Review on the development of truly portable and in-situ capillary electrophoresis systems

Capillary electrophoresis (CE) is a technique which uses an electric field to separate a mixed sample into its constituents. Portable CE systems enable this powerful analysis technique to be used in the field. Many of the challenges for portable systems are similar to those of autonomous in-situ analysis and therefore portable systems may be considered a stepping stone towards autonomous in-situ analysis. CE is widely used for biological and chemical analysis and example applications include: water quality analysis; drug development and quality control; proteomics and DNA analysis; counter-terrorism (explosive material identification) and corrosion monitoring. The technique is often limited to laboratory use, since it requires large electric fields, sensitive detection systems and fluidic control systems. All of these place restrictions in terms of: size, weight, cost, choice of operating solutions, choice of fabrication materials, electrical power and lifetime. In this review we bring together and critique the work by researchers addressing these issues. We emphasize the importance of a holistic approach for portable and in-situ CE systems and discuss all the aspects of the design. We identify gaps in the literature which require attention for the realization of both truly portable and in-situ CE systems

Development of an autonomous lab-on-a-chip system with ion separation and conductivity detection for river water quality monitoring

This thesis discusses the development of a lab on a chip (LOC) ion separation for river water quality monitoring using a capacitively coupled conductivity detector (C⁴D) with a novel baseline suppression technique.Our first interest was to be able to integrate such a detector in a LOC. Different designs (On-capillary design and on-chip design) have been evaluated for their feasibility and their performances. The most suitable design integrated the electrode close to the channel for an enhanced coupling while having the measurement electronics as close as possible to reduce noise. The final chip design used copper tracks from a printed circuit board (PCB) as electrodes, covered by a thin Polydimethylsiloxane (PDMS) layer to act as electrical insulation. The layer containing the channel was made using casting and bonded to the PCB using oxygen plasma. Flow experiments have been conduced to test this design as a detection cell for capacitively coupled contactless conductivity detection (C⁴D).The baseline signal from the system was reduced using a novel baseline suppression technique. Decrease in the background signal increased the dynamic range of the concentration to be measured before saturation occurs. The sensitivity of the detection system was also improved when using the baseline suppression technique. Use of high excitation voltages has proven to increase the sensitivity leading to an estimated limit of detection of 0.0715 μM for NaCl (0.0041 mg/L).The project also required the production of an autonomous system capable of operating for an extensive period of time without human intervention. Designing such a system involved the investigation of faults which can occur in autonomous system for the in-situ monitoring of water quality. Identification of possible faults (Bubble, pump failure, etc.) and detection methods have been investigated. In-depth details are given on the software and hardware architecture constituting this autonomous system and its controlling software

High-voltage capacitively coupled contactless conductivity detection for conventional and microchip capillary electrophoresis

This thesis focuses on the optimisation of capacitively coupled contactless conductivity detection for capillary and microchip electrophoresis and its applications in analytical chemistry. First, the effect of high excitation voltages and operation frequencies on the capacitively coupled contactless conductivity detector cell for conventional capillary electrophoresis is evaluated. The detector electrodes comprised two steel tubes cut from hypodermic needles, through which the capillaries were inserted. It is demonstrated that increasing excitation voltages from 25 V pp, to 250 Vpp improves the detection limits by a factor of 10. The high actuator voltage approach was also investigated for contactless conductivity detection on a glass-microchip device with ancm long channel. The detector electrodes formed part of the microchip and were placed on the microchip directly above the microchannel. In a separate project the simplification of on-microchip contactless conductivity detection was accomplished. This was achieved by integrating the detector electrodes on to a chip-holder specifically designed for this purpose. Thus the electrodes were a part of the holder, an improvement of the previous arrangement whereby the detector electrodes were situated on the microdevice. Finally the applications and advantages of the optimised high-voltage capacitively coupled contactless conductivity detection for inorganic and organic analysis were demonstrated. The separation and detection of 14 metal ions was accomplished in less than six minutes. The compatibility of this detector with non-UV transparent, polymer capillaries has been demonstrated. The detection of native amino acids has been evaluated. Part of the work was dedicated to the on-chip analysis of various classes of organic ions. The two immunoproteins human immunoglobulin M (IgM) and immunoglobulin G (IgG), were analysed in their unlabelled state on both capillary and lab-on-chip platforms. All species involved in an immunological interaction between IgM and IgG could be detected. A method for the analysis of selected basic pharmaceutical drug substances was developed. Detection limits comparable to those supplied by direct UV detection were obtained. Main component assays of selected pharmaceutical preparations have been demonstrated

Monitoring of enzymatic reactions using capillary electrophoresis with conductivity detection

Capillary electrophoresis combined with contactless conductivity detection allows to separate and detect the ionic species, which are neither UV absorbing nor fluorescent. This thesis focuses on the applications of this method on enzymatic reactions in different analytical tasks. First, the non-ionic species ethanol, glucose, ethyl acetate and ethyl butyrate were made accessible for analysis by capillary electrophoresis via charged products or byproducts obtained in enzymatic conversions using hexokinase, glucose oxidase, alcohol dehydrogenase and esterase. The conversion of glucose and that of ethylacetate were also successfully demonstrated on a microchip-device. Quantification of ethyl acetate, was found possible with a detection limit of approximately 7 μM. Then the model of urea catalyzed by urease was chosen for the study of the enzymatic mechanism, the effect of substrate concentration and pH value as well as the Michaelis-Menten constant. The determination of urea in human blood as clinical application of this enzymatic reaction was tested. 10 human blood samples were collected from a hospital and analyzed. The results were comparable with the established methods. The method was then extended to proteome analysis; identification of proteins is generally achieved through proteolytic digestion with enzymes such as pepsin and trypsin. Protein digestion with pepsin and trypsin was successfully monitored by capillary electrophoresis. Minigastrin I, myoglobin, cytochrome C, human serum albumin and bovine serum album were the model proteins digested by pepsin, cytochrome C and myoglobin were the model proteins digested by trypsin. Electrophoretically mediated micro-analysis (EMMA) technique is employed for the tryptic digestion of cytochrome C and apomyoglobin. Finally, the enantioselective hydrolysis of esters of amino acids with lipase was monitored. Porcine pancreas lipase was found to have a better efficiency on hydrolysis and enantioselectivity than wheat germ lipase. L-threonine methyl ester demonstrated stronger enantioselectivity than L-serine methyl ester. Acetylcholinesterase inhibitors can be used as drug against Alzheimer disease or nerve agents. Three compounds, namely galantamine, paraoxon and Huperzine-A, were the model inhibitors to study the behavior and kinetics of the inhibitors. The values of IC50 were obtained through graphical plot. Their dependence on the time course was monitored and graphically illustrated

Optimization of Tetrapolar Impedance Electrodes in Microfluidic Devices for Point of Care Diagnostics using Finite Element Modeling

Electrophoresis is widely applied in the field of biochemistry and molecular biology. Tetrapolar electrical impedance sensing (TEIS) has been shown capable of replacing the conventional detection technology in order to develop a point of care electrophoretic analyzer. Besides the advantages of reduced influence of electrode polarization, TEIS is affected by sensitivity distribution depending on the electrode design. A well reported practice outside of electrophoresis, systematic investigation of the effects of sensitivity distribution on the TEIS in microfluidic devices has not been conducted. Here we utilize finite element modeling, backed by experimental results, to optimize the sensor design within an electrophoretic separation device. Numerous sensor designs were validated regarding detectability, sensitivity and spatial resolution. The results show, that minimizing the distance between the central/pick-up electrodes increases sensitivity and spatial resolution whereas the distance between the central electrodes and the outer electrode do not influence sensitivity and spatial resolution

Contactless conductivity detection for analytical techniques: Developments from 2016 to 2018

The publications concerning capacitively coupled contactless conductivity detection for the 2-year period from mid-2016 to mid-2018 are covered in this update to the earlier reviews of the series. Relatively few reports on fundamental investigations or new designs have appeared in the literature in this time interval, but the development of new applications with the detection method has continued strongly. Most often, contactless conductivity measurements have been employed for the detection of inorganic or small organic ions in conventional capillary electrophoresis, less often in microchip electrophoresis. A number of other uses, such as detection in chromatography or the gauging of bubbles in streams have also been reported

Development of a microchip electrophoresis system for online monitoring of atmospheric aerosol composition

2011 Spring.Includes bibliographical references.Atmospheric aerosols are solid or liquid particles that remain suspended in the environment for an extended time because of their size. Due to their high number concentration, low mass concentration, unique size range, and high temporal and spatial variability, atmospheric aerosols represent a significant unknown in both environmental impact and human health. Despite the importance of aerosols, current instrumentation for monitoring their chemical composition is often limited by poor temporal resolution, inadequate detection limits, lack of chemical speciation, and/or high cost. To help address these shortcomings, microchip electrophoresis (MCE) has been introduced for the semi-continuous monitoring of water-soluble aerosol composition. The MCE instrument was coupled to a water condensation particle collector (growth tube), and the integrated system is termed Aerosol Chip Electrophoresis (ACE). ACE is capable of measuring particle composition with temporal resolution of 1 min and detection limits of ~100 ng m-3. This dissertation covers the development process of the prototype ACE instrument, including the novel separation chemistry, necessary modifications to traditional microfluidic devices, and the interface between the growth tube and the microchip

Selectivity enhancement in capillary electrophoresis – development of a two-dimensional separation and a dual detection system

For the analysis of complex samples such as biological or environmental ones, highly selective analytical strategies are required. The selectivity can be enhanced by means of hyphenation of orthogonal separation techniques or combination of complementary detectors as an alternative concept. In this work, an example for both concepts is presented, respectively. First, methodical and instrumental studies concerning the comprehensive hyphenation of the two most important instrumental techniques in ion analysis namely ion chromatography (IC) and capillary electrophoresis (CE) are described enabling the simultaneous determination of anions and cations. Further, the combination of capacitively coupled conductless conductivity detection (C4D) and mass spectrometry (MS) as dual detection concept for CE is introduced and applied using aqueous and non-aqueous background electrolyte systems. Another concept to enhance the selectivity is rendering the separation technique compatible to a highly selective detector such as MS. Thus, in a third part, an approach is presented enabling the coupling of surfactant-free microemulsion electrokinetic chromatography (SF-MEEKC) to MS. For the comprehensive hyphenation of capillary IC and CE, a modulator was developed ensuring well-controlled injections from the first to the second dimension. This was achieved by periodical injection of the IC effluent from a transfer capillary to the CE separation capillary by capillary batch injection. Important for the coupling was the compatibility of the two systems. Thus, the characteristics of the advanced capillary high performance IC with flow rates in the lower µL/min range facilitated the hyphenation being closer to CE conditions compared to IC with conventional columns. Due to the implemented capillary scale suppressor technology, the IC effluent consisted of analyte and pure water avoiding matrix concerned interferences and enabling exploitation of analyte stacking in CE. Further, fast CE measurements in the time range of seconds enabled the comprehensive coupling. Proof-of-concept measurements were performed using a model system containing nucleotides and their cyclic derivates. It was shown that the separation performance could be enhanced in the two-dimensional ICxCE-MS system compared to the single techniques. The work was further expanded to a methodical study concerning the simultaneous determination of positively and negatively charged analytes. A bypass system for IC was developed using a switching valve. This configuration enabled the cations bypassing the suppressor and thus they were not filtered out by the cation exchange membrane of the suppressor being not selective to eluent cations. Feasibility of the setup was demonstrated separating a model mixture of different arsenic species. The cations were eluted with water from the anion exchange column before driving a KOH gradient elution for the separation of anions. The injection parameters of the modulator introducing the IC effluent into the CE-MS system were studied taking into account the complex transport situation. It was found that the migration times were stable even for highly frequent injections with waiting times between the injections that were a factor four shorter than the migration time itself. Signal intensity of the MS was highest for positively charged species caused by the discriminating effect of the electrokinetic injection and the better ionization of the organic arsenic species by the electrospray ionization (ESI) source. Further, the coupling of two important detectors for CE is presented, namely capacitively coupled contactless conductivity detection (C4D) and electrospray ionization time-of-flight MS (ESI-TOF-MS). An experimental protocol was developed taking into account the requirements of the separation aspects and the compatibility with both detectors. ESI-TOF-MS requires background electrolytes consisting of volatile components such as ammonium acetate or formate. These, however, exhibit a rather high conductivity, which is disadvantageous for C4D. A 10 mM ammonium acetate/ammonia buffer was taken as compromise concerning the detection performance of both detectors. A sample containing various phenolic compounds serving as a model system was determined. The analytical characteristics showed the complementarity and suitability of this dual detection approach as C4D showed better response behavior towards m-cresol (limit of detection (LOD)=3.1 µM), while MS was more sensitive for determinations of m- and p-nitrophenol (LODs=0.8 µM) and 2,4-dinitrophenol (LOD=1.5 µM). The overall separation efficiency was excellent realizing the separation of counter-electroosmotic species with migration times of less than 60 s and illustrating that detector-induced band broadening could be neglected in the CE-C4D/MS system. In a second study, non-aqueous electrolytes were used for CE separations as they are highly compatible with both detectors due to their volatility and low background conductivity. A non-aqueous capillary electrophoresis (NACE)-C4D-MS method was developed using an acetonitrile based background electrolyte containing 2 M HAc and 4 mM NH4Ac enabling fast electrophoretic separations. Concentration changes of the background electrolyte and the choice of the inner diameter (ID) of the separation capillary were more critical for C4D than for MS. To choose a separation capillary with appropriate ID, the dependency of the sensitivity of the C4D on the ID was studied resulting in the use of a capillary with ID of 50 µm. The complementarity of the two detectors was demonstrated determining inorganic anions such as chloride, bromide, and nitrate (C4D) as well as organic biomolecules such as choline, thiamine, and acetylcholine (MS) simultaneously. A calibration was performed and the method applied on an extract of a food supplement quantifying the model analytes using the respective detector. MEEKC is a powerful tool for the separation of neutral species based on differences in their hydrophobic and hydrophilic properties. However, conventionally used SDS-based microemulsions are not compatible with ESI-MS. Enhancing the selectivity of the overall method, a surfactant-free microemulsion (SFME) based on water, ethanol, and 1-octanol was used as background electrolyte ensuring compatibility with ESI-TOF-MS. The small-angle X-ray scattering and dynamic light scattering measurements proved that the addition of ammonium acetate, which was necessary for the electrophoretic separation, was not altering aggregate formation in the SFME in the chosen concentration range. The separation performance of SF-MEEKC was demonstrated separating a model system consisting of hydrophobic and hydrophilic neutral vitamins, namely the vitamins B2 and D3, and the cationic vitamin B1 using UV/VIS detection. The influence of the ammonium acetate concentration on the separation performance was studied in detail. Characterization of the developed method was performed concerning reproducibility of migration times and peak areas and concerning the linearity of the calibration data. Further, the compatibility of SF-MEEKC with ESI-MS was shown determining the content of vitamin D3 in a commercial drug. Comparable sensitivity to aqueous CE-ESI-MS was achieved

Development of capillary electrophoresis with capacitively coupled contactless conductivity detection for clinical analysis and related applications

This thesis was focused on the development of capillary electrophoresis with capacitively coupled contactless conductivity detection (CE-C4D) for quantification of small organic ions that are poorly UV absorbing molecules in clinical analysis and related applications. CE-C4D is suitable for clinical analysis that usually involves small sample volumes in the nanolitre range. Due to the high separation efficiency obtained measuring the analyte ions in the presence of complex matrices is possible. The determination of uric acid was firstly examined to explore our knowledge for the routine compound analysis. We successfully determined uric acid in clinical samples in its anionic form using MES/His buffer. The results obtained were well correlated with the standard enzymatic method. We further investigated the determination of non-UV absorbing molecules, namely lactate, carnitine and acylcarnitines. CE-C4D proved to be suitable for the analysis of these compounds with little sample preparation and without any derivatization steps. Plasma lactate, which is an important parameter to evaluate the anaerobic metabolism and the exercise intensity in terms of the lactate threshold, was quantified. We successfully performed the quantification of plasma lactate in its anionic form using MES/His buffer. The plasma lactate concentrations at different exercise conditions and the lactate threshold curve were achieved. The results obtained by CE-C4D corresponded to the results observed by the enzymatic method. We also demonstrated the determination of carnitine and its derivatives (acylcarnitines) in various clinical samples. Free carnitine, short- and medium-chain acylcarnitines, consisting of acetyl-, propionyl-, isovaleryl-, hexanoyl-, octanoyl-, and valproyl-carnitines in plasma and urine samples were successfully quantified. The similarity of their chemical structures posed a high challenge in this study and required comprehensive optimization of the running buffer. An acetic acid buffer at pH 2.6 in the presence of hydroxypropyl-β- cyclodextrin (HP-β-CD) as a modifier was found to be the optimum. The isomers of valproyl- and octanoyl-carnitines could additionally be distinguished. The results obtained by CE-C4D were compared to standard enzymatic- and LC/MS-methods. In food analysis, the determination of lactic acid and carnitine in various food samples were also achieved. The determination of D- and L-lactic acid in fresh and spoiled milk and yogurt samples were successfully obtained in the presence of Tris/Maleic acid buffer using vancomycin as a chiral selector. In addition, the determination of carnitine in food samples was investigated in the presence of acetic acid buffer at pH 2.6. The carnitine contents in various samples were successfully quantified with good method reproducibility, and the results obtained were acceptable