High-voltage contactless conductivity detection of underivatized amino acids in capillary electrophoresis

High-voltage contactless conductivity detection of underivatized amino acids in both acidic and basic media is demonstrated. The suitability of different acidic buffer solutions at pH values of about 2.5 was investigated with 12 amino acids. Lactic acid as background electrolyte gave the best results in terms of detection limits for arginine, lysine and histidine, which were approximately 2 x 10(-7), 3 x 10(-7) and 4 x 10(-7) M, respectively. However, the sensitivity for other species was not quite as good and the detection limits in the order of 0.5-1 x 10 (-5) M. The use of basic conditions at pH 10-11 generally led to more stable baselines and more consistent sensitivities. A range of 20 amino acids was investigated with alkaline buffers and detection limits were typically about 10(-6) M. Urine and beer samples were analyzed. Nine and eleven amino acids could be identified, respectively.     

Determination of heavy metal ions by microchip capillary electrophoresis coupled with contactless conductivity detection

An integrated detection circuitry based on a lock-in amplifier was designed for contactless conductivity determination of heavy metals. Combined with a simple-structure electrophoresis microchip, the detection system is successfully utilized for the separation and determination of various heavy metals. The influences of the running buffer and detection conditions on the response of the detector have been investigated. Six millimole 2-morpholinoethanesulfonic acid + histidine were selected as buffer for its stable baseline and high sensitivity. The best signals were recorded with a frequency of 38 kHz and 20 V(pp). The results showed that Mn(2+), Cd(2+), Co(2+), and Cu(2+) can be successfully separated and detected within 100 s by our system. The detection limits for five heavy metals (Mn(2+), Pb(2+), Cd(2+), Co(2+), and Cu(2+)) were determined to range from about 0.7 to 5.4 μM. This microchip system performs a crucial step toward the realization of a simple, inexpensive, and portable analytical device for metal analysis.     

Considerations on contactless conductivity detection in capillary electrophoresis

Nearly all analyses by capillary electrophoresis (CE) are performed using optical detection, utilizing either absorbance or (laser-induced) fluorescence. Though adequate for many analytical problems, in a large number of cases, e.g., involving non-UV-absorbing compounds, these optical detection methods fall short. Indirect optical detection can then still provide an acceptable means of detection, however, with a strongly reduced sensitivity. During the past few years, contactless conductivity detection (CCD) has been presented as a valuable extension to optical detection techniques. It has been demonstrated that with CCD detection limits comparable, or even superior, to (indirect) optical detection can be obtained. Additionally, construction of the CCD around the CE capillary is straightforward and robust operation is easily obtained. Unfortunately, in the literature a large variety of designs and operating conditions for CCD were described. In this contribution, several important parameters of CCD are identified and their influence on, e.g., detectability and peak shape is described. An optimized setup based on a well-defined detection cell with three detection electrodes is presented. Additionally, simple and commercially available read-out electronics are described. The performance of the CCD-CE system was demonstrated for the analysis of peptides. Detection limits at the microM level were obtained in combination with good peak shapes and an overall good performance and stability.     

Capacitively coupled contactless conductivity detection in capillary electrophoresis

Capacitively coupled contactless conductivity detection (C(4)D) has become an accepted detection method in capillary electrophoresis (CE) for a variety of analytes. Advantages of this technique over optical detection modes and galvanic contact conductivity detection include great flexibility in capillary handling and rather simple mechanical parts and electronics, as it can be performed in an on-capillary mode. Furthermore, the detection principle can be used with capillaries made of other materials than fused silica (PEEK, Teflon), with chip-based separation technologies, or with capillaries having very small inner diameters. This review presents a discussion of the published literature on C(4)D for CE and capillary electrochromatography.     

Determination of heavy metal ions by capillary electrophoresis with contactless conductivity detection after field-amplified sample injection.

A method has been developed for determining of heavy metal ions by field-amplified sample injection capillary electrophoresis with contactless conductivity detection. The effects of the 2-N-morpholinoethanesulfonic acid/histidine (MES/His) concentration in the sample matrix, the injection time and organic additives on the enrichment factor were studied. The results showed that MES/His with a low concentration in the sample matrix, an increase of the injection time and the addition of acetonitrile improved the enrichment factor. Four heavy metal ions (Zn2+, Co2+, Cu2+ and Ni2+) were dissolved in deionized water, separated in a 10 mM MES/His running buffer at pH 4.9 and detected by contactless conductivity detection. The detection sensitivity was enhanced by about three orders of magnitude with respect to the non-stacking injection mode. The limits of detection were in the range from 5 nM (Zn2+) to 30 nM (Cu2+). The method has been used to determine heavy metal ions in tap water.     

Separation of enantiomers in capillary electrophoresis with contactless conductivity detection

Contactless conductivity detection is successfully demonstrated for the enantiomeric separation of basic drugs and amino acids in capillary electrophoresis (CE). Derivatization of the compounds or the addition of a visualization agent as for indirect optical detection schemes were not needed. Non-charged chiral selectors were employed, hydroxypropylated cyclodextrin (CD) for the more lipophilic basic drugs and 18-crown-6-tetracarboxylic acid (18C6H4) for the amino acids. Acidic buffer solutions based on lactic or citric acid were used. The detection limits were determined as 0.3 microM for pseudoephedrine as an example of a basic drug and were in the range from 2.5 to 20 microM for the amino acids.     

Miniaturized movable contactless conductivity detection cell for capillary electrophoresis

A miniaturized capacitively coupled contactless conductivity detector (mini-C(4)D) cell has been designed which is small enough to allow it to slide along the effective capillary length inside the capillary cassette of an Agilent capiillary electrophoresis system (CE) (or other CE brand of similar construction), including the possibility of positioning it close to the point of optical detection (4 cm), or even putting two such detector cells in one cassette. The cell was tested and the performance characteristics (noise, sensitivity, and peak width) were compared with those obtained with the previously used large C(4)D cell. No significant differences were observed. The mini-C(4)D was used in simultaneous separations of common cations and anions where its advantage over a larger C(4)D cell is the ability to vary the point of detection with the mini-C(4)D cell continuously at any point along the capillary length, so that the optimum apparent selectivity can be chosen. Other applications include providing a convenient second point of detection in addition to photometric detection, such as to measure accurately the linear velocity of a zone, or to allow placement of two mini-C(4)D cells in one capillary cassette simultaneously.     

Monosaccharide profiling of glycoproteins by capillary electrophoresis with contactless conductivity detection

Saccharides form one of the major constituents of biological macromolecules in living organisms. Many biological processes including protein folding, stability, immune response and receptor activation are regulated by glycosylation. In this work, we optimized a capillary electrophoresis method with capacitively coupled contactless conductivity detection for the separation of eight monosaccharides commonly found in glycoproteins, namely D-glucose, D-galactose, D-mannose, N-acetyl-D-glucosamine, N-acetyl-D-galactosamine, D-fucose, N-acetylneuraminic acid, and D-xylose. A highly alkaline solution of 50 mM sodium hydroxide, 22.5 mM disodium phosphate, and 0.2 mM CTAB (pH 12.4) was used as a background electrolyte in a 10 µm id capillary. To achieve baseline separation of all analytes, a counter-directional pressure of –270 kPa was applied during the separation. The limits of detection of our method were below 7 µg/ml (i.e., 1.5 pg or 1 mg/g protein) and the limits of quantification were below 22 µg/ml (i.e., 5 pg or 3 mg/g protein). As a proof of concept of our methodology, we performed an analysis of monosaccharides released from fetuin glycoprotein by acid hydrolysis. The results show that, when combined with an appropriate pre-concentration technique, the developed method can be used as a monosaccharide profiling tool in glycoproteomics and complement the routinely used LC-MS/MS analysis.     

Electromembrane extraction of heavy metal cations followed by capillary electrophoresis with capacitively coupled contactless conductivity detection

Electromembrane extraction (EME) was used as an off-line sample pre-treatment method for the determination of heavy metal cations in aqueous samples using CE with capacitively coupled contactless conductivity detection (CE-C(4) D). A short segment of porous polypropylene hollow fibre was penetrated with 1-octanol and 0.5%?v/v bis(2-ethylhexyl)phosphonic acid and constituted a low cost, single use, disposable supported liquid membrane, which selectively transported and pre-concentrated heavy metal cations into the fibre lumen filled with 100?mM acetic acid acceptor solution. Donor solutions were standard solutions and real samples dissolved in deionized water at neutral pH. At optimized EME conditions (penetration time, 5?s; applied voltage, 75?V; and stirring rate, 750?rpm), 15-42% recoveries of heavy metal cations were achieved for a 5?min extraction time. Repeatability of the EME pre-treatment was examined for six independent EME runs and ranged from 6.6 to 11.1%. Limits of detection for the EME-CE-C(4) D method ranged from 25 to 200?nM, resulting into one to two orders of magnitude improvement compared with CE-C(4) D without sample treatment. The developed EME sample pre-treatment procedure was applied to the analysis of heavy metal cations in tap water and powdered milk samples. Zinc in the real samples was identified and quantified in a background electrolyte solution consisting of 20?mM L-histidine and 30?mM acetic acid at pH 4.95 in about 3?min.     

Quantification of plasma lactate concentrations using capillary electrophoresis with contactless conductivity detection

The quantification of plasma lactate and evaluation of the lactate threshold by CE with capacitively coupled contactless conductivity is demonstrated. The only sample preparation needed was deproteinization with a ACN/methanol mixture. A solution of 10 mmol/L 2-morpholinoethanesulfonic acid monohydrate, 10 mmol/L DL-histidine, 70 μmol/L hexadecyltrimethylammonium bromide, pH 6.0 was found suitable as running buffer. Linearity was achieved for the concentration range of 10-1000 μmol/L with a correlation coefficient of 0.9994. The limit of detection (3 S/N) was determined as 3.2 μmol/L. Intra- and inter-day variabilities were less than 7% RSD. The suitability of the method could be demonstrated by analyzing various clinical samples, where the results correlated satisfactorily with those of an established enzymatic method.     

Evaluation of the detection of biomolecules in capillary electrophoresis by contactless conductivity measurement

The sensitivity of contactless conductivity detection to amino acids, peptides and proteins in CE was studied for BGE solutions of different pH values. The LOD and analytical characteristics were compared for acidic and basic conditions and better results were in most cases found for buffers of low pH values. Linear dynamic ranges varied between two orders of magnitude for amino acids and peptides and three orders of magnitude for larger proteins. The concentration detection limits were found to be between 1.2 and 7.5 microM for the amino acids tested and for the larger molecules they varied between 2.6 microM for leucine enkephalin and 0.2 microM for HSA when using a buffer at pH 2.1.     

Determination of mono- and disaccharides by capillary electrophoresis with contactless conductivity detection

The separation and detection of common mono- and disaccharides by capillary electrophoresis (CE) with contactless conductivity detection (CCD) is presented. At high values of pH, the sugars are converted to anionic species that can be separated by CE and indirectly detected by CCD. The main anionic species present in the running electrolytes are hydroxide and phosphate, which have greater mobility than the ionized sugars, and, thus, the indirect detection is possible. The method was applied to analysis of glucose, fructose, and sucrose in soft drinks, isotonic beverages, fruit juice, and sugarcane spirits. Galactose was used as internal standard in all cases. Plate numbers range from ca. 70,700 to 168,200 and the limits of detection from 13 to 31 microM.     

Separation of fluoroquinolones in acidic buffer by capillary electrophoresis with contactless conductivity detection

A method to determine five fluoroquinolones (FQs), namely, rufloxacin (RUF), ciprofloxacin (CIP), enrofloxacin (ENO), gatifloxacin (GAT) and moxifloxacin (MOX), in acidic buffer by capillary electrophoresis (CE)-capacitively coupled contactless conductivity detection (C⁴D) technique is presented. Separation was carried out in a fused-silica capillary (42 cm × 50 μm) using a buffer composed of 10 mM tartaric acid, 14 mM sodium acetate and 15% (v/v) methanol at pH 3.8. The RSDs of the migration times and peak areas were 0.65% and 12.3% (intraday), 1.28% and 8.8% (interday), respectively. CE-C⁴D in combination with liquid–liquid extraction (LLE) as clean-up and preconcentration procedure, allows detection of the FQs in fortified chicken muscle samples with detection limits of 6.8–11.7 ng/g. This method shows potential in rapid determination of FQs in samples with complex matrix.     

硫酸沙丁胺醇的毛细管电泳非接触电导法检测

建立了毛细管电泳-非接触电导检测分离测定硫酸沙丁胺醇的分析方法。分别考察了分离介质、背景电解质及其浓度和pH、分离电压、进样时间、激发电压、激发频率等因素对实验结果的影响。在优化的实验条件(以15 mmol/L乳酸水溶液(pH 2.7)为电泳介质,10 kV下电动进样3 s,分离电压为10 kV,激发电压为60 V,激发频率为120 kHz)下,硫酸沙丁胺醇的检出限(信噪比为3)为1.92 mg/L,在9.60～48.0 mg/L质量浓度范围内有良好的线性关系(r=0.995),迁移时间的相对标准偏差(RSD)为2.7%。将该方法用于硫酸沙丁胺醇片和硫酸沙丁胺醇气雾剂中的硫酸沙丁胺醇含量的测定,加标回收率为90%～113%,检测结果与药厂的标示值相符合,为硫酸沙丁胺醇的检测提供了一种简便、快速、高灵敏的方法。关键词: 毛细管电泳;非接触电导检测法;硫酸沙丁胺醇;硫酸沙丁胺醇片;硫酸沙丁胺醇气雾剂     

Determination of tobramycin in human serum by capillary electrophoresis with contactless conductivity detection

A study on the determination of the antibiotic tobramycin by CE with capacitively coupled contactless conductivity detection is presented. This method enabled the direct quantification of the non-UV-absorbing species without incurring the disadvantages of the indirect approaches which would be needed for optical detection. The separation of tobramycin from inorganic cations present in serum samples was achieved by optimizing the composition of the acetic acid buffer. Field-amplified sample stacking was employed to enhance the sensitivity of the method and a detection limit of 50 microg/L (S/N = 3) was reached. The RSDs obtained for migration time and peak area using kanamycin B as internal standard were typically 0.12 and 4%, respectively. The newly developed method was validated by measuring the concentration of tobramycin in serum standards containing typical therapeutic concentrations of 2 and 10 mg/L. The recoveries were 96 and 97% for the two concentrations, respectively.     

Improving precision of manual hydrodynamic injection in capillary electrophoresis with contactless conductivity detection

Reproducible injection in capillary electrophoresis has been difficult to achieve with manual injection techniques using simple injection devices, such as gravity injection (siphoning) or hydrodynamic sample splitting. We demonstrate that the injection reproducibility can be improved using very simple means. With hydrodynamic sample splitter, a passive micro-metering valve can be inserted in-line to regulate the sample flow rate through the splitter interface. A significant improvement of both reproducibility and repeatability was achieved. The reproducibility of RSD of the peak areas improved from 25.4% to 4.4%, while the repeatability was below 4.1% when micro-metering valve was used. Additional simple correction that can be used to further improve the variability of injected sample volumes in any hydrodynamic injection mode in CE with conductivity detection was proposed and verified. The measured EOF peak can serve as a simple indicator of the injected volume and can be effectively used for additional correction. By a linear function between the injection volume and the peak area of the EOF, the RSD values of peak areas for both manual gravity injection and hydrodynamic sample splitter were further improved below 2% RSD. The linearity of the calibration curve was also significantly improved. The proposed correction works even with slight differences in matrix composition, as demonstrated on the analysis aqueous soil extract of model mixture of five nerve agent degradation products.     

Fingerprinting postblast explosive residues by portable capillary electrophoresis with contactless conductivity detection

A portable capillary electrophoretic system with contactless conductivity detection was used for fingerprint analysis of postblast explosive residues from commercial organic and improvised inorganic explosives on various surfaces (sand, concrete, metal witness plates). Simple extraction methods were developed for each of the surfaces for subsequent simultaneous capillary electrophoretic analysis of anions and cations. Dual‐opposite end injection principle was used for fast (<4 min) separation of 10 common anions and cations from postblast residues using an optimized separation electrolyte composed of 20 mM MES, 20 mM l ‐histidine, 30 μM CTAB and 2 mM 18‐crown‐6. The concentrations of all ions obtained from the electropherograms were subjected to principal component analysis to classify the tested explosives on all tested surfaces, resulting in distinct cluster formations that could be used to verify (each) type of the explosive.     

A thin cover glass chip for contactless conductivity detection in microchip capillary electrophoresis

A microfabricated thin glass chip for contactless conductivity detection in chip capillary electrophoresis is presented in this contribution. Injection and separation channels were photolithographed and chemically etched on the surface of substrate glass, which was bonded with a thin cover glass (100 μm) to construct a new microchip. The chip was placed over an independent contactless electrode plate. Owing to the thinness between channel and electrodes, comparatively low excitation voltage (20–110 V in V_p–p) and frequency (40–65 kHz) were suitable, and favorable signal could be obtained. This microchip capillary electrophoresis device was used in separation and detection of inorganic ions, amino acids and alkaloids in amoorcorn tree bark and golden thread in different buffer solutions. The detection limit of potassium ion was down to 10 μmol/L. The advantages of this microchip system exist in the relative independence between the microchip and the detection electrodes. It is convenient to the replacement of chip and other operations. Detection in different position of the channel would also be available.     

Optimization of separation of heavy metals by capillary electrophoresis with contactless conductivity detection

The separation of four toxic metal ions (Cr(3+) , Pb(2+) , Hg(2+) , Ni(2+) ) was achieved by optimizing the composition of the histidine/tartaric acid background electrolyte. An on-column preconcentration technique, viz. field amplified sample injection, was performed to improve the sensitivity. This method afforded an enhancement factor of up to 91,800 fold with the LODs ranging from 0.005 to 2.32 μg/L, which were well below the maximum contaminant levels set by the United States Environmental Protection Agency. The robustness of this method was demonstrated with its application to the analysis of real samples including tap water, drain water, and reservoir water with recoveries between 90 and 120%.     

微芯片毛细管电泳非接触电导法快速测定盐酸倍他洛尔

建立了微芯片毛细管电泳非接触电导检测法快速测定盐酸倍他洛尔滴眼液中盐酸倍他洛尔的含量。探讨了缓冲液类型、浓度、分离电压及进样时间等因素对分离检测的影响。实验采用1.5 mmol·L-1HAc-1.5mmol·L-1Na Ac(p H=4.69)为缓冲溶液,分离电压为2.1 k V,进样时间10.0 s。此条件下于0.7 min内实现了盐酸倍他洛尔的快速分离测定。盐酸倍他洛尔的浓度在5.0~200.0μg·m L-1线性良好(r=0.9997,n=6),检出限为1.0μg·m L-1(S/N=3),RSD为0.8%,样品的加标回收率为100.4%~102.0%。滴眼液中的辅料在该条件下不干扰测定,可成功测定盐酸倍他洛尔滴眼液中盐酸倍他洛尔的含量。