高效液相色谱-串联质谱法同时检测血液中3种季铵盐类肌松剂

维库溴铵、罗库溴铵和泮库溴铵是一类广泛使用的非去极化肌松剂,使用过程中引起过敏反应甚至死亡的情况时有发生,快速检测血液中该类肌松剂的浓度,可为临床早期诊断提供有价值的信息.该类肌松剂为强极性的季铵盐类化合物,在反相色谱柱上难以保留,主要采用离子对色谱法进行分离.采用离子对色谱法时,加入的离子对试剂有离子抑制作用,可降低...     

Determination of carbethopendecinium bromide in eye drops by capillary electrophoresis with capacitively coupled contactless conductivity detection

Antiseptic agent carbethopendecinium bromide (septonex) was determined by capillary electrophoresis with capacitively coupled contactless conductivity detection. Optimal separation of this quaternary ammonium ion was achieved in BGE of pH 7.0 containing 30 mM 2-(N-morpholino)ethanesulfonic acid, 12.5 mg/mL of 2-hydroxypropyl-β-cyclodextrin and 20% v/v of acetonitrile. The separation was performed at 25°C in an uncoated fused silica capillary (50 μm id; total length, 60.5 cm; effective length, 50 cm) at 30 kV. Samples were injected hydrodynamically at 50 mbar for 6 s. For quantitative analysis, L-arginine (500 μg/mL) was used as internal standard. The calibration curve was rectilinear for 25-400 μg/mL of septonex (y=0.0113x-0.0063; r(2)=0.9992). The LOD was 7 μg/mL of septonex (at S/N=3). The run-to-run repeatability (n=6) was characterized by the RSDs of 0.18% for the migration time and 1.96% for the analyte/internal standard peak area ratio. Accuracy tested by recovery experiments at three concentration levels gave recoveries of 100.27-104.22% with RSD ≤2.19%. The method was successfully applied to the assay of carbethopendecinium bromide in eye drops. Quaternary ammonium ions having structure and size close to that of carbethopendecinium may not be resolved from the analyte.     

Large volume sample stacking with EOF and sweeping in CE for determination of common preservatives in cosmetic products by chemometric experimental design

This study proposes a capillary electrophoresis method incorporating large volume sample stacking, EOF and sweeping for detection of common preservatives used in cosmetic products. The method was developed using chemometric experimental design (fractional factorial design and central composite design) to determine multiple separation variables by efficient steps. The samples were loaded by hydrodynamic injection (10 psi, 90 s), and separated by phosphate buffer (50 mM, pH 3) containing 30% methanol and 80 mM SDS at -20 kV. During method validation, calibration curves were found to be linear over a range of 5-100 μg/mL for butyl paraben and isobutyl paraben; 0.05-10 μg/mL for ethyl paraben; 0.2-50 μg/mL for dehydroacetic acid; 0.5-70 μg/mL for methyl paraben; 5-350 μg/mL for sorbic acid; 0.02-450 μg/mL for p-hydroxybenzoic acid and 0.05-10 μg/mL for salicylic acid and benzoic acid. The analytes were analysed simultaneously and their detection limits (S/N = 3) were down to 0.005-2 μg/mL. The analysis method was successfully used for detection of preservatives used in commercial cosmetics.     

Analysis of steroids. XLI. Ion-pair high-performance liquid chromatographic separation of quaternary ammonium steroids on silica.

A normal-phase ion-pair chromatographic system has been developed for the high-performance liquid chromatographic investigation of pipecuronium bromide (2 beta,16 beta-bis-(N'-dimethyl-l-piperazinyl)-3 alpha,17 beta-diacetoxy-5 alpha-androstane dibromide) and related quaternary ammonium steroids. The use of silica as the stationary phase and a 96:4 mixture of acetonitrile and water containing 0.1 mol/dm3 sodium perchlorate as the eluent with detection at 213 nm enable the potential impurities as well as the hydrolytic and oxidative degradation products of pipecuronium bromide to be separated and detected down to the 0.01% level. The above system is also applicable to the high-performance liquid chromatographic investigation of other quaternary ammonium steroids (pancuronium bromide, vecuronium bromide).     

Validation of a high-performance liquid chromatography method for the determination of pancuronium in Pavulon injections

A new high-performance liquid chromatography (HPLC) method was developed for the quality control of pancuronium bromide and its degradation products. The HPLC method used a 5-microm Supelcogel ODP-50 (150x4 cm) column with acetonitrile-CH3OH-water-F3CCOOH (20.5:74.9:0.1, v/v) as the mobile phase (pH value 2.0 adjusted with trifluoroacetic acid) at a flow-rate 0.8 ml/min and UV detection at 210 nm. The Beer's law plots were found to be linear over the concentration range 0.4-1.2 mg/ml of pancuronium bromide and 0.04-0.08 mg/ml of desacetyl degradation products (R2=0.9995). The RSD of the peak areas was 1.09% and the recovery was 102.43%. The RSD value shows good precision, acceptable accuracy and reproducibility of the new method for the determination of pancuronium bromide in presence of its desacetyl degradation products. The method is rapid and sensitive enough to be used for Pavulon injection analysis.     

Analysis of therapeutic peptides in human urine by combination of capillary zone electrophoresis-electrospray mass spectrometry with preparative capillary isotachophoresis sample pretreatment

The presented study deals with the off-line coupling of preparative isotachophoresis (pITP) with on-line combination of capillary zone electrophoresis with electrospray mass spectrometric detection (CZE-ESI-MS) used for the analysis of therapeutic peptides (anserine, carnosine, and buserelin) in complex matrix (urine). Preparative capillary isotachophoresis, operating in a discontinuous fractionation mode in column-coupling configuration, served as a sample pretreatment technique to separation, and fractionation of mixture of therapeutic peptides present in urine at low concentration level. The fractions isolated by pITP procedure were subsequently analyzed by capillary zone electrophoresis with electrospray mass spectrometric detection. Acetic acid at 200 mmol L(-1) concentration served as background electrolyte in CZE stage and it is compatible with MS detection in positive ionization mode. In pITP fractionation procedure, sodium cation (10 mmol L(-1) concentration) as leading ion and beta-alanine as terminating ion (20 mmol L(-1) concentration) were used. While using CZE-ESI-MS, the limits of detection were 0.18 μg mL(-1) for carnosine, 0.17 μg mL(-1) for anserine and 0.64 μg mL(-1) for buserelin in water and 0.19 μg mL(-1) for carnosine, 0.50 μg mL(-1) for anserine and 0.74 μg mL(-1) for buserelin in 10 times diluted urine, respectively. The cleaning power of pITP sample pretreatment was proved as the peptides provided the higher MS signals at lower concentration levels resulting from the minimized matrix effects. The quality of obtained MS/MS spectra was very good so that they can provide information about the structure of analytes, and they were used for verification of the analytes identities. The pITP pretreatment improved the detection limits of the analyzed therapeutic peptides at least 25 times compared to the CZE-ESI-MS itself.     

超高效液相色谱法测定虾青素

建立超高效液相色谱法快速检测虾青素的方法。采用UPLC BEH C_8色谱柱(50 mm×2.1 mm,1.7μm),考察了流动相、流量及柱温对虾青素样品分离的影响,确定了最佳色谱条件:等度洗脱,流动相为甲醇–水(体积比为75∶25),流量为0.5 mL/min,柱温为40℃,检测波长为475 nm。虾青素的质量浓度在0.2~10.0μg/mL范围内与其色谱峰面积呈良好的线性关系,线性相关系数r=0.998 8,检出限(S/N=3)为0.1μg/mL,定量限(S/N=10)为0.2μg/mL。测定结果的相对标准偏差为0.41%(n=6),加标回收率为105.8%~110.3%。该方法快速、简单、可靠、灵敏、重复性好,可用于虾青素有关样品的快速检测。     

Simultaneous analysis of biologically active pyridines in pharmaceutical formulations by capillary zone electrophoresis

A capillary zone electrophoretic method using UV detection is developed for the analysis of four biological active pyridines [i.e., nicotine (NIC), cotinine (COT), nicotinic acid (NA), and nicotinamide (NM)]. The separation of the pyridines is achieved in 25 mM sodium dihydrogen phosphate (pH 2.1) using a fused-silica capillary with an effective length of 56 cm and an inner diameter of 50 μm (extended light path), hydrodynamic injection at 50 mbar for 10 s, a temperature of 25°C, applied voltage of 30 kV, and UV detection at 260 nm. These conditions provide baseline separation of all the analytes [resolution (R(s)) > 3.6] in 9.4 min with good linearity (r(2) > 0.998, in ranges of 50-600 μg/mL for NIC, 8-160 μg/mL for NM, and 10-200 μg/mL for COT and NA), precision (relative standard deviation <2.04%), recovery (96.4-101.6%), limits of detection (<3.0 μg/mL), and quantitation (<10 μg/mL). The method is robust upon the alterations of pH of BGE, separating voltage, and injection time [the RSDs of the relative migration time (migration time of the analyte/migration time of the internal standard) and resolution <3.26%]. The method is efficient, reliable, and simple for the routine analysis of NIC, NA, and NM in various products such as gum and tablets and can be applied to determine COT in thermal degradation of NIC gum.     

Mass Spectrometric and Contactless Conductivity Detection Approaches in the Determination of Muscle Relaxants by Capillary Electrophoresis

Novel and rapid capillary electrophoresis-coupled tandem mass spectrometry (CE-MS/MS) and capillary electrophoresis with capacitively coupled contactless conductivity detection (CE-C⁴D) methods have been developed for the separation and determination of three neuromuscular blocking agents: pancuronium, vecuronium, and rocuronium. In both cases, the separation was conducted in background electrolytes based on acidic acetate-ammonium buffers to avoid possible decomposition of the analytes that are known to be unstable in alkaline media. Baseline resolution of the analytes was achieved in the presence of modified γ-cyclodextrin by CE with C⁴D detection. The two detection techniques were compared with regard to analytical figures of merit including linear dynamic range, limit of detection, limit of quantification, precision, and accuracy. The calibration curves showed good linearity for both detection methods examined (characterized by r² ≥ 0.9908). The LODs of the CE-MS/MS and the CE-C⁴D methods differed at least by two orders of magnitude considering all analytes. The differences in precision and accuracy of these methods were evaluated and discussed. The assays of pancuronium, vecuronium, and rocuronium in commercial injection solutions by CE-MS/MS and CE-C⁴D were performed and the results compared.     

毛细管电泳-电致化学发光法测定兔血浆中的羟考酮

基于稀土Eu(Ⅲ)掺杂的类普鲁士蓝膜修饰的铂电极为工作电极,建立了测定羟考酮的毛细管电泳-电致化学发光分析方法。考察了检测电位、运行缓冲溶液的酸度及浓度、分离电压、进样条件等对电泳分离效果及检测灵敏度的影响。在最佳的实验条件下,羟考酮可在4 min内得到分离,其ECL强度值与羟考酮的质量浓度在7.0×10-2～7.0μg/mL和7.0～70.0μg/mL范围内呈良好的线性关系,检出限为4.2×10-2μg/mL(3σ),峰高和迁移时间的相对偏差分别为3.6%和0.48%(n=6)。方法用于兔血浆中羟考酮含量的检测,加标回收率在99.7%～101.0%之间。     

Development and validation of a novel RP-HPLC method for the analysis of reduced glutathione

The objective of this study was the development, optimization, and validation of a novel reverse-phase high-pressure liquid chromatography (RP-HPLC) method for the quantification of reduced glutathione in pharmaceutical formulations utilizing simple UV detection. The separation utilized a C18 column at room temperature and UV absorption was measured at 215 nm. The mobile phase was an isocratic flow of a 50/50 (v/v) mixture of water (pH 7.0) and acetonitrile flowing at 1.0 mL/min. Validation of the method assessed the methods ability in seven categories: linearity, range, limit of detection, limit of quantification, accuracy, precision, and selectivity. Analysis of the system suitability showed acceptable levels of suitability in all categories. Likewise, the method displayed an acceptable degree of linearity (r(2) = 0.9994) over a concentration range of 2.5-60 μg/mL. The detection limit and quantification limit were 0.6 and 1.8 μg/mL respectively. The percent recovery of the method was 98.80-100.79%. Following validation the method was employed in the determination of glutathione in pharmaceutical formulations in the form of a conjugate and a nanoparticle. The proposed method offers a simple, accurate, and inexpensive way to quantify reduced glutathione.     

微芯片毛细管电泳快速测定加替沙星注射液中加替沙星的含量

研究了用微芯片毛细管电泳非接触电导检测系统快速测定加替沙星注射液中加替沙星的方法。对缓冲液的类型、浓度、分离电压以及进样时间等因素进行了优化。最佳条件为:缓冲液5.0 mmol/L HAc,分离电压2.0 kV,进样时间15.0 s。在该条件下,可在1.0 min内实现加替沙星的快速含量测定。线性范围为4.0～150μg/mL,检出限为1.0μg/mL,加标回收率为95.7%～101%,可成功测定注射液中加替沙星的含量。     

Determination of thiol drugs in pharmaceutical formulations as their S-pyridinium derivatives by high-performance liquid chromatography with ultraviolet detection

Acetylcysteine and captopril can be determined in pharmaceutical formulations after precolumn derivatization of the thiol with 1-benzyl-2-chloropyridinium bromide (BCPB) by reversed-phase ion-pair HPLC separation and UV detection. The thiol group of the drugs reacts with BCPB in 0.1 mol/L phosphate buffer (pH 8) to form S-pyridinium derivatives showing an absorption maximum at 314 nm. The S-pyridinium derivative was separated isocratically on an Asahipak ODP-50 column at 45°C with 0.2 mol/L citrate buffer containing 10 mmol/L of sodium octanesulfonate (pH 2.5) and acetone (4:1, v/v). Calibration curves for both analytes were linear over the range of 2–20 μg/mL with variation coefficients of 3.12–1.28% for acetylcysteine and 9.22–1.51% for captopril.     

Determination of Doxepin in human plasma using cloud‐point extraction with high‐performance liquid chromatography and ultraviolet detection

A new straightforward method based on cloud‐point extraction has been developed, optimized, and validated for the determination of doxepin in human plasma by high‐performance liquid chromatography separation and UV detection. The nonionic surfactant Triton X‐114 was chosen as the extraction solvent. Chromatography separation was performed on a μBondapak^R C₁₈ column (4.6 mm id × 300 mm, 3 μm particle size), which was used for isocratic elution at a detection wavelength of 289 nm. Under the optimum conditions, the linear range of doxepin in human plasma was 0.1–0.9 μg/mL. Also, the detection limit, preconcentration factor, and enrichment factor were 0.08 μg/mL, 50, and 49.0, respectively.     

在线扫集-胶束毛细管电动色谱法测定复方氨酚烷胺胶囊中的3种有效成分

以在线扫集-胶束毛细管电动色谱法(Sweeping-MEKC)测定了复方氨酚烷胺胶囊中的马来酸氯苯那敏、咖啡因和对乙酰氨基酚3种有效成分。考察了缓冲溶液pH值、SDS浓度、分离电压及进样时间等对分离效果的影响。优化条件:以未涂层熔融石英毛细管(55 cm×50μm,有效柱长35 cm)为分离柱;环境温度25℃;80 mmol/L十二烷基磺酸钠+20 mmol/L NaH2PO4(pH 2.2)+15%乙腈为缓冲体系,分离电压-20kV,进样时间60 s(H=20.0 cm),测量波长210 nm。在该条件下氯苯那敏、咖啡因和对乙酰氨基酚在25min内出峰,峰面积RSD均小于4%;线性范围分别为2.45～39.17、1.61～25.76、1.58～25.28 mg/L;检出限(S/N=3)分别达139、34、24μg/L,回收率分别为96%～101%、98%～102%、96%～102%。     

Quantification of 4-hydroxy-2,5-dimethyl-3-furanone in fruit samples using solid phase microextraction coupled with gas chromatography-mass spectrometry

Furaneol is an important aroma compound. It is very difficult to extract furaneol from food matrices and separate it on a gas chromatography column due to its high polarity and instability. A new quantitative method was developed to quantify furaneol in aqueous samples by the use of derivatization/solid phase microextraction (SPME) coupled with gas chromatography/mass spectrometry (GC/MS). The derivatization was carried out by reacting pentafluorobenzyl bromide with furaneol in basic solutions at elevated temperatures. The derivative was stable and less polar so that SPME-GC/MS could be applied for extraction, separation and detection. The automated analytical method had a limit of detection (LOD) of 0.5 ng mL(-1), a limit of quantification (LOQ) of 2 ng mL(-1), a repeatability of 9.5%, and a linear range from 2 to 500 ng mL(-1). The method was applied to analyze fruit samples. And it was found that the concentrations of furaneol in tomato ranged from 95 to 173 μg kg(-1), in strawberries ranged from 1663 to 4852 μg kg(-1). The results were verified with a LC procedure. To facilitate analytical method development, some physico-chemical parameters for furaneol were determined in this work. Its solubility in water was determined as 0.315 g mL(-1) (25°C). Its LogD in water and LogP in 0.1 M phosphate buffer were -0.133 and 0.95 (20 °C), respectively. Its pKa was 8.56 (20 °C).     

毛细管电泳-柱端安培检测测定莲子心中荷叶碱、芦丁和金丝桃甙的含量

采用毛细管电泳-柱端安培检测测定莲子心中荷叶碱、芦丁和金丝桃甙的含量．研究了检测电位、运行缓冲液浓度和pH值,分离电压和进样时间对分离和检测的影响．以微碳圆盘电极（Ф=0.5ram）为工作电极,检测电位为＋0．95V（vs．Ag／AgCl）,pH为7．25的50mmol／L Na2B4O7和100mmol／L NaH2PO4缓冲液为运行液,当分离电压为15kV时,3种分析物在15min内完全分离．荷叶碱、芦丁和金丝桃甙的检出限（S／N=3）分别为0．02μg／mL、0．05μg／mL和0．04μg／mL．该方法已成功地应用于莲子心中上述3种活性成分的测定．     

Simultaneous HPLC determination of thiocolchicoside and glafenine as well as thiocolchicoside and floctafenine in their combined dosage forms

Sensitive and accurate high-performance liquid chromatographic methods have been developed for the simultaneous determination of thiocolchicoside (TC)-glafenine (GF) (Mix I) and thiocolchicoside-floctafenine (FN) (Mix II) in their pharmaceutical formulations. The analysis for both mixtures was performed using 250 mm × 4.6 mm i.d., 5 μm particle size C18 Waters Symmetry column. The mobile phase consisted of methanol-0.035 M phosphate buffer (50:50, v/v) of pH 4.5 for Mix I and methanol-0.03 M phosphate buffer (70:30, v/v) of pH 4 for Mix II with flow rate of 1 mL/min and UV detection at 400 nm in both cases. The calibration plots were rectilinear over the concentration range of 0.2-2 μg/mL for TC in both mixtures and 20-200 μg/mL for each of GF and FN . The limits of detection for TC and GF were 0.05 μg/mL and 0.62 μg/mL, respectively, and for TC and FN were 0.02 μg/mL and 0.70 μg/mL, respectively. Additionally, the proposed methods were successfully applied to their combined tablets with average percentage recoveries of 100.35 ± 0.61 and 100.57 ± 0.72% for TC and GF respectively and for TC and FN the percentage recoveries were 101.2 ± 0.72 and 100.36 ± 0.67%, respectively. The results obtained were favorably compared with those given using the comparison methods.     

Fast and sensitive high performance liquid chromatography analysis of cosmetic creams for hydroquinone, phenol and six preservatives

A fast and sensitive HPLC method for analysis of cosmetic creams for hydroquinone, phenol and six preservatives has been developed. The influence of sample preparation conditions and the composition of the mobile phase and elution mode were investigated to optimize the separation of the eight studied components. Final conditions were 60% methanol and 40% water (v/v) extraction of the cosmetic creams. A C18 column (100 mm × 2.1 mm) was used as the separation column and the mobile phase consisted of methanol and 0.05 mol/L ammonium formate in water (pH=3.0) with gradient elution. The results showed that complete separation of the eight studied components was achieved within 10 min, the linear ranges were 1.0-200 μg/mL for phenol, 0.1-150 μg/mL for sorbic acid, 2.0-200 μg/mL for benzoic acid, 0.5-200 μg/mL for hydroquinone, methyl paraben, ethyl paraben and propyl paraben, butyl paraben, and good linear correlation coefficient (≥0.9997) were obtained, the detection limit was in the range of 0.05-1.0 μg/mL, the average recovery was between 86.5% and 116.3%, and the relative standard deviation (RSD) was less than 5.0% (n=6). The method is easy, fast and sensitive, it can be employed to analyze component residues in cosmetic creams especially in a quality control setting.     

大体积进样-乙腈盐堆积-胶束扫集毛细管电动色谱法对马来酸氯苯那敏的测定

建立了大体积进样-乙腈盐堆积-胶束扫集毛细管电动色谱法测定马来酸氯苯那敏片中马来酸氯苯那敏的新方法,并考察了样品中乙腈和NaCl浓度对分离效果的影响.结果表明,以12 mmol/L四硼酸钠-50 mmol/L硼酸- 50 mmol/L十二烷基硫酸钠(SDS)为缓冲液(含10%甲醇,pH9.1),以70%乙腈- 200m...