Sensitive analysis of donepezil in plasma by capillary electrophoresis combining on-column field-amplified sample stacking and its application in Alzheimer's disease

Field-amplified sample stacking (FASS) in capillary electrophoresis (CE) was used to determine the concentration of donepezil, an acetylcholinesterase inhibitor, in human plasma. A sample pretreatment by liquid-liquid extraction with isopropanol/n-hexane (v/v 3:97) and subsequent quantification by FASS-CE was used. Before sample loading, a water plug (0.5 psi, 6 s) was injected to permit FASS. Electrokinetic injection (7 kV, 90 s) was used to introduce sample cations. The separation condition for donepezil was performed in electrolyte solutions containing Tris buffer (60 mM, pH 4.0) with sodium octanesulfonate 40 mM and 0.01% polyvinyl alcohol as a dynamic coating to reduce analytes' interaction with capillary wall. The separation was performed at 28 kV and detected at 200 nm. Using atenolol as an internal standard, the linear ranges of the method for the determination of donepezil in human plasma were over a range of 1-50 ng/mL. The limit of detection was 0.1 ng/mL (S/N=3, sampling 90 s at 7 kV). One female volunteer (54 years old) was orally administered a single dose of 10 mg donepezil (Aricept, Eisai), and blood samples were drawn over a 60 h period for pharmacokinetic study. The method was also applied successfully to monitor donepezil in sixteen Alzheimer's disease patients' plasmas.     

Separation and determination of corynoxine and corynoxine B using chiral ionic liquid and hydroxypropyl‐β‐cyclodextrin as additives by field‐amplified sample stacking in capillary electrophoresis

A sensitive, fast, and effective method, field‐amplified sample stacking (FASS) in capillary electrophoresis, has been established for the separation and determination of corynoxine and corynoxine B. Hydroxypropyl‐β‐CD (HP‐β‐CD) and tetrabutylammonium‐L‐glutamic acid (TBA‐L‐Glu) were used as additives in the separation system. Electrokinetic injection was chosen to introduce sample from inlet at 10 kV for 50 s after a water plug (0.5 psi, 4 s) was injected to permit FASS. The running buffer (pH 6.1) was composed of 40 mM sodium dihydrogen phosphate solution, 130 mM HP‐β‐CD, and 10 mM TBA‐L‐Glu and the separation voltage was 20 kV. Under the optimum conditions, corynoxine and corynoxine B were successfully enriched and separated within 12 min and the sensitivity was improved approximately by 700–900 folds. Calibration curves were in a good linear relationship within the range of 62.5–5.00 × 10³ ng/mL for both corynoxine and corynoxine B. The limits of detection (S/N = 3) and quantitation (S/N = 10) were 14.9, 45.2 ng/mL for corynoxine and 11.2, 34.5 ng/mL for corynoxine B, respectively. Finally, this method was successfully applied for the determination of corynoxine and corynoxine B in the stems with hooks of Uncaria rhynchophylla and its formulations.     

Rapid and sensitive determination of strychnine and brucine in human urine by capillary electrophoresis with field‐amplified sample stacking

A simple, rapid, sensitive and low‐cost method using capillary electrophoresis (CE) coupled with field‐amplified sample stacking (FASS) has been developed and validated for the simultaneous determination of strychnine and brucine residues in human urine. Before sample loading, a water plug (3.5 kPa, 3 s) was injected to contain sample cations and to permit FASS. Electrokinetic injection at a voltage (20 kV, 25 s) was then used to introduce cations. Separation was performed using 20 mM acetate buffer (pH 3.8) with an applied voltage of 20 kV. The calibration curves were linear over a range of 8.00–2.56 ∞ 10² ng/mL (r = 0.9995) for strychnine and 10.0–3.20 × 10² ng/mL (r = 0.9999) for brucine. Extraction recoveries in urine were greater than 79.6 and 82.8% for strychnine and brucine, respectively, with an RSD of less than 4.9%. The detection limits (signal‐to‐noise ratio 3) for strychnine and brucine were 2.00 and 2.50 ng/mL, respectively. A urine sample from one healthy female volunteer (26 years old, 50 kg) was pretreated and analyzed. Strychnine and brucine levels in urine could be detected 24 h after administration. On these grounds, this method was feasible for application to preliminary screening of trace levels of abused drugs for both doping control and forensic analysis. Copyright © 2009 John Wiley & Sons, Ltd.     

Analysis of alendronate in human urine and plasma by magnetic solid-phase extraction and capillary electrophoresis with fluorescence detection

A simple, rapid and sensitive CE-fluorescence (FL) detection method for the analysis of alendronate (ALEN), a bisphosphonate drug, has been developed. Using a buffer solution of 20 mM sodium phosphate (pH 10.0) and a voltage of 24 kV, separation of ALEN in a 55-cm length (35-cm effective length) capillary was achieved in 5 min. FL detection of ALEN was performed via pre-column derivatization with 2,3-naphthalene dicarbox-yaldehyde (NDA). Linear correlation (r=0.9981, n=6) between FL intensity and analyte concentration was obtained in the range of 7-200 ng/mL ALEN. The developed CE-FL method was applied to the analysis of ALEN in human urine and plasma samples. In order to eliminate the interfering matrix components, SPE using magnetic Fe(3) O(4) @Al(2) O(3) nanoparticles as solid sorbents was employed to clean the biological fluids before CE-FL analysis. The linear ranges of ALEN in urine and plasma were 5-100 ng/mL (r = 0.9982, n = 7) and 5-70 ng/mL (r = 0.9954, n = 7), respectively. The LOD and LOQ in both urine and plasma samples were 1.5 and 5 ng/mL ALEN, respectively. Total analysis time including sample pre-treatment and CE separation was less than 1.5 h.     

Multiresidue determination of sulfonamides in chicken meat by polymer monolith microextraction and capillary zone electrophoresis with field-amplified sample stacking

A method based on poly(methacrylic acid-co-ethylene glycol dimethacrylate) (MAA-EGDMA) monolith microextraction (PMME) and online preconcentration technique of field-amplified sample stacking (FASS) was proposed for sensitive capillary electrophoresis-ultraviolet (CE-UV) analysis of 12 sulfonamides (sulfamethazine, sulfamethoxypyridazine, sulfathiazole, sulfamerazine, sulfameter, sulfadoxine, sulfadimethoxine, sulfamonomethoxine sodium, sulfachlorpyridazine, sulfamethoxazole, sulfamethizole, and sulfisoxazole) in chicken samples. The conditions of PMME were optimized for the improvement of extraction efficiency and reduction of the matrix interferences from chicken sample. The best separation was achieved within 15min using a buffer of 100mM phosphate electrolyte (pH 7.3) with temperature and voltage of 20 degrees C and 25kV, respectively. By applying FASS, detection limits of 3.49-16.7ng/g were achieved with satisfactory precision (RSD<==13%) and recovery (96.3-104%) over a linear range of 50-1000ng/g for most analytes.     

An online field-amplification sample stacking method for the determination of diuretics in urine by capillary electrophoresis-amperometric detection

A simple and sensitive online field-amplification sample stacking (FASS) pre-enrichment method following by capillary electrophoresis with amperometric detection has been developed for the determination of diuretics, such as indapamide (IDP), hydrochlorothiazide (HCT) and bumetanide (BMTN) in urine. Under the optimum conditions, it was found that the low concentration buffer solution could be used as the diluents for simultaneous field-amplification injection of three diuretics after electrokinetically injecting a short water plug (15 kV, 3 s). Three analytes could be well separated within 10 min in an uncoated fused-silica capillary with H(3)BO(3)-Na(2)B(4)O(7) (BB) buffer solution (pH 8.98). The detection limits (S/N=3) were 9.0 ng/mL for IDP, 20 ng/mL for HCT and 1.5 ng/mL for BMTN, respectively. The detection limits of three diuretics were much lower by FASS than that by conventional sample injection, of which the detection limits were 340, 890 and 330 ng/mL for IDP, HCT and BMTN, respectively. Especially, for bumetanide the detection limit was 220-time lower by FASS. The linear ranges of three diuretics were all over three orders of magnitude. The proposed method has been successfully applied to analyze the diuretics in human urine samples without off-column sample pre-concentration.     

On-line stacking techniques for the nonaqueous capillary electrophoretic determination of acrylamide in processed food

In the present study, field amplified sample stacking (FASS) techniques in the nonaqueous capillary electrophoresis method (NACE) were introduced for the on-line concentration of the acrylamide to improve acrylamide detection at 210 nm by diode-array detection. Acetonitrile (ACN) as a nonaqueous solvent permits acrylamide to be protonated through the change of its acid-base chemistry, allowing capillary electrophoretic separation of this compound. Choosing 30 mmol L(-1) HClO(4), 20 mmol L(-1) NaClO(4), 218 mmol L(-1) CH(3)COOH in ACN as the separation electrolyte and employing sample stacking methods, the LOD value of acrylamide was decreased to 2.6 ng mL(-1) with electrokinetic injection and 4.4 ng mL(-1) with hydrodynamic injection. Optimized stacking conditions were applied to the determination of acrylamide in several foodstuffs. The method is simple, rapid, inexpensive, and widely applicable for the determination of acrylamide in food samples.     

On-line sample enrichment for the determination of proteins by capillary zone electrophoresis with poly(vinyl alcohol)-coated bubble cell capillaries

Field-amplified sample stacking (FASS) is used to separate basic proteins in a poly-(vinyl alcohol)-coated bubble cell capillary. To our knowledge, this is the first paper describing the on-column stacking of proteins (as cations) using FASS in bubble cell capillary. The bubble cell capillary is fabricated using a one-step method. Cetyltrimethylammonium chloride is added into the running buffer to reverse the EOF and, thus, to pump the water plug out during the sample stacking step. The effect of the water plug lengths and sample injection durations were investigated and optimized. The results obtained were compared with those for the normal capillary without bubble cell in terms of resolution and sensitivity enhancement. Under the optimal condition, this method can improve the sensitivity of the peak areas ranging from 5000- to 26 000-fold. The RSDs (n = 5) of the migration time and peak area are satisfactory (less than 0.6 and 12%, respectively). Application of the capillary electrophoresis method with bubble cell, FASS, and UV detection thereby leads to the determination of these proteins at concentrations ranging from 3 to 10 ng/mL, based on a signal-to-noise ratio of 3:1.     

毛细管电泳场放大进样-高压非接触电导检测测定痕量Zn~(2+)

利用高压电容耦合非接触电导检测器(HV-C4D),结合毛细管电泳场放大进样(FASS),以2-N-吗啡啉乙磺酸(MES)/组氨酸(His)为缓冲溶液,电泳分离测定了Zn2+.考察了样品溶液中MES/His的浓度及电动进样时间对场放大浓缩因子及缓冲溶液浓度对检测灵敏度的影响.在10mmol/LMES/His(pH=4.9)的分离缓冲溶液中,FASS对Zn2+的浓缩因子为1.3×103.Zn2+的浓度在10～1000nmol/L范围内与峰面积有良好线性关系(R=0.9995),检测限为5nmol/L(S/N=3).该方法可用于痕量Zn2+的测定.     

Trace analysis of zotepine and its active metabolite in plasma by capillary electrophoresis with solid phase extraction and head-column field-amplified sample stacking

A sensitive high-performance capillary zone electrophoresis (CZE) with head-column field-amplified sample stacking (FASS) in binary system has been developed for the simultaneous determination of zotepine and its active metabolite, norzotepine, in human plasma. The separation of zotepine and norzotepine was performed using a background electrolyte consisting of 50% ethylene glycol-borate buffer (20mM, pH 8.0) solution with 20% methanol as the running buffer and on-column detection at 200 nm. Under the optimal FASS-CZE condition, good separation with high efficiency and short analysis time is achieved. Several parameters affecting the separation and sensitivity of the drug were studied, including sample matrix, pH and concentrations of the borate buffer, ethylene glycol and methanol. Using clozapine as an internal standard, the linear ranges of the method for the determination of zotepine and norzotepine in human plasma were over 3-100 ng/mL; the detection limits of zotepine and norzotepine in plasma were 2 and 1 ng/mL, respectively. A sample pretreatment by means of solid-phase extraction (SPE) with subsequent quantitation by FASS-CZE was used. The application of the proposed method for determination of zotepine and norzotepine in plasma collected after oral administration of 125 mg zotepine in one schizophrenic patient was demonstrated.     

Steroselective determination of trihexyphenidyl using carboxylmethyl-β-cyclodextrin by capillary electrophoresis with field-amplified sample stacking

A simple, sensitive and low-cost method using capillary electrophoresis coupled with field-amplified sample stacking (FASS) technique has been developed for enantioselective separation and quantification of trihexyphenidyl (THP) enantiomers in human serum. In this work, three kinds of modified β-cyclodextrin were tested as chiral selectors in CE. Among the CDs studied, THP enantiomers could only be separated by carboxylmethyl-β-cyclodextrin (CM-β-CD). A systematic study of the parameters (CD concentration and pH value in CE buffer, separation voltage and temperature, composition of sample solvent, injection voltage and time) affecting chiral separation and on-line concentration of THP enantiomers were investigated and optimized. The optimum FASS method provided a sensitivity enhancement of about 490-fold compared with usual hydrodynamic injection. Limits of detection for each enantiomer were in the low ng ml^− 1 concentration range (0.92 ng ml^− 1 or 3.06 nM). The quantification of each THP enantiomer in human serum was performed after serum sample extraction. To validate this CE-FASS method, linear regression analysis, intra and inter-day precision and recovery were determined with satisfying results.     

Nonaqueous capillary electrophoresis for rapid and sensitive determination of fangchinoline and tetrandrine in radix Stephaniae tetrandrae and its medicinal preparations

A simple, rapid, and sensitive non-aqueous capillary electrophoresis procedure with head-column field-amplified sample stacking concentration for the analysis of fangchinoline and tetrandrine is established. Optimum separation and stacking conditions were obtained when the sample was injected at 8 kV for 50 s after preliminary pressure injection of ethanol (16.9 kPa) for 0.6 s and separated with the buffer containing 50 mM ammonium acetate, 0.5% (v/v) acetic acid, and 50% (v/v) acetonitrile in methanol medium at 24 kV applied voltage. The analytes were detected by UV at 214 nm. The two bisbenzylisoquinoline alkaloids can be separated within 6 min and quantified with high sensitivity. The detection limits were 0.30 ng mL(-1) for fangchinoline and 0.34 ng mL(-1) for tetrandrine, which indicated that the sensitivities were at least 1000-fold enhanced over those reported in the literature as obtained by UV detection. The method was applied to the analysis of fangchinoline and tetrandrine in Radix Stephaniae tetrandrae and its medicinal preparations with good results.     

毛细管电泳电致化学发光检测血浆中布比卡因

布比卡因是一种外科局部麻醉剂,使用过量会导致中枢神经系统和心脏血管系统中毒^[1],可引起心脏停博.高效液相色谱和毛细管电泳(CE)^[2]是该药常用的检测方法.     

Glucose‐β‐CD interaction assisted ACN field‐amplified sample stacking in CZE for determination of trace amlodipine in beagle dog plasma

A simple, sensitive and low‐cost method using CE coupled with glucose‐β‐CD interaction assisted ACN stacking technique has been developed for quantification of trace amlodipine in dog plasma. The plasma samples were extracted with methyl tert‐butyl ether. The separation was performed at 25°C in a 31.2 cm × 75 μm fused‐silica capillary with an applied voltage of 15 kV. The BGE was composed of 6.25 mM borate/25 mM phosphate (pH 2.5) and 5 mg/mL glucose‐β‐CD. The detection wavelength was 200 nm. Because CD could diminish the interaction between drugs and matrix, and derivation groups of CD play an important role in separation performance, the effects of β‐CD, and its derivatives on the separation were studied at several concentrations (0, 2.5, 5.0, 10.0 mg/mL). In this study, organic solvent field‐amplified sample stacking technique in combination with glucose‐β‐CD enhanced the sensitivity about 60–70 folds and glucose‐β‐CD could effectively improve the peak shape. All the validation data, such as accuracy, precision extraction recovery, and stability, were within the required limits. The calibration curve was linear for amlodipine from 1 to 200 ng/mL. The method developed was successfully applied to the pharmacokinetic studies of amlodipine besylate in beagle dogs.     

肌肽类生物活性肽的毛细管电泳在线富集技术

建立了毛细管区带电泳(CZE)在线富集3种肌肽类活性肽(肌肽、鹅肌肽和高肌肽)的两种简便有效的方法。一种是大体积进样反向压力排除基体富集（LVSRP）技术,即通过流体动力学进样,在不改变电源极性的条件下,利用反向压力排除样品基体,电堆积富集后进行CZE分离;另一种是大体积进样电渗流排除基体富集（LVSEP）技术,即通过流体动力学进样,于运行缓冲液中加入溴化十六烷基三甲基铵(CTAB)动态修饰毛细管表面,通过电渗流排除样品基体,改变电源极性后进行CZE分离。与常规CZE相比,LVSRP技术和LVSEP技术使检测灵敏度提高了40~60倍。对影响两种富集过程的一些因素进行了研究,在最优富集条件下考察本方法的线性范围为0.080~5.0 μmol/L。对3种生物活性肽的检测限(S/N=3)分别为LVSRP 41~58 nmol/L,LVSEP 35~43 nmol/L。     

Cation-selective exhaustive injection and sweeping micellar electrokinetic chromatography for analysis of morphine and its four metabolites in human urine

A cation-selective exhaustive injection and sweeping micellar EKC (CSEI-Sweep-MEKC) was established to analyze morphine and its four metabolites, including codeine, normorphine (NM), morphine-3-glucuronide (M3G), and morphine-6-glucuronide (M6G). After SPE, the urine samples were analyzed by this CE method. The phosphate buffer (75 mM, pH 2.5) containing 30% methanol was first filled into an uncoated fused-silica capillary (40 cm, 50 microm id), then a high-conductivity buffer (120 mM phosphate, 10.3 kPa for 99.9 s) followed. The pretreated urine sample was loaded by electrokinetic injection (10 kV, 600 s). The stacking and separation were performed by using phosphate buffer (25 mM, pH 2.5) containing 22% methanol and 100 mM SDS at -20 kV, and detected at 200 nm. During method validation, calibration plots were linear (r > or = 0.998) over a range of 30-3000 ng/mL for morphine, NM, and codeine, 100-2000 ng/mL for M6G, and 80-3200 ng/mL for M3G. The LODs (S/N = 5, sampling 600 s at 10 kV) were 10 ng/mL for morphine, NM, and codeine, 35 ng/mL for M6G, and 25 ng/mL for M3G. This stacking CE method could increase 2500-fold sensitivity of codeine, when comparing with CZE. Five addicts' urine specimens were analyzed. Their results were compared with those of LC-MS-MS, and showed good coincidence. This method could be feasible for monitoring morphine and its metabolites in forensic interest and pharmacokinetic investigations.     

Direct and sensitive analysis of methamphetamine, ketamine, morphine and codeine in human urine by cation-selective exhaustive injection and sweeping micellar electrokinetic chromatography

Cation-selective exhaustive injection and sweeping micellar electrokinetic chromatography (CSEI-Sweep-MEKC) was directly used to test some abuse drugs in human urine, including morphine (M), codeine (C), ketamine (K) and methamphetamine (MA). First, phosphate buffer (50 mM, pH 2.5) containing 30% methanol was filled into uncoated fused silica capillary (40 cm, 50 microm I.D.), then high conductivity buffer (100 mM phosphate, 6.9 kPa for 99.9 s) was followed. Electrokinetic injection (10 kV, 500 s) was used to load samples and to enhance sensitivity. The stacking step and separation were performed at -20 kV and 200 nm using phosphate buffer (25 mM, pH 2.5) containing 20% methanol and 100 mM sodium dodecyl sulfate. Using CSEI-Sweep-MEKC, the analytes could be simultaneously analyzed and have a detection limit down to ppb level. It was unnecessary to have sample pretreatments. During method validation, calibration plots were linear (r>or=0.9982) over a range of 150-3,000 ng/mL for M and C, 250-5,000 n g/mL for MA, and 50-1,000 ng/mL for K. The limits of detection were 15 ng/mL for M and C, and 5 ng/mL for MA and K (S/N=3, sampling 500 s at 10 kV). Comparing with capillary zone electrophoresis, the results indicated that this stacking method could increase 6,000-fold sensitivity for analysis of MA. Our method was applied for analysis of 28 real urine samples. The results showed good coincidence with immunoassay and GC-MS. This method was feasible for application to detect trace levels of abused drugs in forensic analysis.     

Field-amplified sample stacking capillary electrophoresis with electrochemiluminescence applied to the determination of illicit drugs on banknotes

Capillary electrophoresis (CE) with Ru(bpy)3(2+) electrochemiluminescence (ECL) detection system was established to the determination of contamination of banknotes with controlled drugs and a high efficiency on-column field-amplified sample stacking (FASS) technique was also optimized to increase the ECL intensity. The method was illustrated using heroin and cocaine, which are two typical and popular illicit drugs. Highest sample stacking was obtained when 0.01 mM acetic acid was chosen for sample dissolution with electrokinetical injection for 6 s at 17 kV. Under the optimized conditions: ECL detection at 1.2 V, separation voltage 10.0 kV, 20 mM phosphate-acetate (pH 7.2) as running buffer, 5 mM Ru(bpy)3(2+) with 50 mM phosphate-acetate (pH 7.2) in the detection cell, the standard curves were linear in the range of 7.50x10(-8) to 1.00x10(-5) M for heroin and 2.50x10(-7) to 1.00x10(-4) M for cocaine and detection limits of 50 nM for heroin and 60 nM for cocaine were achieved (S/N = 3), respectively. Relative standard derivations of the ECL intensity and the migration time were 3.50 and 0.51% for heroin and 4.44 and 0.12% for cocaine, respectively. The developed method was successfully applied to the determination of heroin and cocaine on illicit drug contaminated banknotes without any damage of the paper currency. A baseline resolution for heroin and cocaine was achieved within 6 min.     

Cation-selective exhaustive injection and sweeping MEKC for direct analysis of methamphetamine and its metabolites in urine

Direct analysis of methamphetamine, amphetamine, and p-hydroxymethamphetamine in urine was achieved by cation-selective exhaustive injection and sweeping micellar EKC. A bare fused-silica capillary (40 cm, 50 microm id) was filled with phosphate buffer (80 mM, pH 3, containing 20% ACN). Then a high-conductivity buffer (100 mM phosphate, pH 3; 6.9 kPa for 2.5 min) was injected. Samples were loaded using electrokinetic injection (10 kV, 600 s) which created long zones of cationic analytes. To enhance sensitivity by sweeping, the stacking step was performed using a phosphate buffer (50 mM, pH 3, containing 20% ACN and 100 mM SDS) at -20 kV before separation by MEKC. This method was capable of detecting the analytes at ppb levels. The calibration plots were linear (r(2) >or= 0.9948) over a range of 100-5000 ng/mL for methamphetamine, and 100-2000 ng/mL for amphetamine and p-hydroxymethamphetamine. The LODs (S/N = 3) were 20 ng/mL for methamphetamine, and 15 ng/mL for amphetamine and p-hydroxymethamphetamine. The method was applied to analysis of 14 urine samples of addicts and is suitable for screening suspected samples for forensic purposes. The results showed good agreement with fluorescence polarization immunoassay and GC-MS.     

Capillary electrochromatographic analysis of barbiturates in serum

A capillary electrochromatographic method was developed for the separation of barbiturates. The separation was optimized in a 75 microm ID capillary, packed with 3-(1,8-naphthalimido)propyl-modified silyl silica gel (NAIP), studying the effect of buffer pH, buffer concentration, and mobile phase composition. Using an applied voltage of 20 kV and the short-end injection method (9 cm capillary effective length), the mobile phase of 1.0 mM citrate buffer (pH 5.0) containing 40% methanol provided the baseline separation of barbital, phenobarbital, secobarbital, and thiopental (internal standard) in less than 4.5 min. The method was successfully applied to the analysis of barbiturates in human serum. Under the optimal conditions, good repeatability and linearity were obtained in the range of 2.90-43.29 microg/mL for barbital, phenobarbital, and secobarbital.