Cation-selective Exhaustive Injection Research Articles

On-Line Preconcentration and Determination of Nuciferine and O-Nornuciferine in Lotus Leaves by Cation-Selective Exhaustive Injection (CSEI)-Sweeping-Micellar Electrokinetic Chromatography (MEKC)

On-line preconcentration incorporating cation-selective exhaustive injection (CSEI) and sweeping followed by micellar electrokinetic chromatography (MEKC) was developed for the simultaneous determination of the bioactive alkaloids nuciferine and O-nornuciferine. The parameters affecting the analytical performance were systematically investigated. Under the optimal conditions, the enrichment factor was significantly improved by 7000 to 13000-fold compared with traditional MEKC. The analytes were well separated within 15 min. The limits of detection for nuciferine and O-nornuciferine were 4.00 and 2.00 ng∙mL−1, and the limits of quantification were 8.00 and 10.00 ng∙mL−1, respectively. The relative standard deviations for intra-day and inter-day precision were less than 3.0%. The recoveries of the analytes were from 98.9% to 102%. Furthermore, the developed method was demonstrated to be suitable for the analysis of nuciferine and O-nornuciferine in lotus leaves, offering high performance without the loss of resolution. CSEI-sweeping-MEKC provides a simple, environmental-friendly option for preconcentration and quantification of nuciferine and O-nornuciferine.

A novel cation-selective exhaustive injection and sweeping micellar electrokinetic chromatography method for the simultaneous determination of second-generation triazoles in human plasma

In this study, we introduce the first capillary electrophoresis method that combines cation-selective exhaustive injection and sweeping micellar electrokinetic chromatography (CSEI-sweeping-MEKC) for the simultaneous and sensitive quantification of voriconazole (VRC), posaconazole (PSC), and isavuconazole (ISV) levels in human plasma. The sensitivity was remarkably improved, with a 5374-fold enhancement compared to the traditional MEKC method. We systematically optimized key parameters affecting sensitivity, resolution, and efficiency. The separation buffer included 100 mM NaH2PO4, 13 % acetonitrile, and 13 % tetrahydrofuran. This was followed by a high-conductivity buffer zone (1 psi, 50 s) of 100 mM NaH2PO4, an extended electrokinetic sample injection (10 kV, 600 s), and analyte sweeping with 100 mM sodium dodecyl sulfate micelles at −20 kV. Under these optimized conditions, the method allowed for an analytical run in 10 min and was validated for linearity (r ≥ 0.9997), precision (%RSD ≤ 8.8 %), and accuracy (%RE ≤ 2.20 %), delivering satisfactory results. The limit of detection was 2.55 ng/mL for ISV, 0.89 ng/mL for PSC, and 1.04 ng/mL for VRC. This method was successfully applied to analyze plasma samples from patients undergoing triazole antifungal treatment, demonstrating its feasibility for clinical testing.

Stereoselective enrichment and determination of citalopram enantiomers by cation-selective exhaustive injection and sweeping coupled with cyclodextrin modified electrokinetic chromatography

An highly sensitive, rapid and enantioselective method was developed for the enantioseparation and determination of citalopram enantiomers by cation selective exhaustive injection-sweeping-cyclodextrin modified electrokinetic chromatography (CSEI-sweeping-CDEKC). The optimized conditions were: 50 mM pH 3.0 phosphate solution with 25 mg·mL−1 S-β-CD used as background buffer, 50 mbar 300 s hydrodynamical injection of 150 mM pH 3.0 NaH2PO4 buffer followed with 5 s water plug, 10 kV 600 s electrokinetic sample injection, -20 kV CDEKC run. Under the optimized conditions, the resolution was Rs=8.04, the enrichment factor as up to 2163 folds, the LOD values were: 3.6 ng·mL−1 for R-citalopram, 4.1 ng·mL−1 for S-citalopram, and 3 ng·mL−1 for both enantiomers in plasma samples. This new method showed good precision, repeatability and stability, which had been successfully applied to the impurity inspection of escitalopram oxalate and the stereoselective pharmacokinetic study of citalopram enantiomers.

Stereoselective Analysis by Design in the Development of a Cation-Selective Exhaustive Injection and Sweeping Coupled with Cyclodextrin Modified Electrokinetic Chromatography Method for the Determination of Citalopram Enantiomers

Stereoselective Analysis by Design in the Development of a Cation-Selective Exhaustive Injection and Sweeping Coupled with Cyclodextrin Modified Electrokinetic Chromatography Method for the Determination of Citalopram Enantiomers

Enantioselective analysis of pheniramine in rat using large volume sample stacking or cation-selective exhaustive injection and sweeping coupled with cyclodextrin modified electrokinetic chromatography

For the aim of simultaneously performing the enantioseparation and determination of the trace enantiomers in plasma samples, enantioseparation by HPLC using five kinds of chiral stationary phases were initially investigated. But unfortunately, enantioseparation could not be detected in reversed mobile phase mode with all the five columns. For this reason, two simple, economical and highly efficient online preconcentration methods, large volume sample stacking and sweeping (LVSS-sweeping) and cation-selective exhaustive injection and sweeping (CSEI-sweeping) both followed by the cyclodextrin modified electrokinetic chromatography (CDEKC) were examined in the present work. Parameters affecting the enantioseparation and enhancement efficiency of these two injection modes were monitored in detail, and migration order of the two enantiomers was identified by circular dichroism (CD) and HPLC. Upon optimization, two enantiomers were best separated with the improvement of sensitivity reaching 160-fold and 4000-fold respectively for LVSS-sweeping and CSEI-sweeping comparing with the normal CDEKC separation. Then the optimal condition of CSEI-sweeping-CDEKC was validated and showed high sensitivity (10 ng/mL for lower limit of quantification, LLOQ), satisfactory accuracy (96.8–111.6%) and precision (relative standard deviation, RSD within 9.4%). This demonstrated it to be a suitable strategy for the rapid enantioselective determination and quantitative analysis of pheniramine enantiomers in plasma samples. Therefore, the method was further applied in the enantiomeric analysis of pheniramine in rat pharmacokinetics and plasma protein binding investigations. Stereoselectivity in pharmacokinetics as well as plasma protein binding were observed, suggesting that the stereoselective protein binding might be responsible for the stereoselectivity in pharmacokinetics.

Determination of brompheniramine enantiomers in rat plasma by cation-selective exhaustive injection and sweeping cyclodextrin modified electrokinetic chromatography method.

A method consisting of cation-selective exhaustive injection and sweeping (CSEI-sweeping) as online preconcentration followed by a cyclodextrin modified electrokinetic chromatography (CDEKC) enantioseparation has been developed for the simultaneous determination of two brompheniramine enantiomers in rat plasma. In this method, analytes were electrokinetically injected at a voltage of 8 kV for 80 s in a fused-silica capillary. Prior to the injection, the capillary was rinsed with 50mM phosphate buffer of pH 3.5, followed by a plug of a higher conductivity buffer (150mM phosphate pH 3.5, 20psi, 6min) and a plug of water (0.5psi, 5 s). Separation was carried out applying -20 kV in 50mM phosphate buffer, pH 3.5, containing 10% v/v ACN and 30mg/mL sulfated-β-cyclodextrin (S-β-CD). Analytical signals were monitored at 210 nm. The detection sensitivity of brompheniramine enantiomers was enhanced by about 2400-fold compared to the normal injection mode (hydrodynamic injection for 3 s at 0.5psi, with a BGE of 50mM phosphate buffer containing 20mg/mL S-β-CD at pH 3.5), and LLOQ of two enantiomers were both 0.0100μg/mL. In addition, this method had fairly good repeatability and showed promising capabilities in the application of stereoselective pharmacokinetic investigations for brompheniramine enantiomers in rat.

The cation-selective exhaustive injection and sweeping capillary electrophoresis method for the analysis of chlorpheniramine enantiomers in rat plasma

Chlorpheniramine is a potent antihistaminic administered as a racemic mixture, while its clinical activity is mainly associated with the S-enantiomer. In this study, a sensitive and rapid on-line preconcentration capillary electrophoresis (CE) method, cation-selective exhaustive injection (CSEI) and sweeping method was established and validated for the analysis of two chlorpheniramine enantiomers. Parameters influencing separation and enhancement efficiency were investigated, including cyclodextrin (CD) types and concentration, background electrolyte pH, type and content of organic modifier, injection time of water plug, injection time and voltage of sample. A zone of 30mM Tris buffer at pH 3.5 without chiral selector was injected into the capillary followed by a 3s water plug, allowing for the analytes to be electrokinetically injected at a voltage of 10kV for 80s. A 30mM Tris buffer at pH 3.5 consisting of 20mM sulfated-β-cyclodextrin (S-β-CD) and 5% methanol was found to be highly efficient for the separation of the two enantiomers. The present method manifested that high sensitivity (0.025μg/mL for the thethe lower limit of quantification), satisfactory accuracy (89.2%–95.0%) and precision (relative standard deviation within 8.4%) were achieved as well as favourable stability. The extraction recoveries of two enantiomers were both above 72.5%. Finally, the developed method was successfully applied to pharmacokinetic study of the two enantiomers in rat plasma after oral administration of racemic chlorpheniramine.

Approaching over 10000-fold sensitivity increase in chiral capillary electrophoresis: Cation-selective exhaustive injection and sweeping cyclodextrin-modified micellar electrokinetic chromatography.

A novel and simple method that combines an online concentration technique with an enantioseparation technique for capillary electrophoresis-namely, cation-selective exhaustive injection and sweeping cyclodextrin-modified micellar electrokinetic chromatography (CSEI-sweeping CD-modified MEKC)-realizes the effective enantioseparation of cationic analytes while keeping a significant increase of detection sensitivity. This technique consists of a slight modification of the basic CSEI-sweeping MEKC. The main idea is to simply add an anionic CD as a chiral selector into the micellar buffer including sodium dodecyl sulfate, but not to change any other buffers in order to preserve the online concentration mechanism. When applied to analysis of the street drug, methamphetamine, the method achieved not only a baseline enantioseparation but also limits of detection (LODs; S/N = 3) of 70-90 pg/mL (ppt) for each isomer. This translates to a more than 10000-fold improvement compared to the LODs by the usual injection method. The present technique, which was made from a slight modification of CSEI-sweeping MEKC, would give an attractive approach that is applicable to almost any analytes for which CSEI-sweeping MEKC is applicable; all that is required is the selection of an appropriate anionic CD to be added to the micellar buffer.

Evaluation of the combination of micellar electrokinetic capillary chromatography with sweeping and cation selective exhaustive injection for the determination of 5-nitroimidazoles in egg samples

A methodology is presented for the sensitive determination of nitromidazole residues in egg by means of micellar electrokinetic capillary chromatography in combination with cation selective exhaustive injection and ultraviolet detection. Six compounds have been considered and the separation has been achieved in less than 12min in a 61.5-cm effective length capillary with 50-μm internal diameter. Phosphate buffer 44mM pH 2.5, containing 8% tetrahydrofurane and 123mM sodium dodecyl sulfate was employed as running buffer. Solid phase extraction has been employed for sample clean-up. The methodology has been successfully validated in hen eggs, obtaining method detection limits in the range of 2.1–5.0ng/g. Precision was studied in terms of repeatability and intermediate precision, with relative standard deviations lower than 18.0%. Recoveries were calculated in quail eggs and a commercial pasteurized egg white product, reaching over 70% for most of the considered 5-nitroimidazoles.

Development of an ultrasensitive stacking technique for 5-nitroimidazole determination in untreated biological fluids by micellar electrokinetic chromatography.

Cation-selective exhaustive injection and sweeping followed by a MEKC separation is evaluated for the sensitive analysis of 5-nitroimidazoles in untreated human serum and urine. Deproteinized serum and urine samples were diluted 76 and 143 times, respectively, in a low-conductivity solvent (5.00 mM orthophosphoric acid containing 5.0% v/v methanol). Samples were electrokinetically injected at 9.8 kV for 632 s in a previously conditioned fused-silica capillary (65.0 cm × 50 μm id). Separation was performed at -30 kV and 20°C using 44 mM phosphate buffer (pH 2.5), 123 mM SDS, and 8% v/v tetrahydrofurane as BGE. Signals were monitored at 276 nm and peak area was selected as analytical response. Good linearity (R(2) ≥ 0.988) and LODs lower than 1.5 and 1.8 μg/mL were achieved in serum and urine, respectively.

Determination of imidazole derivatives by micellar electrokinetic chromatography combined with solid-phase microextraction using activated carbon-polymer monolith as adsorbent

In this study, an effective method for the separation of imidazole derivatives 2-methylimidazole (2-MEI), 4- methylimidazole (4-MEI) and 2-acetyl-4-tetrahydroxybutylimidazole (THI) in caramel colors using cation-selective exhaustive injection and sweeping micellar electrokinetic chromatography (CSEI-sweeping-MEKC) was developed. The limits of detection (LOD) and quantitation (LOQ) for the CSEI-sweeping-MEKC method were in the range of 4.3–80μgL−1 and 14–270μgL−1, respectively. Meanwhile, a rapid fabrication activated carbon-polymer (AC-polymer) monolithic column as adsorbent for solid-phase microextraction (SPME) of imidazole colors was developed. Under the optimized SPME condition, the extraction recoveries for intra-day, inter-day and column-to-column were in the range of 84.5–95.1% (<6.3% RSDs), 85.6–96.1% (<4.9% RSDs), and 81.3–96.1% (<7.1% RSDs), respectively. The LODs and LOQs of AC-polymer monolithic column combined with CSEI-sweeping-MEKC method were in the range of 33.4–60.4μgL−1 and 111.7–201.2μgL−1, respectively. The use of AC-polymer as SPME adsorbent demonstrated the reduction of matrix effect in food samples such as soft drink and alcoholic beverage thereby benefiting successful determination of trace-level caramel colors residues using CSEI-sweeping-MEKC method. The developed AC-polymer monolithic column can be reused for more than 30 times without any significant loss in the extraction recovery for imidazole derivatives.

Determination of Nicotine in Tobacco by Chemometric Optimization and Cation-Selective Exhaustive Injection in Combination with Sweeping-Micellar Electrokinetic Chromatography.

Nicotine is a potent chemical that excites the central nervous system and refreshes people. It is also physically addictive and causes dependence. To reduce the harm of tobacco products for smokers, a law was introduced that requires tobacco product containers to be marked with the amount of nicotine as well as tar. In this paper, an online stacking capillary electrophoresis (CE) method with cation-selective exhaustive injection sweeping-micellar electrokinetic chromatography (CSEI-sweeping-MEKC) is proposed for the optimized analysis of nicotine in tobacco. A higher conductivity buffer (160 mM phosphate buffer (pH 3)) zone was injected into the capillary, allowing for the analytes to be electrokinetically injected at a voltage of 15 kV for 15 min. Using 50 mM sodium dodecyl sulfate and 25% methanol in the sweeping buffer, nicotine was detected with high sensitivity. Thus, optimized conditions adapted from a chemometric approach provided a 6000-fold increase in the nicotine detection sensitivity using the CSEI-sweeping-MEKC method in comparison to normal CZE. The limits of detection were 0.5 nM for nicotine. The stacking method in combination with direct injection which matrix components would not interfere with assay performance was successfully applied to the detection of nicotine in tobacco samples.

Novel cation selective exhaustive injection-sweeping procedure for 5-nitroimidazole determination in waters by micellar electrokinetic chromatography using dispersive liquid–liquid microextraction

A novel method consisting of cation-selective exhaustive injection and sweeping (CSEI-sweeping) as on-line preconcentration followed by a micellar electrokinetic chromatography (MEKC) separation has been developed for the determination of 5-nitroimidazoles (5-NDZ) in environmental waters. Moreover, dispersive liquid–liquid microextraction (DLLME) has been proposed for first time as sample treatment technique prior to CSEI-sweeping-MEKC. DLLME was applied to 5mL of sample. Dibromomethane (1156μL) and 2-butanol (1363μL) were employed as extractant and dispersive solvents, respectively. Salting-out effect was achieved by the addition of 16% (w/v) NaCl to the samples. After DLLME and organic solvent evaporation, the residue was redissolved in a low conductivity solvent (5mM phosphoric acid with 5% of methanol) and electrokinetically injected at 9.8kV for 632s in a bare fused-silica capillary (57.2cm, 50μm I.D.). Prior to the injection, the capillary was rinsed with 50mM phosphate buffer pH 2.5, followed by a plug of a higher conductivity buffer (100mM phosphate pH 2.5, 50mbar, 264s) and a plug of water (50mbar, 2s). Separation was carried out applying −30kV at 20°C in 44mM phosphate buffer pH 2.5, containing 8% tetrahydrofuran and 123mM sodium dodecyl sulfate. Analytical signals were monitored at 276nm. Validation was performed in river and well waters, obtaining satisfactory results in terms of linearity, precision (% RSD generally lower than 10%) and trueness (recoveries higher than 70% in almost all cases). LODs ranged from 0.61 to 2.44ng/mL. The combination of this microextraction technique with the proposed capillary electrophoresis methodology supposes a simple, sensitive and cheap alternative for 5-NDZ analyses, in accordance with the aims of green chemistry.

Chemometric Optimization of Cation-Selective Exhaustive Injection Sweeping Micellar Electrokinetic Chromatography for Quantification of Ractopamine in Porcine Meat

An online stacking capillary electrophoresis (CE) method, cation-selective exhaustive injection sweeping micellar electrokinetic chromatography (CSEI-sweep-MEKC), is developed and optimized for analysis of ractopamine (RP) and its homologue dehydroxyractopamine (DRP) in porcine meat. Chemometric experimental design was used to achieve the best possible optimization and reduce the number of trials and errors. The CSEI-sweep-MEKC method enables nanogram per gram level analysis, with limits of detection (LODs) in meat of 5 ng/g for RP and 3 ng/g for DRP (S/N = 3). A higher conductivity buffer (HCB) zone was injected into the capillary, allowing for the analytes to be electrokinetically injected at a voltage of 9 kV for 12 min. Using 125 mM sodium dodecyl sulfate and 15% methanol in the sweeping buffer, RP and DRP were well-separated. The method was validated with a linear calibration curve of 10-300 ng/g (r > 0.994). In comparison to the normal capillary zone electrophoresis method (1 psi for 10 s), this stacking strategy resulted in 900 times sensitivity enhancement. This technique was further applied for analyzing seven kinds of commercial meats, and the residual RP was detected in one (5.76 ng/g of RP). The data were corresponding to the data analyzed by the commercial testing kit and mass spectrometry spectra. This method was successfully used on real samples and is considered feasible for serving as a tool for routine examination in markets.

Concentration and determination of cotinine in serum by cation‐selective exhaustive injection and sweeping micellar electrokinetic chromatography

Micellar electrokinetic chromatography (MEKC), combined with on-line concentration techniques, cation-selective exhaustive injection (CSEI) and sweeping, was developed for the analysis of cotinine, the primary biomarker for exposure to secondhand smoke. Experimental parameters including sample matrix, surfactant concentration, injection length and concentration of high-conductivity buffer, and sample electrokinetic injection time were optimized for electrophoretic enrichment and separation processes. Under the optimal conditions, the detection sensitivity of cotinine was enhanced by about 5000-fold using CSEI-sweeping MEKC compared to normal MEKC. The limit of detection for cotinine was found to be 0.2 ng/mL using ultraviolet absorbance detection. Furthermore, the developed method was successfully applied to the detection of cotinine in mouse serum samples.

A chemometric approach for the elucidation of the parameter impact in the hyphenation of field‐enhanced sample injection and sweeping in capillary electrophoresis

The aim of this work was to elucidate the impacts of parameters influencing cation-selective exhaustive injection coupled to sweeping and micellar electrokinetic chromatography (MEKC). A chemometric approach using cationic compounds, acidic conditions (phosphate buffer, pH 2.3) and polyacrylamide-coated capillaries to suppress electroosmotic flow were used. It was demonstrated that the water plug was not useful because of long electrokinetic injections. If conductivity of the high conductivity buffer (HCB) and the HCB to sample conductivity ratio are sufficiently high (>1.66 S/m and >30, respectively), variations of HCB conductivity do not impact sensitivity. The length of the HCB must be long enough so that the most mobile cation remains stacked in this zone for a given injection time. SDS concentration should be as high as possible (the maximum concentration is dictated by MEKC, here 90 mM), so sensitivity is not impacted. We have shown analytes can be lost after electrokinetic injection, when the polarity of the voltage is reversed. Introducing a plug of micellar electrolyte before polarity reversal avoids these losses. Following these recommendations only injection time and sample conductivity impacted sensitivity enhancement. Sample conductivity had to be the lowest as possible and controlled in real case analyses to obtain repeatable enrichment factors.

Quantitative determination of isoniazid in biological samples by cation-selective exhaustive injection–sweeping–micellar electrokinetic chromatography

Tuberculosis (TB), an infectious disease caused by Mycobacterium tuberculosis, infects approximately one third of the current world population. Isoniazid is one of the most frequently used first-line anti-TB drugs. In this study, we developed a sensitive cation-selective exhaustive injection-sweeping-micellar electrokinetic chromatography method (CSEI-Sweep-MEKC) for analyzing isoniazid in human plasma. Parameters including acetonitrile (ACN) percentage in the separation buffer; the injection time, and concentration of the high-conductivity buffer; sodium dodecyl sulfate (SDS) concentration; phosphate concentration in the sample matrix; and the sample injection time were all optimized to obtain the best analytical performance. The optimal background electrolyte comprised 50 mM phosphate buffer, 100 mM SDS, and 15% ACN. Non-micelle background electrolyte, containing 75 mM phosphate buffer and 15% ACN, was first injected into the capillary, followed by a short plug of 200 mM phosphate (high-conductivity buffer). Run-to-run repeatability (n=3) and intermediate precision (n=3) of peak area ratios were found to be lower than 8.7% and 11.4% RSD, respectively. The accuracy of the method was within 98.1-106.9%. The limit of detection of isoniazod in human plasma was 9 ng mL(-1). Compared with conventional MEKC, the enhancement factor of the CSEI-Sweep-MEKC method was 85 in plasma samples. The developed method was successfully used to determine isoniazid concentration in patient plasma. The results demonstrated that CSEI-Sweep-MEKC has the potential to analyze isoniazid in human plasma for therapeutic drug monitoring and clinical research.

Cation-selective exhaustive injection and sweeping micellar electrokinetic chromatography for the analysis of methadone and its metabolites in serum of heroin addicts

Methadone (MET) metabolism has been largely demonstrated with a high inter-individual variability and, therefore, quantification of MET is very important for therapeutic drug monitoring. A cation-selective exhaustive injection and sweeping MEKC (CSEI-Sweeping) was first developed to analyze MET and its two metabolites, 2-ethylidene-1,5-dimethyl-3,3-diphenylpyrrolidine (EDDP) and 2-ethyl-5-methyl-3,3-diphenyl-1-pyrroline (EMDP), in human serum. After pretreatment, the samples were electrokinetically injected into capillary (10 kV, 500 s) and swept by the separation phosphate buffer (100 mM, pH 4.0) containing 20% tetrahydrofuran and 100 mM SDS at −15 kV. The LODs were 200 pg/mL for MET and EMDP, and 400 pg/mL for EDDP. Ten volunteers were administered MET (5.0–120.0 mg/day) orally for 84 days and serum samples were taken after the daily dose of MET (days 1, 2, 7, 14, 28, 56 and 84) individually. This method was used for monitoring MET and its metabolites in heroin addicts and for pharmacokinetic investigations.

Determination of cypromazine and its metabolite melamine in milk by cation‐selective exhaustive injection and sweeping‐capillary micellar electrokinetic chromatography

In this study, we described a high-sensitive on-line preconcentration method for cypromazine (CYP) and melamine (MEL) analysis using cation-selective exhaustive injection (CSEI) combined with sweeping-MEKC. The optimum conditions of on-line concentration and separation were discussed. The BGE contained 100 mM SDS, 50 mM phosphoric acid (pH=2.0) and 15% acetonitrile (v/v). The sample was injected at 10 kV for 600 s, separated at -20 kV, and detected at 210 nm. The sensitivity enhancements were 6222 for CYP and 9179 for MEL. The linear dynamic ranges were 0.4-25 ng/mL for CYP (r=0.9995) and 0.2-12 ng/mL for MEL (r=0.9991). The LODs (signal-to-noise ratio, 3) were 43.7 and 23.4 pg/mL for CYP and MEL, respectively. The proposed method was applied to analyze CYP and MEL in dairy products pretreated using off-line SPE to minimize the influence of the matrix. The recoveries of CYP and MEL were satisfactory (ca. 74-83%). The experimental results suggest that the CSEI-sweeping-MEKC method is feasible for the application to simultaneously detect trace levels of CYP and its metabolite MEL in real milk samples.

On-line concentration and determination of tobacco-specific N-nitrosamines by cation-selective exhaustive injection–sweeping–micellar electrokinetic chromatography

In this paper, a micellar electrokinetic chromatography (MEKC) method combined with cation-selective exhaustive injection (CSEI) and sweeping was developed to separate and concentrate four tobacco-specific N-nitrosamines (TSNAs) including N′-nitrosoanabasine (NAB), 4-(methylnitrosamino)-1-(3-pyridyl)-1-butanone (NNK), 4-(methylnitrosamino)-1-(3-pyridyl)-1-butanol (NNAL) and 4-(methylnitrosamino)-4-(3-pyridyl)-1-butanol (iso-NNAL). Experimental parameters affecting separation efficiency and enhancement factors were investigated in detail. Under the optimum MEKC condition, NAB, NNK, NNAL and iso-NNAL were baseline separated with high separation efficiencies and good peak shapes. Furthermore, with the preconcentration by CSEI–sweeping–MEKC, the sensitivity enhancement factors for NAB, NNK, NNAL and iso-NNAL in terms of peak areas ranged from 6.0 × 10 3 to 1.5 × 10 4, and the detection limits (LOD, S/N = 3) of four TSNAs were in the range of 0.004–0.016 μg/mL. In addition, this method had fairly good repeatability, and the RSDs of retention time and peak area were less than 1% and 5%, respectively. Finally, this method showed promising capabilities in the application of detecting and analyzing TSNAs in human urine samples.