best analysis conditions were obtained with 100 mmol/L ammonium bicarbonate at pH 8 as BGE and methanol:H(2)O 25:75 + 0.05% formic acid as sheath liquid. CZE-ESI-MS analysis revealed two possible siderophores, according to bacterium species, presenting M(r) of 1004.3 and 798.3 Da.

This work brings together some contributions for the use of nonaqueous media for proteomic analysis, for both capillary electrophoresis (CE) separation and the preparation of tryptic digests. First, a ternary nonaqueous buffer consisting of 60/30/10 v/v methanol/acetonitrile/acetic acid with 12.5 mmol/L ammonium acetate was optimized for CE separation of the tryptic digest of lysozyme. Lysozyme was chosen as a model system for the protein digestion, which has also been prepared in an organic-rich medium with methanol/50 mmol/L NH(4)HCO(3), pH 8.0 (60/40 v/v). The separation results were compared to in silico (PeptideCutter program) digestion conditions, and high-efficiency peak separation (18 peaks) was obtained in 20 min with an electric field of 350 V/cm. In addition, we have evaluated the stability of a coated capillary with poly-N,N-dimethylacrylamide (60/30 cm total/effective length and 75 microm ID) for over 100 runs of tryptic digest with the nonaqueous background electrolyte solvent system. The migration times for ten selected peptide peaks presented 3-7% relative standard deviation.

The separation of DNA molecules in polymeric solution by capillary electrophoresis involves the optimization of several variables, such as polymer solution concentration, electric field separation, temperature, etc. The optimization of each variable individually usually is a time-consuming process and the results may reach a false optimum point. Chemometric methods are suitable to be applied in such cases in which a number of variables can be optimized simultaneously. The simplex is a chemometric method that can perform such a task easily and efficiently. In this study, a simplex method was carried out to maximize the correlation coefficient (r(2)) of a logarithmic plot of mobility (mu) vs. base pair (bp), which was obtained from the separation of DNA fragments of size between 75 and 4072 bp. The simplex showed three vertexes with r(2) > 0.98 and the vertex 21 showing the highest resolution. For the fragments between 201 and 2036 bp, the r(2) increased to 0.992 with and relative standard deviation (RSD) lower than 0.2% (inter- and intra-day variation). The precision of the method in determining the size of a PCR DNA fragment was carried out using a 1 kbp DNA ladder. With the addition of an internal standard to the sample, the precision could be further improved.

In DNA analysis by capillary electrophoresis with polymer solutions there are many variables that can be optimized. However, electric field strength, polymer solution concentration and temperature of analysis are the most relevant ones. These are the variables most responsible for the fragment resolution and analysis time. Optimization of such parameters can be obtained simultaneously using chemometric techniques, reaching the optimum working conditions with few experiments. In this work, we have studied the influence of the sample composition and electrokinetic injection conditions in the reproducibility and the quality of the DNA separation results. A simplex optimization has been carried out and the optimum condition was reached with nine experiments.

We developed a method for the analysis of multiplexed double-stranded DNA (dsDNA) samples complexed to various intercalating dyes using entangled polymer solution. A commercial single-column capillary electrophoresis (CE) instrument with diode array detection was used for multiplexed detection of DNA samples by addition of intercalating fluorescent molecules. A Phi X174HinfI and a pGEM DNA ladder (1 mg/mL) were used for the electrophoretic separation of dsDNA fragments ranging in size from 24 to 726 and 36 to 2645 bp, respectively. The results suggested that simultaneous electrophoretic separation of different DNA ladders multiplexed with different dyes could be performed in the same capillary yielding fast DNA sizing separations. CE analysis, which is often overpowered by slab gel in sample throughput, could now overcome this disadvantage by allowing multiplexed sample analysis in a fraction of the time needed for slab gel analysis. The separation efficiency of stained DNA molecules with both dyes were dramatically improved with buffers containing a large cation such as tetrapentylammonium ion (Npe(4) (+)) as the only cation in the buffer.

To comply with the current needs for high-speed DNA sequencing analysis, several instruments and innovative technologies have been introduced by several groups in recent years. This review article discusses and compares the issues regarding high-throughput DNA sequencing by electrophoretic methods in miniaturized systems, such as capillaries, capillary arrays, and microchannels. Initially, general features of several capillary array designs (including commercial ones) will be considered, followed by similar analyses with microfabricated array electrophoretic devices and how they can contribute to the success of large sequencing projects.

In a previous paper, a 2% w/w replaceable high molecular mass linear polyacrylamide solution (high molecular mass LPA) was used to achieve long read-lengths for DNA sequencing by capillary electrophoresis (E. Carrilho et al., Anal. Chem. 1996, 68, 3305-3313). In that work, the polymer was prepared by polymerization in water at 6% w/w, followed by dilution to 2% w/w. In this study, an improved method for preparation of high molecular mass LPA was developed, based on inverse emulsion polymerization. With this polymerization procedure, the LPA results in a molecular mass of approximately 9 MDa, with characteristics of a fine powder of high purity and practically unlimited shelf life. Using size exclusion chromatography (SEC) and viscosity measurements to characterize the polymer, good batch-to-batch reproducibility was found. It was observed that the viscous polymer solutions made from these high molecular mass polymers require careful preparation and handling because the method of dissolution could affect the molecular mass distribution and the resultant separation of DNA components. Solutions containing 2% w/w of LPA made by emulsion polymerization were simple to prepare, resulting in excellent performance as a replaceable matrix for DNA sequencing by capillary electrophoresis. The viscosity of the polymer decreased exponentially when pressure was applied, allowing easy replacement from a capillary using a syringe. With a properly prepared matrix, a read-length of more than 1000 bases in 80 min with an accuracy better than 97%, and better than 99% for the first 800 bases, could be achieved.

Short and medium-chain fatty acids (SMCFAs) are known as essential metabolites found in gut microbiota that function as modulators in the development and progression of many inflammatory conditions as well as in the regulation of cell metabolism. Currently, there are few simple and low-cost analytical methods available for the determination of SMCFA. This report focuses on SMCFA analysis utilizing CE with indirect photometric detection (CE-IPD). A ribonucleotide electrolyte, 5'-adenosine mono-phosphate (5'-AMP), is investigated as an IPD reagent due to its high molar absorptivity and dynamic reserve compatible with separation and detection of SMCFA. The operating parameters like the composition of organic solvent, millimolar concentrations of the complexing agent (alpha-cyclodextrin), 5'-AMP and non-absorbing electrolyte (boric acid), as well as the applied voltage, are optimized for resolution, efficiency, and signal-to-noise ratio. A baseline resolution of all nine SMCFAs is achieved in less than 15 min. Additionally, the developed CE-IPD method shows promising potential to identifying SMCFA in rat fecal supernatant. The presented analytical assay is simple, economical, and has considerably good repeatability. The intraday and interday RSD of less than 1 and 2% for relative migration time, as well as less than 14 and 15% for peak area, respectively, were obtained for SMCFA in fecal solution., (© 2021 Wiley-VCH GmbH.)

Ideal-filter CE (IFCE) is a method for the selection of affinity binders for protein targets from oligonucleotide libraries, for example, random-sequence oligonucleotide libraries and DNA-encoded libraries, in a single step of partitioning. In IFCE, protein-oligonucleotide complexes and unbound oligonucleotides move in the opposite directions, facilitating very high efficiency of their partitioning. For any given protein target and oligonucleotide library, protein-oligonucleotide complexes and unbound oligonucleotides move in the opposite directions only for a limited range of EOF mobilities, which, in turn, corresponds to a limited range of pH and ionic strength values of the running buffer. Rational design of IFCE-based partitioning requires a priori knowledge of this range of pH and ionic strength values, and here we introduce an approach to predict this range for a given type of the running buffer. The approach involves measuring EOF mobilities for a relatively wide range of pH and ionic strength (I) values and finding an empirical predictor function that related the EOF mobility with pH and ionic strength. In this work, we developed a predictor function for a running buffer (Tris-HCl) that is commonly used in CE-based partitioning of affinity binders for protein targets. This predictor function can be immediately used for the rational design of IFCE-based partitioning in this running buffer, while the described approach will be used to develop predictor functions for other types of running buffer if needed., (© 2020 WILEY-VCH Verlag GmbH & Co. KGaA, Weinheim.)

Application of hollow fiber-based electromembrane extraction was studied for extraction and quantification of phenytoin from exhaled breath condensate (EBC). Phenytoin is extracted from EBC through a supported liquid membrane consisting of 1-octanol impregnated in the walls of a hollow fiber, and into an alkaline aqueous acceptor solution inside the lumen of the fiber. Under the obtained conditions of electromembrane extraction, that is, the extraction time of 15 min, stirring speed of 750 rpm, donor phase pH at 11.0, acceptor pH at 13.0, and an applied voltage of 15 V across the supported liquid membrane, an enrichment factor of 102-fold correspond to extraction percent of 25.5% was achieved. Good linearity was obtained over the concentration range of 0.001-0.10 µg/mL (r 2 = 0.9992). Limits of detection and quantitation were 0.001 and 0.003 µg/mL, respectively. The proposed method was successfully applied to determine phenytoin from EBC samples of patients receiving the drug. No interfering peaks were detected that indicating excellent selectivity of the method. The intra- and interday precisions (RSDs) were less than 14%., (© 2020 WILEY-VCH Verlag GmbH & Co. KGaA, Weinheim.)

In this paper, the development of a simple dilute-and-shoot method for quantifying urinary creatinine by CE-ESI-MS was described. The creatinine analysis time was about 7 min/sample by conventional single injection (SI) method and can be significantly reduced to less than 2 min/sample with multi-segment injection (MSI). In addition, the standard addition analysis of 5-hydroxyindole-3-acetic acid (5-HIAA) and creatinine normalization was performed within one run by the MSI technique, and the total analysis time was 14-min faster compared to the SI method for analyzing the same set of samples. The uses of isotopic and non-isotopic internal standards (ISs) were compared. Creatinine-(methyl- 13 C) and 5-hydroxyindole-4,6,7-D 3 -3-acetic-D 2 acid (5-HIAA-D 5 ) used as isotopic ISs can provide both accurate and precise results. In contrast, 1,5,5-trimethylhydantoin (1,5,5-TH) used as the non-isotopic IS for creatinine may cause a bias of over 13% in SI method and even worse when the MSI technique was used. Another compound, 2-methyl-3-indoleacetic acid (2-MIAA), was determined not suitable for MSI analysis of 5-HIAA due to endogenous interferences despite its acceptable performance in conventional methods of analysis., (© 2019 WILEY-VCH Verlag GmbH & Co. KGaA, Weinheim.)

The review presents a comprehensive survey of recent developments and applications of high performance capillary and microchip electroseparation methods (zone electrophoresis, isotachophoresis, isoelectric focusing, affinity electrophoresis, electrokinetic chromatography, and electrochromatography) for analysis, micropreparation, and physicochemical and biochemical characterization of peptides since 2017 up to about the middle of 2019. Progress in the study of electromigration properties of peptides and in the methodology of their analysis (sample preseparation, preconcentration and derivatization, adsorption suppression, EOF control, and detection) are described. Advances in CE and CEC methods are demonstrated and their applications in the following areas are presented: qualitative and quantitative analysis, determination in complex (bio)matrices, monitoring of chemical and enzymatical reactions and physical changes, amino acid, sequence and chiral analysis, and peptide mapping of proteins. In addition, micropreparative separations and determinations of important physicochemical characteristics of peptides by CE and CEC methods are reported., (© 2019 WILEY-VCH Verlag GmbH & Co. KGaA, Weinheim.)

We present a method for the speciation of chromium by capillary electrophoresis. Cr(III) was complexed with diethylenetriaminepentaacetic acid (DTPA) to form a negatively charged complex. Using 20 mM phosphate buffer of pH 8 containing 0.5 mM tetradecyltrimethylammonium hydroxide (TTAOH) at a separation voltage of -15 kV, both forms of chromium CrDTPA(2-) and CrO(4) (2-) were separated in less than 6 min. Direct UV detection at 214 nm was used. The effect of the presence of interfering ions was investigated. The application of the developed method to speciation of chromium in tanning liquor is demonstrated. The obtained results have shown a good correlation with those of flame atomic absorbance spectrometry (FAAS), inductively coupled plasma-mass spectrometry (ICP-MS) and UV/VIS spectrophotometry.

The growing awareness of the strong development of the toxicity of heavy metals upon their chemical forms has led to an increasing interest in the qualitative and quantitative determination of specific metal species. Speciation has therefore become an important topic of present-day analytical research. The development in the elemental speciation analysis by capillary electrophoresis (CE) is reviewed. Various CE separation modes and detection techniques applied are discussed. A comprehensive description of reported methods to date in CE speciation analysis including metals, metalloids and nonmetallic elements is demonstrated. Some examples are presented to demonstrate CE's ability to solve real-world speciation analysis with emphasis on the applications in biological and environmental samples. Further, some issues concerning the limitations and the future of CE with regard to speciation studies are also discussed.