Capillary Electrophoresis Device Research Articles

Dextrin-assisted gel electromembrane extraction of chiral drugs: Improving the extraction efficiency and investigation of enantioselectivity of extraction

The present study investigates the use of dextrins (maltodextrin, β-cyclodextrin, and hydroxypropyl-β-cyclodextrin) to improve the efficiency of the agarose-based gel electromembrane extraction technique for extracting chiral basic drugs (citalopram, hydroxyzine, and cetirizine). Additionally, it examines the enantioselectivity of the extraction process for these drugs. To achieve these, dextrins were incorporated into either the sample solution, the membrane, or the acceptor solution, and then the extraction procedure was performed. Enantiomers were separated and analyzed using a capillary electrophoresis device equipped with a UV detector. The results obtained under the optimal extraction conditions (sample solution pH: 4.0, acceptor solution pH: 2.0, gel membrane pH: 3.0, agarose concentration: 3 % w/v, stirring rate: 1000 rpm, gel thickness: 4.4 mm, extraction voltage: 62.3 V, and extraction time: 32.1 min) indicated that incorporating dextrins into either the sample solution, membrane or the acceptor solution enhances extraction efficiency by 17.3–23.1 %. The most significant increase was observed when hydroxypropyl-β-cyclodextrin was added to the acceptor solution. The findings indicated that the inclusion of hydroxypropyl-β-cyclodextrin in the sample solution resulted in an enantioselective extraction, yielding an enantiomeric excess of 6.42–7.14 %. The proposed method showed a linear range of 5.0–2000 ng/mL for enantiomers of model drugs. The limit of detection and limit of quantification for all enantiomers were found to be < 4.5 ng/mL and <15.0 ng/mL, respectively. Intra- and inter-day RSDs (n = 4) were less than 10.8 %, and the relative errors were less than 3.2 % for all the enantiomers. Finally, the developed method was successfully applied to determine concentrations of enantiomers in a urine sample with relative recoveries of 96.8–99.2 %, indicating good reliability of the developed method.

Using capillary electrophoresis to identify Anopheline species in routine sampling sites.

In the Anopheles genus, various mosquito species are able to transmit the Plasmodium parasites responsible for malaria, while others are non-vectors. In an effort to better understand the biology of Anopheles species and to quantify transmission risk in an area, the identification of mosquito species collected in the field is an essential but problematic task. Morphological identification requires expertise and cannot be checked after processing samples in a destructive treatment, while sequencing of numerous samples is costly. Here, we introduce a method of Species identification via Simple Observation Coupled with Capillary Electrophoresis Technology (SOCCET). This molecular technique of species identification is based on precise determination of ITS2 length combined with a simple visual observation, the colour of mosquito hindleg tip. DNA extracted from field-collected Anopheles mosquitoes was amplified with universal Anopheles ITS2 primers and analysed with a capillary electrophoresis device, which precisely determines the size of the fragments. We defined windows of amplicon sizes combined with fifth hind tarsus colour, which allows discrimination of the major Anopheles species found in our collections. We validated our parameters via Sanger sequencing of ITS2 amplicons. Using the SOCCET method, we characterised the composition of Anopheles populations in five locations of French Guiana, where we detected a total of nine species. Anopheles braziliensis and Anopheles darlingi were detected in four locations each and represented 13 and 67% of our samples, respectively. The SOCCET method can be particularly useful when working with routine sampling sites with a moderate species diversity, that is, when the number of local species is too high to define species-specific primers but low enough to avoid individual ITS2 sequencing. This tool will be of interest to evaluate local malaria transmission risk and this approach may be further implemented for other mosquito genera.

Improvement of limit of detection in primer extension-based multiplexed mutation assay using capillary electrophoresis

One of the challenges in liquid biopsy for early cancer detection is ascribed to the fact that mutation DNA often represents an extremely small ratio of less than 1% compared to wild-type genes in blood. However, in conventional fragment analysis with capillary electrophoresis (CE), the detectable allele frequency could be about 5%. In this work, we developed an original reagent-based fragment analysis with single base extension (SBE) reactions for cancer-associated mutation assay using a commercially available CE device, and investigated on a possibility of improvement of limit of detection (LOD) for genetic mutation. First, after adjustment of reagent conditions for the SBE reactions, the linear relationship between gene template concentration and fluorescence intensity was obtained from 1 to 100 fmol of target genes. Next, from the results of an experiment to detect mutation EGFR L858R at abundance ratios of mutant type to wild type (100-fmol template) of 0, 1, 5, and 10%, it was shown that the target gene can be detected with LOD of 0.33%. This high sensitivity was realized in part by separating fluorescently labeled substrates into an individual tube for an each-colored SBE reaction. Moreover, mutations EGFR L858R and KRAS G12V were simultaneously detected at sensitivities equivalent to LODs of 0.57 and 0.47%, respectively. These results indicate that < 1% of mutations in multiplex gene mutations can be simultaneously detected, and that possibility suggests that the developed method can be used in clinical practice for detecting cancers.Graphical abstract

Developmental validation of a complementary Y-STR system for the amplification of forensic samples

In this study, a new complementary Y-STR system that includes 31 loci was developed (DYS522, DYS388, DYF387S1a/b, DYS510, DYS587, DYS645, DYS531, DYS593, DYS617, GATA_A10, DYS622, DYS552, DYS508, DYS447, DYS527a/b, DYS446, DYS459a/b, DYS444, DYS557, DYS443, DYS626, DYS630, DYS526a, DYF404S1a/b, DYS520, DYS518, and DYS526b). This 31-plex Y-STR system, SureID® Y-comp, is designed for biological samples from forensic casework and reference samples from forensic DNA database. To validate the suitability of this novel kit, many developmental works including size precision testing, sensitivity, male specificity testing, species specificity, PCR inhibitors, stutter precision, reproducibility, suitability for use on DNA mixture and parallel testing of different capillary electrophoresis devices were performed. Mutation rates were investigated using 295 DNA-confirmed father-son pairs. The results demonstrate that the SureID® Y-comp Kit is time-efficient, accurate, and reliable for various case-type samples. It possessed a higher discrimination power and can be a stand-alone kit for male identification. Moreover, the simply acquired additional Y-STR loci will be conductive to construct a robust database. Even if various commercial Y-STR kits are used in distinct forensic laboratories, a wider trans-database retrieval will become feasible with the effort of the SureID® Y-comp Kit.

Molecular characterisation of Hb Akron [β52 (D3) Asp→Val] combined with thalassaemia in a Chinese family

AimsHb Akron (HBB:c.158A>T) is a rare β-chain variant and many characteristics about its clinical features still remain unclear. In this study, we aimed to explore the molecular and haematological characterisations...

Using Multi-Material Fused Deposition Modeling (Fdm) for One-Step 3d Printing of a Microfluidic Capillary Electrophoresis Device with Integrated Electrodes for Capacitively Coupled Contactless Conductivity Detection

Using Multi-Material Fused Deposition Modeling (Fdm) for One-Step 3d Printing of a Microfluidic Capillary Electrophoresis Device with Integrated Electrodes for Capacitively Coupled Contactless Conductivity Detection

Two Original Experimental Setups for Staircase Frontal Affinity Chromatography at the Miniaturized Scale.

Frontal affinity chromatography is a powerful, underappreciated technique for the qualitative (screening) and quantitative (Kd determination) evaluation of biological interactions. Its development has been previously hampered by its sample consumption, limited throughput, and lack of dedicated instrumentation especially at a miniaturized scale. This work describes two original experimental devices allowing nano-frontal affinity chromatography titrations (nano-FAC) to be automatically implemented in the time-saving staircase mode. The first nano-FAC system utilizes a capillary electrophoresis device (7100 CE Agilent system) in the pressurization mode with in situ UV detection. The second nano-FAC experimental setup implements a nano-LC device (Ultimate 3000 Thermo) modified with a 10-port valve equipped with two superloops (loop volume, 5 μL) operating alternatively and automatically in a single run. The benefits and drawbacks of each approach are exemplified using two model protein-ligand interactions (concanavalin A-mannose and concanavalin A-glucose). The two methods result in concordant dissociation constants (Kd) and number of active site (Bact) values, obtained in a fully automated manner, with low sample consumption and good throughput.

Determination of two quaternary ammonium salts in disinfector by portable capillary electrophoresis device based on smartphone

现有的小型毛细管电泳(CE)装置仍采用平板或计算机进行数据处理和分析,其便携性仍存在明显不足。针对这一问题,发展了一种基于智能手机的CE装置,实现了真正的便携式定量分析。该装置集成了电容耦合式非接触式电导检测(C4D)和蓝牙通信功能,并提供了手机界面软件。通过手机界面软件,不仅可以控制CE装置的电泳运行,还可以实时接收C4D检测器发出的数据信息,显示电泳图谱和进行数据处理。该装置尺寸为20 cm×20 cm×15 cm,重量为2 kg。为了验证所设计装置的性能,采用季铵盐(QAs)消毒剂(十二烷基二甲基苄基溴化铵(DDBAB)和十二烷基三甲基溴化铵(DTAB))作为分析对象。实验数据表明,DDBAB和DTAB线性范围分别为20~1000和30~1000 μmol/L,线性回归方程的相关系数(R2)分别为0.9995和0.9989,检出限(LOD)分别为10和13 μmol/L,日内相对标准偏差(RSD, n=3)分别为1.9%和2.7%。另外实验对DDBAB和DTAB混合离子液进行了测试,在8 min内可实现基线分离。最后,对现场使用的新洁尔灭消毒液中QAs进行了加标回收试验,DDBAB和DTAB的回收率分别为100.5%~101.5%和96.2%~99.3%。研究结果表明,所开发CE装置具有线性好、LOD低、重复好、准确性高,尤其便携好等优点,可用于消毒液中QAs现场定量检测。

Assessment of 13 Single Nucleotide Polymorphisms Loci for Identification in Forensic Sciences for Turkish Population

Single nucleotide polymorphisms (SNPs) are used for genetic identification in forensic science. In this study, 13 loci (rs876724, rs1357617, rs2046361, rs717302, rs917118, rs735155, rs901398, rs729172, rs740910, rs1493232, rs719366, rs1031825, rs722098) selected among 52 SNP plex’s whose importance in forensic sciences were determined before, were studied for the Turkish population. In addition to the discrimination power of the Turkish population as selected loci in this study, it was aimed to calculate the separation of powers showed in their Turkey geographic regions. For this purpose, blood samples taken from 50 volunteers' DNA samples were duplicated by polymerase chain reaction (PCR) and typed in a capillary electrophoresis device and genotype information was determined for 13 SNP loci. The data obtained showed parallelism with values published by the SNPforID Consortium. Obtained results proved the separation forces of the studied loci suitable for the Turkish population. The existence of Hardy-Weinberg Equation was tested for all studied loci and Hardy-Weinberg Equation was found in all loci except one loci (rs876724). Because of the fact that the number of samples is insufficient for geographical studies, no statistically significant difference was observed in other loci in the regional context except for the significant results obtained for rs729172 and rs740910 loci.

Design-Based Learning Framework for Introducing Factorial Experimental Design and Lab-on-a-Chip Separations in an Instrumental Chemistry Laboratory

Lab-on-a-chip devices are a critical and emerging chemical technology. New pedagogical tactics that effectively capture both analytical chemistry concepts along with the design elements unique to these devices are necessary for effective integration in the undergraduate chemistry curriculum. This article develops a design-based learning laboratory exercise to concurrently introduce analytical miniaturization, chemical separations, and formal factorial design of experiment methods in an Introduction to Instrumental Analysis course. A new student-accessible lab-on-a-chip fabrication platform using a benchtop laser cutter allows for rapid, iterative, and safe production of student-designed microchip capillary electrophoresis devices in minutes. Student teams used a full factorial experimental design approach to efficiently screen three experimental variables of their choice when evaluating the separation speed of their own lab-on-a-chip devices. This laboratory exercise has been successfully implemented on three occasions over the past several years at Colorado College. Both reflective and performance-based assessment methods suggest this exercise is an effective pedagogical approach for realizing the intended learning outcomes.

Study of antagonism between some intestinal bacteria with high-speed micellar electrokinetic chromatography.

In this work, high-speed micellar electrokinetic chromatography with LIF detection was applied to study the antagonism between three intestinal bacteria, Escherichia coli (E. coli), Bacillus licheniformis (B. licheniformis) and Bacillus subtilis (B. subtilis). The fluorescent derivatization for the bacteria was performed by labeling the bacteria with FITC. In a high-speed capillary electrophoresis (HSCE) device, the three bacteria could be completely separated within 4min under the separation mode MEKC. The BGE was 1 × TBE containing 30mM SDS and 1.5×10-5 g/mL polyethylene oxide. The limits of detection for E. coli, B. licheniformis and B. subtilis were 2.80×106 CFU/mL, 1.60×106 CFU/mL and 1.90×106 CFU/mL respectively. Lastly, the method was applied to investigate the antagonism between the three bacteria. The bacteria were mixed and cultured for 7 days. The samples were separated and determined every day to study the interaction between bacteria. The results showed that B. licheniformis and B. subtilis could not inhibit each other, but they could effectively inhibit the reproduction of E. coli. The method developed in this work was quick, sensitive and convenient, and it had great potential in the application of antagonism study for bacteria.

Pre-Concentration and Analysis of Mycotoxins in Food Samples by Capillary Electrophoresis.

Mycotoxins are considered one of the most dangerous agricultural and food contaminants. They are toxic and the development of rapid and sensitive analytical methods to detect and quantify them is a very important issue in the context of food safety and animal/human health. The need to detect mycotoxins at trace levels and to simultaneously analyze many different mycotoxin types became mandatory to protect public health. In fact, European Commission regulations specified both their limits in foodstuffs and official sample preparation protocols in addition to analytical methods to verify their presence. Capillary Electrophoresis (CE) includes different separation modes, allowing many versatile applications in food analysis and safety. In the context of mycotoxins, recent advances to improve CE sensitivity, particularly pre-concentration techniques or miniaturized systems, deserve remarkable attention, as they provide an interesting approach in the analysis of such contaminants in complex food matrices. This review summarizes the applications of CE combined with different pre-concentration approaches, which have been proposed in the literature (mainly) in the last ten years. A section is also dedicated to recent microchip–CE devices since they represent the most promising CE mode for this application.

Fabrication of high-quality glass microfluidic devices for bioanalytical and space flight applications

Microfabricated glass microfluidic and Capillary Electrophoresis (CE) devices have been utilized in a wide variety of applications over the past thirty years. At the Berkeley Space Sciences Laboratory, we are working to further expand this technology by developing analytical instruments to chemically explore our solar system. This effort requires improving the quality and reliability of glass microfabrication through quality control procedures at every stage of design and manufacture. This manuscript provides detailed information on microfabrication technology for the production of high-quality glass microfluidic chips in compliance with industrial standards and space flight instrumentation quality control.•The methodological protocol provided in this paper includes the scope of each step of the manufacturing process, materials and technologies recommended and the specific challenges that often confront the process developer.•Types and sources of fabrication error at every stage have been identified and their solutions have been proposed and verified.•We present robust and rigorous manufacturing and quality control procedures that will assist other researchers in achieving the highest possible quality glass microdevices using the latest apparatus in a routine and reliable fashion.

A three-dimensional printed electromembrane extraction device for capillary electrophoresis

A concentric electromembrane extraction preconcentration device was designed and fabricated using fused deposition modelling 3D printing. The device consisted of a hemispherical electrode sample vial printed from a filament of conductive polylactic acid (PLA), into which sat a smaller hemispherical 3D printed porous membrane acceptor vial. Application of voltage between a point-electrode placed in the center of 20 μL solution inside the acceptor vial and the electrode vial containing 1 mL of sample, enabled anion migration through the 3D printed porous material into the acceptor solution. Electromembrane extraction was proved using fluorescein for imaging of the extraction process, with preconcentration rates of 0.833 μM/sec at 120 V with close to 95% recovery. The performance of the fabricated porous 3D printed device was evaluated for the preconcentration of anions from water prior to capillary electrophoresis detection. Preconcentration factors ranging between 36–44 were obtained for chloride, nitrate, perchlorate and sulfate, while a lower performance was observed for weaker acids as fluoride and phosphate (3–4). The limit of detection (LOD) was determined to be 0.16 μM, 0.18 μM and 0.18 μM for NO3−, ClO4− and SO42- respectively. The extraction and quantitation of ions from a soil slurry without filtration, namely NO3− and SO42- content was determined to be 0.24 and 0.03 mmol/100 g of soil, respectively which are values in the range of those typically reported in soil samples.

A novel thread-based microfluidic device for capillary electrophoresis with capacitively coupled contactless conductivity detection

This work describes a novel thread-based microfluidic device to perform capillary electrophoresis separation and capacitively coupled contactless conductivity detection. Polyester threads with a diameter of about 315 μm were assembled (stretched) in a 3D-printed platform containing solution reservoirs and platinum electrodes. Separation of potassium, sodium, and lithium ions was achieved within 1 min with good peak resolutions and limits of detection close to 1 μmol L−1. Inversion of the electroosmotic flow direction by adding a cationic surfactant in the running electrolyte was demonstrated. The applicability of the thread-based microfluidic devices was successfully evaluated by the determination of sodium and potassium ions in commercial samples of diet soft drinks.

A bi-end injection capillary electrophoresis method for simultaneous determination of 37 cations and anions in beers.

Capillary electrophoresis (CE) is excellent at separating all the ions in a sample but is rarely used as a result of its detection issue and easy loss of very fast ions by common one-end injection methods. Herein we propose a newly developed method aimed at simultaneous determination of positive and negative ions with a home-made CE device, featuring bi-end injection and contactless conductivity detection at the middle. By simply using 2.5M acetic acid as a running buffer, the method can separate 37 ions (3 inorganic anions, 8 inorganic cations, 10 biogenic amines, and 16 amino acids) per run, with linearity between 10 and 2000μM (R2 > 0.99), limit of detection of 1.0-16.6μM, and limit of quantification of 2.3-31.7μM. The recovery measured by spiking standards into samples at high, middle, and low levels was between 73% and 110%. The intra- and interday repeatability of the 37 analytes ranged from 0.69% to 8.97% and from 0.68% to 11.04%, respectively. The proposed method was evaluated by analysis of 21 beers and, in addition to acquiring the concentration information, the brands of the tested beers were distinguished. This method is of high throughput, fast, and cost-effective. It could be a promising tool for ionomic analysis. Graphical abstract ᅟ.

Determination of inorganic cations in biological fluids using a hybrid capillary electrophoresis device coupled with contactless conductivity detection

We describe the assembly of a hybrid electrophoresis device that contains fused silica capillaries interconnected to a microfabricated interface in a cross format for the determination of inorganic cations in biological samples. The sample transport in the proposed hybrid device was performed under gated injection mode and the separations were monitored with a capacitively coupled contactless conductivity detector. The capillary extremities were inserted into polypropylene tubes to create solution reservoirs. Sensing electrodes were produced using stainless-steel hypodermic needles previously cut with 2.0mm length. The running composition and injection time were optimized and the best results were found using 50mmol/L lactic acid, 20mmol/L histidine and 3mmol/L 18-crown-6 ether, and an electrokinetic injection time of 15s. The separation of six inorganic cations was achieved with baseline resolution, and efficiencies were between 9.1×103 and 5.4×104 plates/m. The proposed hybrid device was explored for determining the concentration levels of inorganic cations in urine, saliva, and tear samples, employing Li+ as an internal standard. The achieved results were in good agreement with the data reported in the literature. The reliability of the proposed method ranged from 93 to 98%, thus suggesting satisfactory accuracy for bioanalytical applications.

Chromatographic behavior of selected dyes on silica and cellulose micro-TLC plates: Potential application as target substances for extraction, chromatographic, and/or microfluidic systems

ABSTRACTThe main goal of this research communication focuses on screening of chromatographic behavior of 18 colorants including amaranth, black PN, bromophenol blue, bromocresol green, bromocresol purple, bromothymol blue, carmine, dimethyl yellow, erythrosine, fluorescein, methyl red, naphthalene black, patent blue, phenol red, Sudan II, Sudan III, Sudan IV, and p-xylenol blue. They are commonly used as food and industrial colorants or sensing molecules for analytical and biomedical applications. We studied retention (RF), peak base width (Wb), and peak asymmetry factor (AS) data collected under thermostatic conditions (303 K) using silica and cellulose-coated microplates (5 × 5 cm) and simple as possible mono/binary eluents. Presented retention database may help to select the proper conditions for, for example, chromatographic separation, extraction to solid phase (e.g. bar-type extraction devices) or application of such chemicals as the internal standard substances for planar/column chromatography and/or microfluidic devices (e.g., microfluidic paper-based analytical devices—μPADs). Furthermore, the microplates peak capacity (maximum number of resolvable spots, n) was calculated and discussed. As the second goal, we presented results of literature concerning dyes quantification in complex samples. We compared latest methodologies involving simple spectrophotometric analysis and more sophisticated protocols like high-throughput separation/detection procedures based on high-performance planar/liquid chromatography, capillary electrophoresis and microfluidic devices.

Integrating Internal Standards into Disposable Capillary Electrophoresis Devices To Improve Quantification.

To improve point-of-care quantification using microchip capillary electrophoresis (MCE), the chip-to-chip variabilities inherent in disposable, single-use devices must be addressed. This work proposes to integrate an internal standard (ISTD) into the microchip by adding it to the background electrolyte (BGE) instead of the sample—thus eliminating the need for additional sample manipulation, microchip redesigns, and/or system expansions required for traditional ISTD usage. Cs and Li ions were added as integrated ISTDs to the BGE, and their effects on the reproducibility of Na quantification were explored. Results were then compared to the conclusions of our previous publication which used Cs and Li as traditional ISTDs. The in-house fabricated microchips, electrophoretic protocols, and solution matrixes were kept constant, allowing the proposed method to be reliably compared to the traditional method. Using the integrated ISTDs, both Cs and Li improved the Na peak area reproducibility approximately 2-fold, to final RSD values of 2.2–4.7% (n = 900). In contrast (to previous work), Cs as a traditional ISTD resulted in final RSDs of 2.5–8.8%, while the traditional Li ISTD performed poorly with RSDs of 6.3–14.2%. These findings suggest integrated ISTDs are a viable method to improve the precision of disposable MCE devices—giving matched or superior results to the traditional method in this study while neither increasing system cost nor complexity.

Analysis of Methanol in the Presence of Ethanol, Using a Hybrid Capillary Electrophoresis Device with Electrochemical Derivatization and Conductivity Detection.

Concurrently with ethanol, many other compounds can be formed during the fermentation of grains and fruits. Among those, methanol is particularly important (because of its toxicity) and is typically formed at concentrations much lower than ethanol, presenting a particular challenge that demands the implementation of separation techniques. Aiming to provide an alternative to traditional chromatographic approaches, a hybrid electrophoresis device with electrochemical preprocessing and contactless conductivity detection (hybrid EC-CE-C4D) is herein described. The device was applied to perform the electro-oxidation of primary alcohols, followed by the separation and detection of the respective carboxylates. According to the presented results, the optimum conditions were obtained when the sample was diluted with 2 mmol L-1 HNO3 and then electro-oxidized by applying a potential of 1.4 V for 60 s. The oxidation products were then electrokinetically injected by applying a potential of 3 kV for 4 s and separated using a potential of 3 kV and a background running electrolyte (BGE) consisting of 10 mmol L-1 N-cyclohexyl-2-aminoethanesulfonic acid (CHES) and 5 mmol L-1 sodium hydroxide (NaOH). n-Propanol was used as an internal standard and the three carboxylate peaks were resolved with baseline separation within <3 min, defining linear calibration curves in the range of 0.10-5.0 mmol L-1. Limits of detection (LODs) of 20, 40, and 50 μmol L-1 were obtained for ethanol, n-propanol, and methanol, respectively. To demonstrate the applicability of the proposed strategy, a laboratory-made sample (moonshine) was used. Aliquots collected along the beginning of the fractional distillation presented a decreasing methanol ratio (from 4% to <0.5%) and a growing ethanol ratio (from 80% to 100%) in the collected volume.