Determination of nikethamide by micellar electrokinetic chromatography

A simple, fast, sensitive and reproducible micellar electrokinetic chromatography (MEKC)–UV method for the determination of nikethamide (NKD) in human urine and pharmaceutical formulation has been developed and validated. The method exhibits high trueness, good precision, short analysis time and low reagent consumption. NKD is an organic compound belonging to the psychoactive stimulants used as an analeptic drugs. The proposed analytical procedure consists of few steps: dilution of urine or drug in distilled water, centrifugation for 2 min (12,000 g ), separation by MEKC and ultraviolet‐absorbance detection of NKD at 260 nm. The background electrolyte used was 0.035 mol/L pH 9 borate buffer with the addition of 0.05 mol/L sodium dodecyl sulfate and 6.5% ACN. Effective separation was achieved within 5.5 min under a voltage of 21 kV (~90 μA) using a standard fused‐silica capillary (effective length 51 cm, 75 μm i.d.). The determined limit of detection for NKD in urine was 1 μmol/L (0.18 μg/mL). The calibration curve obtained for NKD in urine showed linearity in the range 4–280 μmol/L (0.71–49.90 μg/mL), with R² 0.9998. The RSD of the points of the calibration curve varied from 5.4 to 9.5%. The analytical procedure was successfully applied to analysis of pharmaceutical formulation and spiked urine samples from healthy volunteers.     

Determination of heterocyclic compounds by micellar electrokinetic capillary chromatography

Micellar electrokinetic capillary chromatography (MECC) based on sodium cholate (NaCh) and sodium dodecyl sulphate (SDS) was developed for the determination of aromatic amino acids and heterocyclic legume constituents. The influence of temperature, voltage, micellar system, pH, zwitterion and modifier concentrations in the buffer on migration times, peak areas, resolution and number of theoretical plates was investigated. This MECC method makes possible the sensitive determination of the individual compounds with detection limits in the picomole range. Up to 300 000 theoretical plates per metre of capillary were obtained together with satisfactory linearity and repeatability of the NaCh method. The applicability of MECC to samples prepared from plant material, following a fast and simple technique of isolation, purification and group separation, is illustrated by selected examples.     

Determination of plasma heparin by micellar electrokinetic capillary chromatography

The micellar electrokinetic capillary chromatography (MECC) method is reported for the separation of heparin, and for the possibility of direct determination of free heparin in plasma. The conditions for MECC were: pH 8.5, 25 mM sodium dodecyl sulfate (SDS), 25 mM borate buffer, with a 30 cmx50 mum ID fused-silica capillary. The sample was detected with a UV-detector at 270 nm with heparin as external standard. The recovery rate was 95.6-98.7%. This method was linear in the range 80-7000 U l(-1). The within-run and between-run relative standard deviations were lower than 3.1 and 4.5%, respectively. It is suggested that this MECC method may be used to determine blood samples containing high levels of heparin.     

Determination of thyreostatics in animal feed by micellar electrokinetic chromatography

The determination of the thyreostatics 2-thiouracil, its derivatives (4-methyl-2-thiouracil, 4-propyl-2-thiouracil and 4-phenyl-2-thiouracil) and methimazole in manufactured dried animal feed by micellar electrokinetic chromatography (MEKC) is described. A 99 +/- 5% extraction yield at the 20 micrograms g-1 level (n = 8) was achieved by shaking the milled fodder with methanol-1 M NaOH (80 + 20). Aliquots of the supernatant were injected in a 75 microns x 33.5 cm uncoated silica capillary using pressure; separation was performed at 23 degrees C with 15 kV (positive polarity) in a background electrolyte (BGE) containing 40 mM sodium dihydrogenphosphate, 50 mM sodium dodecyl sulfate and 15 mM Tween 20 at pH 9. When the surfactants were added to the BGE, all the thyreostatics were well resolved and the fodder extracts showed lower backgrounds. The peaks appeared within the 2.25-5.2 min range with efficiencies in the 2.5 x 10(4)-8 x 10(4) range; methimazole appeared in the vicinity of the electroosmotic migration time. Calibration curves were linear within the studied range (20-200 micrograms ml-1, r2 > 0.998). Limits of detection in the extracts of spiked fodder samples ranged from 0.25 to 0.4 microgram ml-1, which corresponded to 0.6-1.0 microgram of drug per gram of fodder. Peak area repeatabilities were about 4% at the 20 micrograms ml-1 level.     

Determination of steroids in biological samples by micellar electrokinetic chromatography

The possibility of determining eight adrenocorticotropic steroid hormones (cortisol, cortisone, corticosterone, 11-dehydrocorticosterone, 17-hydroxyprogesterone, 11-deoxycortisol, and progesterone) by micellar electrokinetic chromatography (MEKC) using urea as an organic additive to the working electrolyte (a 25 mM phosphoric acid solution and 10 mM sodium dodecyl sulfate) was demonstrated. The use of online preconcentration (stacking and sweeping) allowed us to lower the detection limit for steroids to ～3 ng/mL. The total analysis time was 15 min.     

Determination of 20-hydroxyecdysone content by thin-layer chromatography and micellar electrokinetic chromatography

Summary Seasonal dependence of 20-hydroxyecdysone content ofSerratula tinctoria andSerratula wolffii (Asteraceae) was investigated by micellar electrokinetic chromatography (MEKC). Samples were collected each month through the vegetation period. The leaves were dried, milled and extracted with methanol. Clean-up of the extracts was by solid-phase extraction using a polyamide micro-column to remove flavonoids and other plant phenolics which can interfere with the analysis. This work deals with the separation of 20-hydroxyecdysone from polypodine B and the seasonal variation of 20-hydroxyecdysone concentration. Determinations have been performed by both thin-layer chromatography and capillary electrophoresis using micellar electrokinetic chromatography. Presented at: Balaton Symposium on High-Performance Separation Methods, Siófok, Hungary, September 3–5, 1997.     

场放大进样-胶束扫集毛细管电脉法测定痕量泼尼松

建立了胶束毛细管电动色谱在线富集技术测定药品中痕量的泼尼松的方法。在胶束扫集的基础上联用场放大进样,使泼尼松的富集倍数提高了136倍;检出限由原来的2.7mg/L降至20μg/L。胶束扫集毛细管电泳缓冲体系为120mmol/LSDS、10mmol/LNaH2PO4(pH2.5)10%乙腈(V/V)。分离电压-20kV,进样电压-20kV,进样时间70s,进水时间180s,检测波长250nm。同时讨论了SDS浓度、样品基质pH、进样电压、进水时间和进样时间对分离效果的影响。实验结果显示:在优化实验条件下,样品的检测仅需8min,泼尼松在0.05～10mg/L的范围内线性关系良好(r=0.998)。回收率在89.4%～106%之间,相对标准偏差在2.1%～2.6%之间,可用于各种中药制剂中泼尼松的含量测定。     

Determination of piribedil in pharmaceutical formulations by micellar electrokinetic capillary chromatography

A fast and simple micellar electrokinetic capillary chromatographic method was developed for the analysis of piribedil in pharmaceutical formulations. The effects of buffer concentration, buffer pH, sodium dodecyl sulphate (SDS) concentration, organic modifier, applied voltage and injection time were investigated. Optimum results were obtained with a 50 mM borate buffer at pH 8.0 containing 50 mM SDS by using a fused silica capillary (50 m internal diameter, 72 cm effective length). The sample was injected hydrodynamically for 4 s at 50 mbar pressure and the applied voltage was +30 kV. The detection wavelength was set at 205 nm. Diflunisal was used as an internal standard. The analysis was performed at 25 °C and the total run time was 14 min. The method was suitably validated with respect to linearity range, limit of detection and quantification, precision, accuracy, specificity and robustness. The linear calibration range was 5–100 g mL^–1 and the limit of detection was determined as 1 g mL^–1. The method developed was successfully applied to the determination of piribedil in pharmaceutical formulations. The results were compared with a spectrophotometric method reported in the literature and no significant difference was found statistically.     

Determination of alkylphenol ethoxylates by micellar electrokinetic chromatography with bile salts

Octyl- and nonylphenol ethoxylates (OPEs and NPEs) with different numbers of ethoxy units (average values: n = 10 and N = 40 for OPEs, and n = 10 for NPEs) were separated by micellar electrokinetic chromatography under positive polarity using an 80 mM borate buffer of pH 8.5 containing sodium deoxycholate (SDC) or sodium cholate (SC). When sodium dodecyl sulfate (SDS) was added to the background electrolyte (BGE) in the absence of the bile salt, a single peak at a migration time longer than that of the EOF was obtained. Substituting the SDS by a bile salt, the homologues were resolved. At the same bile salt concentration, resolution between the homologues was higher with SDC than using SC. Optimum resolution between consecutive homologues was obtained with 50 mM SDC. In the presence of low or moderate amounts of acetonitrile or n-propanol, the background line improved significantly, whereas resolution may increase or decrease slightly. We propose a procedure for the determination of OPEs and NPEs with optimum resolution between the homologues as well as a modified procedure with improved selectivity for the single-run determination of other absorbing nonionic, cationic, and anionic (such as linear alkylbenzene sulfonates) surfactants in industrial and household cleaning products and its application to a variety of samples. The detection limit was ca. 28 microg x mL(-1) of total NPE (n = 10), and peak area repeatabilities at 50 microg x mL(-1) were 1.7% (intraday) and 5.6% (interday).     

Determination of pesticides in drinking water by micellar electrokinetic capillary chromatography

A new analytical procedure using a two-step sample preconcentration (solid-phase extraction (SPE) and field-amplified sample stacking) prior to separation by micellar electrokinetic capillary chromatography was developed for the determination of 14 pesticides such as aldicarb, carbofuran, isoproturon, chlorotoluron, metolachlor, mecoprop, dichlorprop, MCPA, 2,4-D, methoxychlor, TDE, DDT, dieldrin, and DDE in drinking water. Good recoveries of pesticides were obtained using SPE with sample pH adjusted to 2-3. Field-amplified sample stacking was found to give enrichment factors up to 30-fold preconcentration of various pesticides under reversed polarity at -2 kV for 50 s. The optimized background electrolyte (BGE) consisted of 50 mM sodium dodecyl sulfate (SDS), 10 mM borate buffer, 15 mM beta-cyclodextrin (beta-CD), and 22% acetonitrile at pH 9.6, running was under 25 kV and detection at 202 nm. Good linearity was obtained for all pesticides with detection limits down to 0.04-0.46 ng/mL and a working range of 0.1-40 ng/mL. The repeatabilities of migration time and peak area were satisfactory with relative standard deviations (RSDs) between 0.66 and 13.6% and 4.1 and 28%, respectively. All pesticides except dieldrin were found to be detected at concentrations at least tenfold lower than the World Health Organization (WHO) guideline values. The analytical procedure developed offers an economic method for fast screening of multiple pesticide residues in drinking water for health protection. It had been applied to determine carbofuran and MCPA in agricultural run-off water samples, giving satisfactory repeatabilities of 10 and 12%, respectively, with n=5 for the determination of pesticides in contaminated water samples.     

胶束电动毛细管色谱法测定植物中的水杨酸

建立了以胶束电动色谱为分离模式测定植物中水杨酸的新方法。在一定范围内，随着硼酸和甲醇浓度的升高，苯甲酸内标和水杨酸的分离度以近似线性关系升高；随缓冲液pH的升高分离度呈非线性升高；随十六烷基三甲基溴化铵浓度的升高分离度呈非线性下降。在优化的条件下，两可在12min内分离。测定了苹果和梨样品，并做了回收率试验，回收率在97.1%-102%之间。     

Profiling of cocaine by micellar electrokinetic chromatography

The potential of micellar electrokinetic chromatography (MEKC) for the profiling of cocaine samples is described. An MEKC system containing sodium dodecyl sulfate (SDS) and methanol was optimized using a test mixture of cocaine, its common impurities (benzoylecgonine, norcocaine, tropacocaine, and trans-cinnamoylcocaine), and several degradation products. The effect of pH, percentage modifier, and concentration surfactant on the separation has been investigated. The optimal separation buffer for cocaine samples consisted of 75 mM SDS, 17.5% methanol, and 25 mM borate (pH 8.3) and was well suited to separate components of diverse polarity in one run. Various cocaine seizures have been analyzed with the MEKC system and their signatures were compared. The electrokinetic chromatograms obtained were characteristic, and differences and similarities among the samples could easily be observed. Several impurities were identified in the samples by means of migration times and comparison of recorded and library UV spectra. The composition of the samples was determined semiquantitatively using relative corrected peak areas.     

胶束电动毛细管色谱法测定硫酸多粘菌素E药物中的多粘菌素E1和E2

采用胶束电动毛细管色谱法(MECC)对硫酸多粘菌素E药物中的主要成分多粘菌素E1和E2进行了分离,并测定了多粘菌素E1、E2的含量。分别考察了电泳电压、表面活性剂种类、Brij-35(月桂醇聚氧乙烯醚)浓度、乙腈含量、磷酸盐缓冲液的pH值、氯化钠浓度等实验参数对实验结果的影响,从而确定了最佳的分离条件: 电泳电压为10 kV,运行缓冲液为含有30 mmol/L Brij-35、5%(体积分数)乙腈、0.167 mol/L氯化钠的磷酸二氢钠缓冲液(0.01 mol/L,pH 4.1)。在优化的实验条件下,E1和E2得到了较好的分离,分离度达到1.94。以多粘菌素E1为例,柱效和峰面积的日间及日内测定的相对标准偏差(RSD)均小于5%。E1和E2在硫酸多粘菌素E药物中的含量分别为67%和32%。该方法简便、快速、准确、重现性好。     

Determination of ethinylestradiol and gestodene in oral contraceptives by micellar electrokinetic chromatography

Summary A micellar electrokinetic chromatographic method is presented which permits quantification of ethinylestradiol and gestodene in pharmaceutical products. Separation was carried out at 25°C and 25 KV, using a 20 mM borate buffer (pH 9.2), 15 mM sodium dodecylsulfate in 30% acetonitrile-water (v/v). Under these conditions analyses were carried out in 7 min. Four different oral contraceptives were analysed and the results compared favourably with those of a reference liquid chromatographic method.     

Pesticide analysis by micellar electrokinetic capillary chromatography

On-capillary sample concentration using sample stacking for the improvement of detection limits for various pesticides separated by micellar electrokinetic capillary chromatography was examined. The dependence of the stacking on different parameters was investigated. An approximately 30-fold preconcentration was achieved by applying sample stacking. Employing a two-step enrichment process (solid-phase extraction combined with sample stacking), detection limits were improved and the sample volume for SPE was reduced. In addition, the total time for the analysis was considerably reduced. Detection limits were between 0.01 and 0.1 ng/ml under these enrichment conditions.     

胶束在线扫集毛细管电泳法测定三聚氰胺

研究胶束在线扫集毛细管电泳法测定三聚氰胺的可行性,结果表明,与区带毛细管电泳相比,胶束在线扫集毛细管电泳法富集倍数提高约60倍。缓冲体系为140 mmol/L SDS+20 mmol/L NaH2PO4(pH 2.20)+10%(体积分数)甲醇,分离电压-18 kV,进样时间30 s,测量波长214 nm。考察了SDS浓度、pH、进样时间、运行电压等因素对分离测定的影响情况。在优化条件下,三聚氰胺在9 min时出峰,峰面积RSD≤3.7%。方法检出限、线性范围、相关系数分别为:0.13μg/mL、0.50~32.0μg/mL、0.9997。方法可用于奶粉中三聚氰胺的分离测定。     

Determination of bioactive diterpenoids from Andrographis paniculata by micellar electrokinetic chromatography.

The present paper describes the development of a micellar electrokinetic chromatographic (MEKC) method for simultaneous determination of andrographolide, deoxyandrographolide and neoandrographolide in ethanol extracts of Andrographis paniculata. Separations were carried out in a fused-silica capillary tube with UV detection at 214 nm. Good separation was achieved using a 20 mM borate buffer, containing 20 mM sodium dodecyl sulphate and 10 mM sodium cholate, adjusted to pH 8.3 at an operating voltage of 25 kV, temperature of 35°C and a hydrodynamic injection of 5 s. The method was validated with good correlation coefficients obtained (0.9986–0.9989) while relative standard deviation (RSD) of migration time was between 1.14 and 2.42. It is concluded that this method could be used for speedy and accurate qualitative and quantitative analysis of bioactive diterpenoids in andrographis herb and its derived products.     

Determination of amino acid ratios in natural products by micellar electrokinetic chromatography

Summary Separation of dansyl and di-dansyl derivatives of amino acids present in natural products (corn seed flour, wheat flour fraction gliadine) is described. Problems with coelution of derivatization by-products and reagent with some dansyl amino acids (DNS-AA) were solved. A preconcentration step after derivatization of real sample is described. DNS-AA peak areas for different varieties of corn seed flour were compared. Di-dansyl histidine is not separated from di-dansyl lysine and didansyl tyrosine. Additional separation mechanisms have to be introduced to achieve separation of these three species.     

Determination of antitubercular drugs by micellar electrokinetic capillary chromatography (MEKC)

A method for the determination of isoniazid (ISO), pyrazinamide (PYR) and rifampicin (RIF) in pharmaceutical products, by micellar electrokinetic capillary chromatography (MEKC) with ultraviolet detection is described. The influence of pH, concentration of surfactants, buffer and organic solvents, over the separation were studied as experimental variables. The optimal separation was carried out at 30 degrees C and 20 kV, using a 40 mM borate buffer and 100 mM sodium dodecylsulphate (SDS) adjusted to pH 8.5. Under these conditions, the analysis is accomplished in about 8 min. The method was applied to the determination of these compounds in different pharmaceuticals with good results when compared with a reference liquid chromatographic (LC) method.     

Determination of melatonin in commercial preparations by micellar electrokinetic chromatography and spectrofluorimetry

Melatonin was determined in pharmaceutical preparations by means of two simple and reliable analytical methods based on micellar electrokinetic chromatography (MEKC) and spectrofluorimetry. The fluorescence emission values were measured at λ=350 nm when exciting at λ=275 nm. The MEKC analysis was achieved using a system consisting of 40 mM SDS in phosphate buffer (20 mM, pH 7.5). The extraction of melatonin from the tablets was achieved by means of a simple one-step dissolution with methanol/water. Both methods were applied for the determination of melatonin in commercial formulations and galenic preparations. The MEKC procedure allows the quantitative determination of melatonin in all pharmaceutical preparations tested. On the contrary, the spectrofluorimetric method is not suitable for tablets which also contain tryptophan; this interference can be eliminated by a suitable liquid-liquid extraction procedure. The results obtained with the two methods are in good agreement and satisfactory in terms of precision and accuracy.