Capillary electrophoresis-electrospray ionization ion trap mass spectrometry for analysis and confirmation testing of morphine and related compounds in urine

Using an aqueous background electrolyte containing 25 mM ammonium acetate and NH3 (pH 9), CE-tandem MS and CE-triple MS with atmospheric pressure electrospray ionization in the positive ion mode are shown to represent attractive approaches for analysis and confirmation testing of morphine (MOR) and related opioids in human urine. Injection of plain or diluted urine permits monitoring of solutes at concentrations above 2-5 microg/ml. For the recognition of lower concentrations, solute extraction and concentration is required. Liquid-liquid extraction at alkaline pH is shown to be suitable for analysis of free opioids only whereas solid-phase extraction using a mixed-mode polymer phase is demonstrated to permit analysis of both free and glucuronidated opioids. The former sample preparation approach, however, requires about half of the time only. Commencing with 2 ml of urine, reconstitution to provide a sample volume of 0.2 ml and hydrodynamic sample injection, detection limits for free opioids are shown to be on the 100-200 ng/ml drug level. Much improved (ppb) sensitivity is obtained by infusing the extract directly into the source of the MS system. However, solutes that produce equal fragments (such as the two glucuronides of MOR) can thereby not be distinguished. CE-tandem MS and CE-triple MS are demonstrated to be suitable to confirm the presence of MOR, MOR-3-glucuronide, 6-monoacetylmorphine, codeine, codeine-6-glucuronide, dihydrocodeine, methadone and 2-ethylidene-1,5-dimethyl-3,3-diphenylpyrrolidine in a toxicological quality control urine. The same is shown for selected metabolites of codeine and dihydrocodeine in urines collected after administration of pharmaceutical preparations.     

Identification of diphenhydramine metabolites in human urine by capillary electrophoresis-ion trap-mass spectrometry

The identification of diphenhydramine (DH) metabolites that are frequently observed in the capillary zone electrophoresis (CZE) and micellar electrokinetic capillary chromatography (MEKC) analyses of alkaline liquid/liquid and solid-phase extracts of patient urines is demonstrated. Having standards for DH and diphenhydramine-N-oxide (DHNO), the presence of these two compounds could be confirmed in urines that were collected overnight after administration of 25 mg DH chloride. Using CZE coupled to ion-trap mass spectrometry (CE-MS(n)) with positive electrospray ionization and an acetate buffer at pH 5.6, the [M+H](+) ions of DH (m/z = 256), DHNO (m/z = 272), and nordiphenhydramine (NDH, m/z = 242) and their fragmentation to a common m/z 167 product ion (diphenylcarbinol moiety) was monitored. The data indicate that all three compounds are cations in an acidic environment, the migration order being NDH, DH, and DHNO. Data obtained under negative electrospray ionization conditions suggest the presence of diphenylmethoxyacetic acid-glycine amide ([M-H](-) ion of m/z 298 and fragmentation to m/z 254, loss of CO(2)), a metabolite that could tentatively be assigned to a characteristic peak observed in the MEKC electropherogram at alkaline pH. The data presented in this paper illustrate the value of using CE-MS(n) for identification of urinary drug metabolites for which no standards are available.     

Head-column field-amplified sample stacking in presence of siphoning. Application to capillary electrophoresis-electrospray ionization mass spectrometry of opioids in urine

Capillary electrophoresis (CE) with head-column field-amplified sample stacking (FASS) in presence of a water plug inserted at the capillary tip is a robust approach providing a more than 1000-fold sensitivity enhancement when applied to low-conductivity samples that are analyzed in an integrated instrument. Employing modular systems comprising a small hydrodynamic buffer flow (siphoning) towards the capillary end and featuring UV absorption or electrospray ionization mass spectrometric (MS) detection, insertion of a water plug is demonstrated to deteriorate the performance of head-column FASS or making it unfunctional. Electroinjection in the absence of the water plug can be employed instead and is shown to provide a ng/ml sensitivity when applied to low conductivity samples. With some suction of sample into the capillary during electroinjection, contamination of the sample vial with buffer is thereby largely avoided. Electroinjection applied to the CE-ion trap MS-MS and MS-MS-MS analysis of twofold diluted urines, urinary solid-phase extracts and urinary liquid-liquid extracts is shown to provide much improved sensitivity compared to hydrodynamic injection of these samples. With electroinjection from diluted urine and urinary solid-phase extracts, the presence of free opioids and their glucuronic acid conjugates can be unambiguously confirmed in urines that were collected after single-dose administration of small amounts of opioids (tested with about 7 mg codeine and 25 mg dihydrocodeine, respectively). Thus, CE-multiple MS with direct electroinjection of opioids from untreated urines could prove to become a rapid and simple approach for unambiguous urinary testing of drug abuse. Procedures leading to the reduction of siphoning in modular CE setups are briefly discussed as well.     

Analysis of oxycodol and noroxycodol stereoisomers in biological samples by capillary electrophoresis

A capillary electrophoresis (CE) method for the separation of the diastereoisomers of 6-oxycodol (6OCOL) and nor-6-oxycodol (N6OCOL), the 6-keto-reduced metabolites of oxycodone (OCOD) and noroxycodone (NOCOD), respectively, is reported and employed to assess the stereoselectivity of these metabolic steps in vivo, in vitro, and in chemical synthesis. CE in an untreated fused-silica capillary with acidic buffers containing 2-hydroxypropyl-beta-cyclodextrin, randomly sulfated beta-cyclodextrin, or single isomer heptakis(2,3-diacetyl-6-sulfato)-beta-cyclodextrin (HDAS-beta-CD) is shown to permit the simultaneous separation of the stereoisomers of 6OCOL and N6OCOL. A 100 mM phosphate buffer of pH 2.0 containing 2.05% w/v HDAS-beta-CD provides a medium for rapid analysis and unambiguous identification of these stereoisomers in solid-phase extracts of (i) urines stemming from patients under pharmacotherapy with OCOD, (ii) incubations of OCOD and NOCOD with human liver cytosol and the human liver S9 fraction, and (iii) after chemical synthesis from OCOD and NOCOD using NaBH(4). In all cases, alpha-N6OCOL is shown to be the predominant stereoisomer of N6OCOL. For 6OCOL, the same is true for in vitro formation and for chemical synthesis. In urine, however, beta-6OCOL is observed to be excreted in a higher amount than alpha-6OCOL. For the urinary alpha-/beta-isomer ratio of 6OCOL and N6OCOL, there are no differences between the data obtained for nonhydrolyzed and enzymatically hydrolyzed urines. The data document the stereoselectivity of the 6-keto-reduction of OCOD and NOCOD in man.     

Stereoselective screening for and confirmation of urinary enantiomers of amphetamine, methamphetamine, designer drugs, methadone and selected metabolites by capillary electrophoresis.

Data presented in this paper demonstrate that a competitive binding, electrokinetic capillary-based immunoassay previously used for screening of urinary amphetamine and analogs cannot be employed to distinguish between the enantiomers of amphetamine and methamphetamine. However, capillary zone electrophoresis with a pH 2.5 buffer containing (2-hydroxypropyl)-beta-cyclodextrin as chiral selector is shown to permit the enantioselective analysis of urinary extracts containing methamphetamine, amphetamine, 3,4-methylenedioxymethamphetamine (Ecstasy) and other designer drugs, and methadone together with its major metabolite, 2-ethylidene-1,5-dimethyl-3,3-diphenylpyrrolidine. In that approach, enantiomer identification is based upon comparison of extracted polychrome UV absorption data and electropherograms obtained by rerunning of spiked extracts with spectra and electropherograms monitored after extraction of fortified blank urine. The suitability of the described chiral electrokinetic capillary method for drug screening and confirmation is demonstrated via analysis of unhydrolyzed quality control urines containing a variety of drugs of abuse. Furthermore, in a urine of a patient under selegiline pharmacotherapy, the presence of the R-(-)-enantiomers of methamphetamine and amphetamine could be unambiguously identified. Direct intake of an R-enantiomer or ingestion of drugs that metabolize to the R-enantiomers can be distinguished from the intake of S-(+)-enantiomers (drug abuse) or prescribed drugs that metabolize to the S-enantiomers of methamphetamine and amphetamine. The described approach is simple, reproducible, inexpensive and reliable (free of interferences of other major basic drugs that are frequently found in toxicological urines) and could thus be used for screening for and confirmation of urinary enantiomers in a routine laboratory.     

Analysis of lorazepam and its 30-glucuronide in human urine by capillary electrophoresis: evidence for the formation of two distinct diastereoisomeric glucuronides

Lorazepam (LOR) is a 3-hydroxy-1,4-benzodiazepine that is chiral and undergoes enantiomerization at room temperature. In humans, about 75% of the administered dose of LOR is excreted in the urine as its 30-glucuronide. CE-MS with negative ESI was used to confirm the presence of LOR-30-glucuronide in urines that stemmed from a healthy individual who ingested 1 or 2 mg LOR, whereas free LOR could be detected in extracts prepared from enzymatically hydrolyzed urines. As the 30-glucuronidation reaction occurs at the chiral center of the molecule, two diastereoisomers can theoretically be formed, molecules that can no longer interconvert. The stereoselective formation of LOR glucuronides in humans and in vitro was investigated. MEKC analysis of extracts of the nonhydrolyzed urines suggested the presence of the two different LOR glucuronides in the urine. The formation of the same two diastereoisomers was also observed in vitro employing incubations of LOR with human liver microsomes in the presence of uridine 5'-diphospho-glucuronic acid as coenzyme. The absence of other coenzymes excluded the formation of phase I or other phase II metabolites of LOR. Both results revealed a stereoselectivity, one diastereoisomer being formed in a higher amount than the other. After enzymatic hydrolysis using beta-glucuronidase, these peaks could not be detected any more. Instead, LOR was monitored. Analysis of the extracts prepared from enzymatically hydrolyzed urines by MEKC in the presence of 2-hydroxypropyl-beta-CD revealed the enantiomerization process of LOR (observation of two peaks of equal magnitude connected with a plateau zone). The data presented provide for the first time the evidence of the stereoselectivity of the LOR glucuronidation in humans.     

不同缓冲体系对毛细管电泳法分离15种核苷类化合物效果的比较

对毛细管电泳法分离15种核苷类化合物所用的不同缓冲液体系进行了系统比较,确定不同模式毛细管电泳法分析多种核苷类化合物的最适合背景缓冲液体系(BGE)。分别以四硼酸钠、磷酸氢二钠、乙酸钠、碳酸氢钠、乙酸铵和乙二胺(DEA)为背景电解质,对毛细管区带电泳(CZE)、毛细管电泳-电喷雾飞行时间质谱(CE-ESI-TOF/MS)以及胶束电动毛细管电泳(MEKC)3种模式进行比较,并对其中几种优势缓冲体系进行了优化。结果表明,CZE模式下使用四硼酸钠和磷酸氢二钠缓冲体系无法同时分离15种核苷类化合物,因此只适用于分析核苷类化合物数量较少的样品。而使用含有2%丙酮的300 mmol/L DEA能完全分开15种核苷类化合物,且分辨率和峰形良好。MEKC模式下,以25 mmol/L磷酸氢二钠(添加70 mmol/L十二烷基磺酸钠(SDS))为缓冲盐的分离结果最佳,并且此方法能成功应用于海洋生物海葵中核苷类化合物的分离。CE-ESI-TOF/MS分析中,以20 mmol/L乙酸铵(pH 10.0)为背景电解质,正离子模式检测,15种核苷类化合物的质谱信号均良好,检测灵敏度明显优于文献中报道的使用DEA缓冲体系的结果。本研究阐明了不同缓冲体系对15种核苷类化合物分离的适用性,为毛细管电泳技术在复杂基质中多种核苷类化合物的分离方法中的应用奠定了基础。     

Identification of kappa and lambda chains of the major immunoglobulin G subclasses by capillary zone electrophoresis

Immunoglobulins are present in most tissues and plasma and play crucial role in immune system. Alteration of the levels of the immunoglobulin G (IgG) subclasses (IgG1, IgG2, IgG3 and IgG4) is an indication of a disturbed immunological response. The aim of the present study was the development of a capillary electrophoresis (CE) method for the analysis of IgG subclasses in respect to their variable kappa and lambda chains. Various analytical conditions and CE modes, including capillary zone electrophoresis (CZE), capillary isoelectric focusing (CIEF) and micellar electrokinetic capillary chromatography (MECC) have been thoroughly studied. CZE was found to be the most convenient way to separate IgG subclasses. Three of the human IgG subclasses were resolved using uncoated fused-silica and 50 mM phosphate, pH = 9.3, as operating buffer at 20 kV and detection at 214 nm. IgG1kappa was completely separated from IgG2kappa and IgG3kappa, whereas IgG2kappa co-migrated with IgG4kappa, which is the minor IgG subclass. Under the same conditions IgG4lambda was completely separated from IgG1lambda, IgG2lambda and IgG3lambda, enabling the identification of the various lambda chains. The developed CE method is rapid and can be applied to the identification of the major immunoglobulin G subclasses in respect to their variable kappa and lambda chains.     

Quantitative gas chromatographic/mass spectrometric analysis of morphine glucuronides in human plasma by negative ion chemical ionization mass spectrometry

A sensitive and specific method for the determination of morphine glucuronides in human plasma is presented. Morphine glucuronides, namely morphine-6-glucuronide (M6G) and morphine-3-glucuronide (M3G), were extracted from plasma by solid-phase extraction on C(18) cartridges at pH 9.3 and derivatized to their pentafluorobenzyl ester trimethylsilyl ether derivatives. The compounds were measured by gas chromatography/negative ion chemical ionization mass spectrometry without any further purification. Using this detection mode, a diagnostic useful fragment ion at m/z 748 was obtained at high relative abundance for both target compounds. [(2)H(3)]-labeled morphine glucuronides were used as internal standards. Calibration graphs were calculated by polynomial fit within a range of 10-1280 and 15-1920 nmol l(-1) for the 6- and 3-glucuronide, respectively. At the limit of quantitation (LOQ), the inter-assay precision was 2.21% (M3G) and 2.23% (M6G) and the GC/MS assay variability was 1.8% (M3G) and 0.9% (M6G). The accuracy at the LOQ showed deviations of +4.92% (M3G) and +1.5% (M6G). The sample recovery after solid-phase extraction was 84.7% for both M3G and M6G. The method is rugged, rapid and robust and has been applied to the batch analysis of morphine glucuronides during pharmacokinetic profiling of the drugs.     

Qualitative determination of urinary morphine by capillary zone electrophoresis and ion trap mass spectrometry

A rapid, sensitive method for the determination of morphine and amphetamine was developed using capillary zone electrophoresis coupled with electrospray interface (ESI), ion-trap tandem mass spectrometry (CE-ESI-MS2). Morphine and amphetamine were separated in 20 mM ammonium acetate buffer (pH 6.6) and detected by ion-trap mass detector in different analytical time segments (0-6.25 min for amphetamine and 6.25-12.0 min for morphine) in which the tune file for each compound was used separately. Molecular ions of morphine (m/z 286) and amphetamine (m/z 136) were detected at 5.77 and 6.83 min, respectively, while product ions of MS2 for each compound (m/z 229, 201 for morphine and m/z 119 for amphetamine) were detected almost exactly at the same time with their parent compounds. The limits of detection (LOD) for MS2 determination were 30 and 50 ng/mL for amphetamine and morphine, respectively, with an S/N ratio of 3. For more sensitive detection of morphine, the sample was injected for a longer time (i.e., 80 s) and hydrodynamically transported into a CE capillary for MS detection. Morphine and its product ion appear at 0.36 and 0.39 min on the ion chromatogram, respectively, with a five-fold increase of detection sensitivity (LOD, 10 ng/mL). The CE-MS system thus established was further applied for forensic urine samples screened as morphine-positive by fluorescence polarization immunoassay (FPIA). These results indicated the feasibility of CE-ESI-MS2 for confirmative testing of morphine in urine sample.     

Determination of ethyl sulfate in human serum and urine by capillary zone electrophoresis

The use of capillary zone electrophoresis (CZE) with indirect absorbance detection for the analysis of ethyl sulfate (EtS) in serum and urine was investigated. EtS is a direct metabolite of ethanol employed as marker for recent alcohol consumption. Fused-silica capillaries of 60 cm total length were either coated with cetyltrimethylammonium bromide (CTAB, 50 microm I.D. capillary) or poly(diallyldimethylammonium chloride) (PDADMAC, 100 microm I.D. capillary) to allow CZE analyses to be performed with reversed polarity. At pH 2.2 with a maleic acid/phthalic acid background electrolyte, both approaches provided reliable EtS serum levels down to 0.2 mg L(-1) (1.6 microM) for the analysis of solid-phase extracts that were prepared after chloride precipitation. Analysis of urines diluted to a conductivity of 5 S m(-1) and analyzed in the two capillary formats resulted in limits of quantification (LOQs) of 2 and 1 mg L(-1), respectively. With urines adjusted to 10 S m(-1) via dilution or condensation, an LOQ of 0.6 mg L(-1) (4.8 microM) was obtained in the CTAB coated capillary whereas in the PDADMAC-coated capillary of equal length not all matrix components were resolved from EtS. The developed assays are robust and suitable to monitor EtS in samples of individuals who consumed as little as one standard drink of an alcoholic beverage containing about 14 g of ethanol.     

Application of capillary electrophoresis to the determination of various benzylpenicillin salts

The application of capillary electrophoresis for separation of benzylpenicillin, procaine, benzathine and clemizole was investigated. Phosphate-borate buffer supplemented with sodium dodecyl sulphate 14.4 g/l and electrophoresis voltage 18 kV seem to provide optimal conditions for micellar electrokinetic chromatographic assay of penicillin salts. This method is selective and precise. The results obtained from CE method recovery assay (above 98% for all but procaine-97% substances) and from determination of benzylpenicillin by CE compared with HPLC results, confirmed good accuracy.     

Separation and characterization of humic acids from Antarctica by capillary electrophoresis and matrix-assisted laser desorption ionization time-of-flight mass spectrometry. Inclusion complexes of humic acids with cyclodextrins

A new capillary electrophoresis procedure based on micellar electrokinetic capillary chromatography for the separation of humic acids (HAs) isolated from Antarctica soil was developed. The HAs were separated and characterized using a background electrolyte containing 0.09 M borate+0.09 M Tris+0.001 M EDTA (BTE) of pH 8.3, modified with alpha-, beta-, or gamma-cyclodextrins (CDs) and sodium dodecyl sulfate. It was found that from alkaline solution of HAs in the presence of CDs, the HAs are not completely precipitated with a strong acid and a certain part (some fractions) remains soluble. Mass spectrometry shows that HAs contain 15-25 simple low-Mr compounds and several families of compounds with similar structure (m/z approximately 800-1200). Comparison of HA analysis from Antarctica soil with those of soil HAs from the American continent show a high similarity between the samples and confirm several identical compounds and some with very similar structural units.     

Key fragments for identification of positional isomer pair in glucuronides from the hydroxylated metabolites of RT-3003 (Vintoperol) by liquid chromatography/electrospray ionization mass spectrometry.

The mass spectral properties of glucuronides of the 9- and 10-hydroxylated metabolites of RT-3003 (Vintoperol; (-)-1beta-ethyl-1alpha-hydroxymethyl-1,2,3,4,6,7, 12balpha-octahydroindolo[2,3-a]quinolizine), which were fractionated by high-performance liquid chromatography with fluorescence detection, were investigated using the positive ion electrospray ionization mode. These glucuronides showed predominantly the protonated molecular ion ([M + H](+) ion), and the [M + H](+) ion provided a characteristic product ion spectrum in which abundant ions were obtained at m/z 301, 160 and 142. The first ion, corresponding to the [aglycone + H](+) ion, was produced by neutral loss of the glucuronic acid moiety from the [M + H](+) ion. The product ion spectrum of the [M + H](+) ion of hydroxy-RT-3003 revealed a number of ions common to the glucuronide spectra, suggesting that other two ions observed most likely represent fragmentation of hydroxy-RT-3003. In turn, these glucuronides were positional isomers with respect to the binding site of glucuronic acid. The structures of the isomer pairs were discriminated by the presence of the ion of m/z 318 or 336 in the product ion spectrum. These ions were produced by fission of the C-ring, the same as for the formation of the ions of m/z 160 and 142, as were observed in the product ion spectrum from the [M + H](+) ion of hydroxy-RT-3003. For the formation of these ions, an unusual fragmentation process was proposed, and these ion structures were supported by evidence from the accurate mass measurement data. Additionally, in the sulfates of hydroxylated metabolites, a similar product ion corresponding to the ion of m/z 336 found in the phenolic glucuronides was observed, and was applied for identification of the sulfate metabolites.     

Possibilities and limitations of different modes of capillary electrophoresis for the quantitative determination of catechols and caffeine in black and green tea

Methods of capillary electrophoresis (capillary zone electrophoresis, micellar electrokinetic chromatography, and microemulsion electrokinetic chromatography) were compared in determining polyphenolic antioxidants and alkaloids in black and green tea. It was shown that reverse-flow micellar electrokinetic chromatography (RF-MEKC, pH < 2.5) using reversed pola rity is most preferable for the rapid simultaneous determination of catechols and caffeine. The detection limit was 1 mg/L.     

On-line sample concentration in micellar electrokinetic chromatography with cationic micelles in a coated capillary

The electroosmotic flow was successfully suppressed even in the presence of cationic surfactants, when a polyacrylamide-coated capillary was employed. Two on-line sample concentration techniques of sample stacking and sweeping were evaluated in micellar electrokinetic chromatography (MEKC) using the polyacrylamide-coated capillary. Cationic surfactants were used as pseudostationary phases in MEKC. At least 60-fold and about 600-fold increases in detection sensitivity were obtained in terms of peak heights by sample stacking and sweeping, respectively.     

Determination of enzymatic activity and properties of secretory phospholipase A2 by capillary electrophoresis.

A capillary electrophoretic (CE) system coupled with a diode array UV detector was used for the assay of secretory phospholipase A2 (sPLA2) activity. This method is based on monitoring both the breakdown of substrates and the formation of products simultaneously using micellar electrokinetic chromatographic techniques. Under our developed separation conditions, we analyzed the substrates and products quantitatively, and investigated enzyme activity as a function of reaction time and presence of enzyme activator or inhibitor. Matrix-assisted laser desorption/ionization time-of-flight mass spectrometry was also utilized to confirm the phosphatidylcholine, a substrate of sPLA2. In order to test the feasibility of the developed method for measurement of enzymatic activity, we compared it to the conventional radioactive assay method for sPLA2. On the basis of our results, the conventional method can be complemented, or even replaced, by this new CE method which possesses the advantages of short analysis time, use of non-radiolabeled and inexpensive substrates, simple measurement of enzymatic activity, and exact quantitation of substrate and product.     

Total steroid profiling with capillary GC. Details on sample preparation

The accuracy and reproducibility of steroid and steroid conjugate determinations using capillary GC are evaluated. It is possible to analyze many unconjugated steroids, steroid sulfates, and some steroid glucuronides simultaneously, simply by using group fractionation based on successive hydrolysis and solvent extraction steps. However, a reliable determination of corticosteroid glucuronides can only be obtained after previous isolation of the glucuronides as a group, with ion exchange chromatography. In complex chromatograms, the combined use of nitrogen specific detection and flame ionization detection facilitates peak identification.     

In‐capillary approach to eliminate SDS interferences in antibody analysis by capillary electrophoresis coupled to mass spectrometry

Capillary electrophoresis is an important technique for the characterization of monoclonal antibodies (mAbs), especially in the pharmaceutical context. However, identification is difficult as upscaling and hyphenation of used methods directly to mass spectrometry is often not possible due to separation medium components that are incompatible with MS detection. Here a CE‐MS method for the analysis of mAbs is presented analyzing SDS‐complexed samples. To obtain narrow and intensive peaks of SDS‐treated antibodies, an in‐capillary strategy was developed based on the co‐injection of positively charged surfactants and methanol as organic solvent. For samples containing 0.2% (v/v) of SDS, recovered MS peak intensities up to 97 and 95% were achieved using cetyltrimethylammonium bromide or benzalkonium chloride, respectively. Successful removal of SDS was shown in neutral coated capillaries but also in a capillary with a positively charged coating applying reversed polarity. The usefulness of this in‐capillary strategy was demonstrated also for other proteins and for antibodies dissolved in up to 10% v/v SDS solution, and in other SDS‐containing matrices, including the sieving matrix used in a standard CE‐SDS method and gel‐buffers applied in SDS‐PAGE methods. The developed CE‐MS approaches enable fast and reproducible characterization of SDS‐complexed antibodies.     

毛细管电泳在线聚焦原理、技术及应用

毛细管电泳以其分析速度快、分离效率高、操作简便、能够实现高通量而获得了广泛的应用,但由于检测窗口小而导致其检测灵敏度低。为了提高检测灵敏度,目前已发展了多种毛细管电泳在线聚焦和样品预浓缩技术,如场放大样品堆积、pH调节浓缩、胶束电动毛细管色谱、等速电泳等。这些技术由于能够在毛细管内同时实现样品的聚焦和分离、操作简便而获得了广泛的兴趣和关注。本文针对毛细管电泳的在线聚焦的原理、技术和应用做一简要的介绍和总结。