Capillary electrochromatographic analysis of aliphatic mono- and polycarboxylic acids

The parameters influencing the electrochromatographic separation of aliphatic organic acids in a capillary column with a wall-coated macrocyclic polyamine have been studied. Indirect detection using chromate, pyromellitate, trimellitate, o-phthalate, benzoate and acetate as background electrolytes has been tested. A complete separation of polyprotic acids could be achieved with pyromellitate buffer (7.5 mM, pH 6.5), and satisfactory results for the simultaneous separation of monoprotic acids and polyprotic acids were found using a capillary column of 70 cm (50 cm effective length)x75 microm inner diameter, electrokinetic injection (-10 kV, 10 s), benzoate buffer (6 mM, pH 4.6), separation voltage of -10 kV, and detection at 220 nm. For the separation of the geometric isomers fumarate and maleate, acetate buffer was found the best choice among the background electrolytes tested. The method so established has been applied to the determination of organic acids in soy sauce, brandy, lemon juice, spinach juice and cigarette. From the retention behavior, it was found that the separation mechanism on the bonded phase was influenced by the macrocyclic effect, electrostatic attraction, hydrogen bonding, van der Waals forces, and anion exchange, in addition to the differences in electrophoretic mobility.     

Nucleoside monophosphates recognition using macrocyclic polyamine bonded phase in capillary electrochromatography

An open-tubular wall-coated macrocyclic polyamine capillary column (70 cm x 75 microm ID) with 50 cm effective length for the separation of nucleoside monophosphates is described. Some parameters with respect to concentration, pH, composition of the buffer, and voltage in order to optimize the separation were studied. The coated capillary showed reversed electroosmotic flow (EOF), allowing anions to be separated in the co-EOF mode. Baseline separations were achieved for the eight nucleotides in less than 26 min using a background electrolyte consisting of H(3)PO(4)-NaH(2)PO(4) (30 mM, pH 3.10), an applied voltage of -15 kV, and detection at 254 nm. The macrocyclic polyamine on the capillary wall introduced anion coordination for the interaction with the analytes, the strength of which could be moderated by the type and concentration of the competing ion used in the background electrolyte (BGE). With a low concentration of the competing ion (phosphate ion), the migration behavior followed that obtained in the electrophoretic system. Increasing the concentration of the competing ion resulted in a faster migration and more complete elution of the analyte. The method established was also employed for the analysis of nucleotides in mushrooms. Aqueous extracts of mushrooms from different species and various extraction methods were injected directly for the analysis. Uridine 5'-monophosphate, guanosine 5'-monophosphate, adenosine 5'-monophosphate, and cytidine 5'-monophosphate, were found in the sample tested.     

Gold nanomaterials based pseudostationary phases in capillary electrophoresis: A brand‐new attempt at chondroitin sulfate isomers separation

In this work, a CE method with bare gold nanorods (GNRs) based pseudostationary phase was developed and applied for the separation of chondroitin sulfate (CS) isomers, CS, and dermatan sulfate (DS). The separation efficiency was investigated by varying the experimental parameters such as concentration and pH of the BGE, separation voltage, internal diameter of capillary, different size, and morphology of gold nanomaterials. Results showed that different size and morphology of gold nanomaterials had different effects on the separation of CS and DS. The best separation of CS and DS was achieved in the BGE composed of aqueous 150 mmol/L (mM) ethylenediamine + 20 mM sodium dihydrogen phosphate + 30% v/v GNRs, pH 4.5, at the separation voltage of ?10 kV. Capillary was 59.2 cm in length (effective length 49 cm), 50 μm id capillary thermostated at 25°C. CE with bare GNRs used as pseudostationary phase was shown to be a suitable technique for the separation of CS and DS mixtures with wider peaks. RSD of migration time and peak area of CS and DS were 0.13, 0.14 and 0.86, 1.07%, respectively.     

逍遥丸的毛细管电泳指纹图谱的建立

采用毛细管区带电泳法建立了逍遥丸(Xiaoyao Pill, XYP)的毛细管电泳指纹图谱(CEFP)。运用正方形优化法,以色谱指纹图谱分离量指数(RF)为优化的目标函数,对建立指纹图谱的实验条件进行了优化,确定了最佳背景电解质(BGE)溶液50 mmol/L硼砂-50 mmol/L磷酸氢二钠-150 mmol/L磷酸二氢钠-50 mmol/L碳酸氢钠(1:1:1:5, v/v/v/v; pH 7.40)、紫外检测波长228 nm、运行电压12 kV、重力进样25 s (高度14 cm)的分离检测条件。采用未涂层石英毛细管(70 cm×75 μm,有效分离长度57 cm)分离,以咖啡酸色谱峰为参照,确定13批逍遥丸样品的21个共有指纹峰。通过聚类分析确定用其中10批样品生成对照CEFP,以此为标准用系统指纹定量法鉴别13批逍遥丸的质量,结果显示: S3号样品的化学成分数量和分布比例不合格,S10和S12号样品含量明显偏高,其余批次质量均合格。所建立的正方形优化法操作简便,适用于中药的毛细管区带电泳BGE的选择;所建立的逍遥丸CEFP具有较好的精密度和重现性,可以为逍遥丸的质量控制提供新的参考。     

Tungstate as complex-forming reagent facilitating separation of selected polyphenols by capillary electrophoresis and its comparison with borate

A novel electrophoretic BGE containing tungstate as complex-forming reagent is suitable for the separation of polyphenols. Similar to molybdate-containing BGE reported earlier (cf. M. Polásek, et al.., Talanta 2006, 69, 192) addition of tungstate to BGE affects significantly migration of compounds/ligands with vicinal -OH groups due to the formation of negatively charged complexes involving W(VI) as central ion. Baseline separation of mixtures of flavonoids (apigenin, luteolin, hyperoside, quercetin, and rutin) and phenolic acids (chlorogenic and p-coumaric acid) was achieved within 15 min with optimized BGE of pH 7.4 containing 50 mM N-(2-hydroxyethyl)piperazine-2'-(2-ethanesulfonic acid) (HEPES), 2.2 mM tungstate, and 25% v/v of methanol. The separation was performed in a 75 cm (effective length 42 cm)x 75 microm id uncoated fused-silica capillary at 30 kV with spectrophotometric detection at 275 nm. The calibration curves were rectilinear for 25-175 microg/mL of all analytes (cinnamic acid as the internal standard). The LODs ranged from 1.8 to 6 microg/mL for all analytes except for chlorogenic acid. Intraday precision (n = 6) of migration times (RSD < or = 1.2%) and peak areas (RSD < or = 5.6%) was evaluated. The tungstate-based BGEs can be alternatively utilized for the analysis of polyphenols at considerably lower pH than with conventional alkaline borate-based BGEs.     

毛细管区带电泳法分离白花丹参中主要的水溶性活性成分

为建立一种快速分离白花丹参水溶性有效成分的毛细管区带电泳体系,分别考察了缓冲液浓度、缓冲液pH、运行电压、检测波长对样品的分离度、迁移时间等因素的影响。最终优化的分离条件为:5 mmol/L硼砂缓冲液（pH 7.5）;毛细管柱75 μm×60.2 cm,有效长度50 cm,压力进样(3.45 kPa×4 s),27.5 kV恒压分离,210 nm波长下检测,柱温25 ℃。在优化的条件下,8 min内使白花丹参样品中的原儿茶醛、丹参素、原儿茶酸组分达到完全基线分离。     

Optimization of the chiral resolution of baclofen by capillary electrophoresis using beta-cyclodextrin as the chiral selector

The chiral resolution of baclofen was achieved by capillary electrophoresis using a fused-silica capillary (60 cm x 75 microm ID). The background electrolyte (BGE) was phosphate buffer (pH 7.0, 50 mM)-acetonitrile (95:5 v/v) containing 10 mM beta-cyclodextrin. The applied voltage was 15 kV. The values of alpha and R(s) were 1.06 and 1.00, respectively. The electrophoretic conditions were optimized varying the pH and the ionic strength of the BGE, concentrations of beta-cyclodextrin and acetonitrile and the applied voltage.     

柏子养心丸的毛细管电泳指纹图谱

建立了柏子养心丸(Baizi Yangxin Wan, BZYXW)毛细管区带电泳指纹图谱(CEFP)。采用三棱柱优化法优化背景电解质(BGE),以色谱指纹图谱分离量指数(RF)为实验条件优化的目标函数,最终确定BGE为50 mmol/L硼砂-50 mmol/L磷酸氢二钠-200 mmol/L硼酸-150 mmol/L碳酸氢钠(体积比为7:7:1:1,含4%乙腈,pH 9.70)。在其他优化的毛细管电泳条件为紫外检测波长228 nm,运行电压12 kV,重力进样25 s (高度14 cm)条件下,采用未涂层石英毛细管(70 cm(有效分离长度57 cm)×75 μm)分离,以阿魏酸峰为参照,确定了17个共有指纹峰。对样品聚类分析后用其中9批生成对照CEFP(RCEFP),以其为标准,用系统指纹定量法鉴别出12批BZYXW质量为: 3批好,1批良好,3批中,1批一般,4批劣。三棱柱优化法为BGE选择提供了重要参考,建立的BZYXW-CEFP精密度好、重现性高,可用于BZYXW的质量控制。     

A capillary zone electrophoresis for determination of thiolic peptides in biological samples

A new method to improve the analyses of thiolic peptides (cysteine, γGlu-Cys, glutathione, phytochelatins and desglycyl-phytochelatins) derivatized with monobromobimane (mBrB) in complex biological samples by CZE is described. The method involves a SPE using Sep-Pak Light C18 Cartridges after derivatization and a later CZE analysis. Elution of mBrB-thiols was achieved with 10 mM HCl + 70% methanol v/v in deionised water. Electrophoretic parameters, such as BGE pH and concentration, different organic additives (methanol and trifluoroethanol), applied voltage and capillary length were studied in order to establish suitable analytical conditions. Optimum separation of the mBrB-thiolic peptides was obtained with 100 mM sodium borate buffer at pH 7.60. The electrophoretic conditions were +15 kV, capillary length of 90 cm from inlet to detector (98 cm total length, 50 μm ID), samples were loaded into the capillary by hydrodynamic injection (50 mbar, 20 s) and detection was performed at 390 nm. The improved method showed good reproducibility, linearity and sensitivity. The LODs and LOQs estimated using a standard of GSH were 1.41 and 4.69 μM respectively.     

Application of reverse-flow micellar electrokinetic chromatography for the simultaneous determination of flavonols and terpene trilactones in Ginkgo biloba dosage forms

A reverse-flow micellar electrokinetic chromatographic (RF-MEKC) method was developed for the simultaneous qualitative determination of 10 components consisting of the flavonol glycosides, rutin and quercitrin, the flavonol aglycones, isorhamnetin, kaempferol and quercetin, the terpene trilactones, ginkgolides A, B, C and J and the sesquiterpene, bilobalide. This method was used to fingerprint Ginkgo biloba solid oral dosage forms and validated for the quantitation of the marker compounds, rutin and quercetin in some commercial products. In addition to the usual variables, the influence of some essential background electrolyte (BGE) components such as sodium dodecyl sulphate (SDS) and -cyclodextrin concentrations were investigated. A polyimide fused-silica square capillary column (75 microm I.D. x 360 microm O.D.) with a total length of 60.0 cm and effective length of 45.0 cm was used for the separation. The final BGE consisted of 20 mM phosphoric acid, 40 mM SDS and 12 mM -cyclodextrin (pH 2.2) using reverse polarity with a voltage of -17.5 kV. Samples were injected electrokinetically at -5 kV for 3 s for the qualitative analysis and hydrodynamically at 20 mbar for 0.6 s for the quantitative assay. The total run time was 22 min and the limits of detection were 3.13 microg/ml and 1.88 microg/ml for rutin and quercetin, respectively. Fingerprint profiles of the solid oral dosage forms and the results of the quantitative analysis indicated that there were major discrepancies in the marker content between products and illustrates the value of this method for use as a procedure to assess product quality of commercially available Ginkgo biloba products.     

胶束电动毛细管色谱法分析头孢哌酮与其S-异构体等杂质

以十二烷基硫酸钠(SDS)胶束为准固定相,考察了头孢哌酮、头孢哌酮S-异构体、头孢哌酮杂质A及其他未知杂质在胶束电动毛细管色谱(MECC)分离模式下的分离行为。研究了运行缓冲液的pH值、磷酸盐浓度、SDS浓度、甲醇体积分数、分离电压、分离温度等因素对头孢哌酮、S-异构体、头孢哌酮杂质A及其他杂质的迁移时间、分离度以及可分离出的杂质个数的影响。结果发现,这些因素对头孢哌酮与诸杂质间的分离及检测有显著的影响,尤以pH值为最。它不仅影响它们的迁移时间和分离效率,还直接影响头孢哌酮及其杂质峰的检测。优化后的分离条件:运行缓冲液为70 mmol/L磷酸盐-100 mmol/L SDS (pH 6.5),分离电压为15 kV,分离温度为25 ℃。在此条件下,用非涂渍石英毛细管51.0 cm×75 μm(有效长度42.5 cm),压力进样5 kPa×5 s,在254 nm波长下进行检测,可分离出28个杂质,诸杂质彼此间及与头孢哌酮间可得到有效分离。并将该方法成功地用于测定注射用头孢哌酮钠的含量和有关物质,结果令人满意。     

糖类衍生物在毛细管区带电泳下的分离研究

以新合成的1-萘基-3-甲基-5-吡唑啉酮(NMP)为柱前衍生试剂,采用毛细管区带电泳模式考察并优化了糖类衍生物的分离条件。实验采用58.5cm×50μmi.d.毛细管(有效柱长50cm),55mmol/L硼酸盐缓冲溶液(pH9.46),柱温20℃,分离电压22kV,进样10s,在不加任何添加剂的情况下,高效、快速地实现了9种糖的基线分离,并在最优化条件下进行了唐古特白刺实际样品的分离分析,结果令人满意。     

Simultaneous determination of erlotinib and metabolites in human urine using capillary electrophoresis

The purpose of this study was to develop a simple and sensitive CE‐UV method to quantify erlotinib and metabolites in urine. Following liquid–liquid extraction, erlotinib, and metabolites were separated with a BGE whose composition was phosphate buffer (pH 2.5, 65 mM) with 0.5% Tween 20. The applied voltage was 22 kV, capillary temperature 25°C and the sample injection was performed in the hydrodynamic mode. All the analyses were carried out in a fused silica capillary with an internal diameter of 75 μm and a total length of 37 cm. The detection of target compounds was performed at 240 nm. The calibration was linear in the range 0.15–20 mg/L for erlotinib and metabolites. Inter‐and intraday imprecision were less than 4%. This simple, sensitive, accurate, and cost‐effective method can be used in routine clinical practice to monitor erlotinib concentrations in urine from nonsmall cell lung cancer patients.     

Development of a gas diffusion probe for the rapid measurement of pCO2 in aquatic samples

A capillary electrophoresis (CE) procedure with contactless conductivity detection (C(4)D) has been developed for monitoring of neutral mono- and disaccharides in drinks and foodstuffs. The separation of a mixture of seven neutral saccharides (glucose, fructose, galactose, mannose, ribose, sucrose and lactose) employed a quartz capillary, 5 μm i.d., with an effective length of 18.3 cm, and 75 mM NaOH (pH 12.8) as the background electrolyte (BGE). The limit of detection (LOD) values obtained lied within a range from 0.4 μmol L(-1) for lactose to 0.9 μmol L(-1) for ribose, with a separation time shorter than 140 s. The procedure was successfully applied to determinations of saccharides in fruit juices, Coca-Cola, milk, red and white wines, yoghurts, honey and a foodstuff additive.     

藏药蕨麻多糖水解单糖的毛细管区带电泳分离研究

以1-萘基-3-甲基-5-吡唑啉酮(NMP)为柱前衍生试剂,探讨了毛细管区带电泳模式下对藏药蕨麻多糖水解液中单糖的分离条件。实验采用58.5 cm×50μm i.d.毛细管(有效长度50 cm),55 mmol/L硼酸盐缓冲溶液(pH=9.46),柱温20℃,分离电压22 kV,进样10 s。该法不加任何添加剂,9种单糖可高效、快速基线分离,实现了对藏药蕨麻多糖水解液中单糖的分离和定量分析,结果令人满意。     

毛细管电泳法测定阿霉素脂质体药物中的微量硫酸根离子

建立了以硝酸钾作为背景电解质测定阿霉素脂质体药物中微量硫酸根离子的毛细管电泳分析法。考察了分离电压、背景电解质、电渗流改性剂浓度、pH值对分离测定的影响。结果表明,当毛细管长度为60 cm(有效长度51.5 cm)、分离电压为~15 kV、缓冲溶液采用20 mmol/L硝酸钾(pH 7.0)、电渗流改性剂采用0.4 mmol/L十六烷基三甲基氯化铵(CTAC)、检测波长为202 nm时,阿霉素脂质体破乳液中硫酸根离子和氯离子在3 min内得到了基线分离,硫酸根离子迁移时间和峰面积的相对标准偏差分别小于0.01%和1.0%,检出限为5 μg/L。用该方法对阿霉素脂质体样品中的微量硫酸根离子进行了分析测定,结果令人满意。     

1 μm硅胶颗粒固定相的加压电色谱分离性能

制备了1 μm无孔硅胶颗粒。通过电动填充法得到总长度为45 cm(固定相填充长度为20 cm)、内径为100 μm的毛细管色谱柱。以乙腈-水体系作为流动相,详细考察了碱性化合物在该色谱柱上的加压电色谱(pCEC)分离性能,讨论了流动相比例、缓冲液浓度、pH值及操作电压等因素对分离的影响。实验结果表明,裸硅胶柱在乙腈-水体系分离碱性样品中表现出典型的反相色谱分离性能;缓冲液浓度的改变则对分离影响不大。当pH值改变时,碱性化合物的解离程度发生变化,它们与固定相之间的作用力发生变化,使得分离度发生相应的变化。分离柱效随施加电压的增加而增加,在1 kV电压下,裸硅胶柱对邻甲苯胺的柱效为35000理论塔板/m。     

毛细管电泳法同时测定血清中的左旋多巴和甲基多巴

建立了毛细管电泳-紫外检测同时测定血清中左旋多巴和甲基多巴的方法。以40 mmol/L硼砂（pH 9.5）为分离缓 冲溶液,在3.45 kPa（0.5 psi）压力下进样7 s、分离电压22 kV、检测波长200 nm、温度25 ℃的条件下进行测定,两 种物质获得了较好的分离。甲基多巴和左旋多巴分别为1.0~64.0 mg/L和1.0~71.0 mg/L时与峰面积呈良好的线性关系, 线性相关系数分别为0.9998和0.9994,检出限分别为0.6和0.8 mg/L(以信噪比为3计)。将该法用于血清中甲基多巴和左 旋多巴的测定,回收率为82.8%~88.8%,相对标准偏差为2.10%~2.63%。     

高效毛细管电泳法同时测定多种肽类药物的方法研究

  以血管紧张素Ⅰ,Ⅱ,Ⅲ和P物质、神经激肽、生 长激素释放的抑制因子、神经紧张肽等7种肽类药物混合物为测定对象,用胺涂层的毛细管 柱(57 cm×75 μm i．d．,有效长度为50 cm)对上述混合物的分离条件进行了研究。在 f工作电压为10 kV［（－）→(＋)］、电流为42 μA、检测波长为214 nm、测定温度为2 5 ℃、虹吸进样10 s的条件下,在50 mmol/L的醋酸铵缓冲液(pH 4.5)中,上述混合物可在8 min内得以完全分离。 关键词：     

Validation and Further Optimisation of a Cyclodextrin-Modified Micellar Electrokinetic Capillary Chromatography Method for Urine Profiling

Metabonomic studies require efficient and high-resolution analytical probes to monitor changes in the ‘metabolic fingerprint’. The advantageous characteristics of Capillary Electrophoresis, enabling highly efficient separations of diverse components present in minute sample volumes, may therefore prove a useful tool in biofluid analysis. This paper describes the optimisation and validation of a sulphated β-cyclodextrin-modified MECC method for urine profiling. Cyclodextrin substitution, experimental conditions including capillary length, injection mode and time, applied voltage, temperature and capillary pre-conditioning procedures were investigated and optimised. Precision, linearity, sensitivity and robustness of the method were assessed, as well as urine stability. The validated sulphated β-cyclodextrin-modified MECC method allows for the separation of over 80 urinary analytes in under 25 min, using a sodium borate/SDS/sulfated β-cyclodextrin (25/75/6.25 mM) electrolyte and an 18 kV applied voltage in a 40 cm (effective length), 50 μm i.d. fused silica capillary at 20 °C, pre-conditioned with HCl for 5 min and BGE for 1 min, and UV diode array detection (190–600 nm). Such methodology should prove invaluable in the rapid comparison of urine profiles and indication of metabolic disorders or abnormalities.