Analysis of pharmaceutical preparations containing local anesthetics by micellar liquid chromatography and spectrophotometric detection

Summary An HPLC procedure for the determination of six local anesthetics, bupivacaine, lidocaine, mepivacaine, procaine, propanocaine and tetracaine, in pharmaceutical silane ODS-2 C₁₈ analytical column and spectrophotometric detection at 230 nm were used. The chromatographic tographic behaviour of local anesthetics with different micellar eluents of sodium dodecyl sulphate (SDS) is described. Selection of the adequate composition of the micellar mobile phase (SDS and 1-propanol concentrations) for the analysis of pharmaceuticals was studied. Adequate retention was achieved with an eluent containing 0.15 M SDS +10% 1-propanol at pH 3. Application of the proposed method to the analysis of eight pharmaceutical formulations gave recoveries between 93 and 100.2% of the values declared by the manufacturers. The proposed procedure for the determination of local anesthetics is rapid, reliable and free from interferences.     

Determination of amobarbital and secobarbital in plasma samples using micellar liquid chromatography

A new liquid chromatographic procedure for the determination of amobarbital and secobarbital in plasma samples is proposed. The method uses a Spherisorb octadecylsilane ODS-2 C(18) analytical column, a guard column of similar characteristics and 0.04 M CTAB solution buffered at pH 7.5 containing 3% 1-propanol as micellar mobile phase. The UV detection was carried out at 250 nm. Butabarbital was used as internal standard. Plasma samples preparation only required adequate dilution with the mobile phase before injection into the chromatographic system. The limits of detection were 0.2 and 0.4 mg/L for amobarbital and secobarbital, respectively. The proposed method allows the determination of amobarbital and secobarbital in plasma at therapeutic levels.     

微乳液相色谱法测定人血浆中丙泊酚浓度

建立了基于微乳液相色谱(MELC)的人血浆中丙泊酚浓度的测定方法。采用Hypersil BDS C18色谱柱分离,并考察了微乳流动相中各组分对溶质洗脱的影响。优化的色谱条件: 以0.5%醋酸（含有3.0%十二烷基硫酸钠（SDS）,0.8%正庚烷,6.0%正丁醇）微乳为流动相,流速为1.0 mL/min,荧光检测器激发波长(λex)为274 nm、发射波长(λem)为312 nm,柱温为室温。人血浆样品用流动相稀释并离心后,直接进样分析。丙泊酚在0.25～10 μg/mL质量浓度范围内呈良好的线性关系,方法的回收率为(98.2±1.9)%～(104.6±2.2)%;日内测定峰面积的相对标准偏差(RSD)为1.42%～2.43%,日间测定峰面积的RSD为2.75%～4.79%。该方法简便可行、重复性好,可用于人血浆中丙泊酚浓度的测定。     

High-performance liquid chromatographic determination of diltiazem in human plasma after solid-phase and liquid–liquid extraction

A selective, sensitive, and accurate high-performance liquid chromatographic method for determination of diltiazem in plasma samples has been developed and validated. The effects of mobile phase composition, buffer concentration, mobile phase pH, and concentration of organic modifiers on retention of diltiazem and internal standard were investigated. Solid-phase and liquid–liquid extraction were examined and proposed for isolation of the drug and elimination of endogenous plasma interferences. A 5 m Lichrocart Lichrospher 60 RP-select B chromatographic column was used; the mobile phase was acetonitrile–0.025 mol L^–1 KH₂PO₄ (pH 5.5), 35:65 ( v / v) at a flow-rate of 1.5 mL min^–1. The detection wavelength was 215 nm. The calibration plots were linear in the concentration range 20.0–500.0 ng mL^–1. The method has been implemented to monitor diltiazem levels in patient samples.     

Chromatographic analysis of penstemide and serrulatoloside by high-performance liquid chromatography and gradient thin-layer chromatography

Summary A sensitive capillary gas chromatographic method is described for the simultaneous determination of lidocaine, tetracaine, procaine and dibucaine. The method was applied to the determination of anesthetics in tissue homogenates incubated at 38°C at doses between 10 and 400 mg/kg. In the liver tissue thein vitro metabolization of the studied anesthetics is most rapid for tetracaine, also fast for procaine, while for lidocaine and dibucaine the metabolization is very slow. In brain tissue thein vitro metabolization of anesthetics is very slow.The method shows good analytical parameters: linearity between 5 and 40 g/ml; day-to-day reproducibility ca. 8% for a concentration of 20 g/ml, precision ca. 7% for a concentration of 20g/ml. Accuracy is also very good.     

Chromatographic determination of caffeine in pharmaceutical formulations using micellar mobile phases

Summary An HPLC procedure is described for the determination of caffeine in pharmaceutical preparations. A Spherisorb octadecylsilane ODS-2 C₁₈ analytical column and spectrophotometric detection at 273 nm were used. The chromatographic behaviour of caffeine with different micellar eluents containing sodium dodecyl sulphate (SDS) is described. The determination of caffeine in pharmaceutical preparations was performed by use of a mobile phase containing 0.05 M sodium dodecylsulphate (SDS) and 1.5% propanol at pH7. At a 6.0 g mL^–1 concentration level the peak area and peak height repeatability were 2.6 and 2.4%, respectively. The application of the proposed method to the analysis of five pharmaceutical formulations, using peak heights as the dependent variable, gave recoveries between 85 and 104% of the values declared by the manufacturers. The proposed procedure for the determination of caffeine is rapid (15 min per sample), reliable and free from interferences.     

Development of an Ion-pair HPLC method for the determination of 2,4- and 2,6-toluendiamine in human plasma

Summary An ion-pair HPLC method is presented for the determination in plasma of 2,4- and 2,6-toluendiamine (TDA), known carcinogens. The chromatographic conditions consisted in isocratic elution on a reversed phase C₁₈ column with 5 mM octanesulfonic acid in methanol-water, 45∶55, as mobile phase. UV detection was performed at 235 nm. Samples were analyzed after a simple single step liquid-liquid extraction and the method was validated by measurement of precision (interassay and intraassay), sensitivity, specificity, linearity, and recovery. The detection limit for both TDAs in plasma was 20 ng mL⁻¹. The correlation coefficients based on the intrassay calibration curve were 0.998 and 0.997 for 2,6- and 2,4-TDA, respectively. The intraassay accuracy, expressed in terms of recovery, was found to be up to 89.91% and 97.05% for 2,6- and 2,4-TDA, respectively.     

High-performance liquid chromatographic analysis of azlocillin in serum

Summary A rapid and sensitive high performance liquid chromatographic method is described for the determination of azlocillin in serum. This method involves a short manual protein precipitation of the sample followed by an injection into a PR 18 column for separation and quantitation. The mobile phase was a 22% (V/V) solution acetonitrile in phosphate buffer pH 4.8 at a flow rate of 2,5 ml/min. The spectrophotometer detector was set at 220 nm with a sensitivity of 0.08 AUFS.     

Optimization and Validation of an HPLC-UV Method for Determination of Tranexamic Acid in a Dosage Form and in Human Urine

A simple and reliable high-performance liquid chromatographic method with UV detection at 245 nm has been developed and validated for determination of tranexamic acid (TA) in a dosage form and in human urine. Before injection samples were derivatized with phenyl isothiocyanate (PITC). The reaction temperature, reaction time, and concentration of PITC used for derivatization were optimized. Chromatographic separation was on a C₁₈ column with a 65:35 (v/v) mixture of 10 mM phosphate buffer, pH 3.6, and acetonitrile as mobile phase. Heptaminol hydrochloride was used as internal standard.     

Determination of Veterinary Pharmaceuticals in Production Wastewater by HPTLC-Videodensitometry

An SPE-HPTLC method for simultaneous identification and quantification of seven pharmaceuticals in production wastewater was optimized and validated. The studied compounds were enrofloxacine, oxytetracycline, trimethoprim, sulfamethazine, sulfadiazine, sulfaguanidine and penicillin G/procaine. The method involves solid-phase extraction on hydrophilic-lipophilic balance cartridges with methanol and HPTLC analysis of extracts on CN modified chromatographic plates followed by videodensitometry at 254 and 366 nm. Optimization of chromatographic separation was performed by systematic variation of the mobile phase composition using genetic algorithm approach and the optimum mobile phase composition for TLC separation was 0.05 M H₂C₂O₄:methanol = 0.81:0.19 (v/v). Linearity of the method was demonstrated in the ranges from 1.5 to 15.0 μg L⁻¹ for enrofloxacine, 100–500 μg L⁻¹ for oxytetracycline, 150–600 μg L⁻¹ for trimethoprim, 300–1100 μg L⁻¹ for sulfaguanidine and 100–400 μg L⁻¹ for sulfamethazine, sulfadiazine and penicillin G/procaine with coefficients of determination higher than 0.991. Mean recoveries ranged from 74.6 to 117.1% and 55.1 to 108.0% for wellspring water and production wastewater, respectively. Only sulfaguanidine showed lower results. The described method has been applied to the determination of pharmaceuticals in wastewater samples from pharmaceutical industry.     

高效液相色谱法同时测定假尿核苷和肌酐的研究及应用

本文用高效液相色谱法同时测定人尿中假尿核苷和肌酐,研究了流动相的pH值变化对色谱行为的影响,方法简便、快速,已成功地用于对正常人、肺部感染及肺癌患者的尿样分析,其结果有显著性差异。     

应用质量源于设计的理念优化大鼠血浆中大黄蒽醌的分析方法

应用质量源于设计理念建立一种高效液相色谱-荧光检测法(HPLC-FLD)用于测定大鼠血浆中5种大黄蒽醌。用Plackett-Burman设计考察流动相中甲醇含量、pH值、流速、柱温和进样体积对色谱峰的分离度、理论塔板数、最末洗脱峰的保留时间和拖尾因子的影响,结果显示流动相中甲醇含量、流速和柱温对色谱系统的影响显著(p<0.05)。继而采用Box-Behnken设计结合响应面法考察上述三因素对分离度、保留时间和理论塔板数的影响。用Derringer渴求函数评价了响应值的综合作用。得出最优色谱条件为:以甲醇-0.1%(v/v)磷酸水溶液(81.4:18.6, v/v)为流动相等度洗脱,流速1.1 mL/min,柱温31℃,荧光检测激发波长为440 nm,发射波长为540 nm。建立的模型显示良好的预测性。结果表明:质量源于设计的理念可有效地应用于优化高效液相色谱分析方法。     

Development and Validation of a Micellar Liquid Chromatographic Method with UV Detection for Determination of Azithromycin in Tablets and Capsules

A rapid and simple micellar liquid chromatographic method that does not require use of specific chromatographic columns has been developed and validated for azithromycin determination. The method uses a Hypersil C₁₈ column at 60 °C, 1-butanol–pH 6.86 phosphate buffer solution–water, 15:25:60 (v/v), containing 0.10 M sodium dodecyl sulfate, as mobile phase, and UV-detection at 215 nm. Different characteristics of the method were validated satisfactorily. The specificity, accuracy, linearity, precision (repeatability), and robustness of the method were demonstrated. The method proved suitable for determination of the azithromycin content of capsules and uncoated tablets.Revised: 5 February and 11 March 2004     

Separation and determination of Ni(II), Co(II) and Cu(II) by RP-HPLC with hexadecyltrimethylammonium bromide/n-propanol mobile phases

Summary The effect of the presence of a cationic surfactant, hexadecyltrimethylammonium bromide (CTAB), and a short chain alcohol,n-propanol, in the mobile phase on the chromatographic retention of Co(II), Ni(II) and Cu(II) diethylammonium diethyldithiocarbamate complexes, has been studied. A simple isocratic reversephase method for the determination of the metals is proposed, using a mobile phase of composition CTAB 0.03 M/n-propanol 45 %v/v. Detection limits at pg levels were obtained for all solutes. The method was applied to the determination of the complexes in tap water samples at three different concentrations, with recoveries close to 100%. In order to evaluate the interaction between the metal complexes and the aggregates formed, the values of solute binding constants are calculated.     

Development and validation of a rapid HPLC method for the determination of five banned fat-soluble colorants in spices using a narrow-bore monolithic column

This work reports a fast and simple liquid chromatographic method for the simultaneous determination of five banned fat-soluble synthetic colorants, namely Sudan I-IV and Para-Red, in spice samples. The analytes were successfully separated isocratically in less than 5 min on the new narrow bore monolithic column, FastGradient^® Chromolith (50 mm × 2.0 mm i.d.) using a mobile phase of 0.1% (v/v) HCOOH/acetonitrile (35/65%, v/v) at a flow rate of 1.5 mL min⁻¹. All colorants were detected at 506 nm. The main parameters (mobile phase composition, flow rate, injection volume) affecting the separation were studied. The proposed method was thoroughly validated in terms of linearity, LODs, precision and accuracy. The method was applied to the determination of the studied azo-dyes in various spices (paprika, chilli and mixed spice powders) after ultrasound-assisted extraction. Satisfactory recoveries, ranging from 92% to 109% were obtained.     

HPLC Determination and Pharmacokinetic Study of Valdecoxib in Human Plasma

A sensitive, simple, and accurate high-performance liquid chromatographic method has been developed for determination of valdecoxib and the internal standard rofecoxib in human plasma. Protein was precipitated from plasma samples by addition of perchloric acid (HClO₄); the drug was then extracted with diethyl ether. Separation was performed on a Cosmosil C₁₈ column (150 mm × 4.6 mm i.d., 5 m particles) with ammonium acetate buffer-acetonitrile, 60:40 (v/v), containing 0.1% TEA, pH 6.5, as mobile phase. Detection and quantification were performed by UV-visible detection at 239 nm. Detection and quantification limits were 3 and 5 ng mL^–1, respectively. The linear concentration range for valdecoxib was 5–400 ng mL^–1. The validated RP HPLC method was used for determination of the pharmacokinetic data for the drug in humans.     

HPLC with Diode-Array Detection for Determination of Leflunomide in Tablets

We have developed and validated a simple HPLC method for analysis of leflunomide in tablets. Method conditions were determined by assay of a photodegraded sample of leflunomide. Optimum chromatographic performance was obtained with a C₁₈ column and acetonitrile-water as mobile phase. Comparison of spectra recorded with a diode-array detector during elution of the leflunomide peak enabled determination of method specificity. The method is highly sensitive (detection limit 10 ng mL⁻¹) and robust to deliberate variation of the conditions (RSD of peak area < 2.0%). Precision and accuracy were adequate over the concentration range 10 to 100 μg mL⁻¹. These results show the proposed method is suitable for its intended use.     

High-Performance Liquid Chromatographic Determination of Cefepime and Cefazolin in Human Plasma and Dialysate

A simple high-performance liquid chromatographic (HPLC) method was developed for the simultaneous determination of cefepime and cefazolin in human plasma and dialysate. For component separation, the method utilized a C₁₈ column with an aqueous mobile phase of dibasic potassium hydrogen phosphate (pH 7.0) and methanol gradient at a flow rate of 1 mL min⁻¹. The method demonstrated linearity from 2.0 to 100.0 μg mL⁻¹ (r > 0.999) with detection limit of 1 μg mL⁻¹ for both cefepime and cefazolin. The method was utilized for evaluation of plasma and dialysate samples in a clinical study evaluating the dialyzer clearance of cefepime and cefazolin using high-flux hemodialysis with varying blood flow rates in chronic kidney failure patients undergoing hemodialysis and peritoneal dialysis treatment.     

高效液相色谱法测定人血浆中总高半胱氨酸含量

建立了快速检测人血浆中总高半胱氨酸含量的高效液相色谱（HPLC）方法。采用反相HPLC分析前,以巯基特异性荧光试剂ABDF对血浆中巯基进行衍生．方法简便、灵敏、准确,无干扰峰影响。平均回收率为97．75％,日内和日间精密度分别为4．49％和9．79％。     

Hydrophilic interaction liquid chromatographic tandem mass spectrometric determination of atenolol in human plasma

A hydrophilic interaction liquid chromatographic method with tandem mass spectrometry for the determination of atenolol, a beta-blocking agent, in human plasma has been developed and validated over the curve range of 10--2000 ng/mL. The assay was based on protein precipitation followed by evaporation of the extraction solvent, reconstitution with acetonitrile, and chromatography on an Hypersil silica column (50 x 4.6 mm) using a low aqueous--high organic mobile phase. The mobile phase consists of 85% acetonitrile, 15% water, 0.5% acetic acid and 0.04% trifluoroacetic acid and runs isocratically at a flow rate of 2.0 mL/min. The column ef fluent was split so that 50% of it was transferred into the LC-MS/MS interface operated in positive electrospray ionization mode. The chromatographic run time was 2.0 min per injection. Atenolol and the internal standard, atenolol-d(7), showed a retention time of 1.0 min. The inter-day and intra-day precision and accuracy of the quality control samples were <5.3% relative standard deviation and <8.0% relative error, respectively. To explore the application of the current method for the analysis of other beta-blocking agents, propranolol and metoprolol were tested under the same chromatographic conditions with retention times of 0.68 and 0.75 min, respectively. The present method could be used for therapeutic drug monitoring, pharmacokinetic and drug--drug interaction studies of beta-blocking agents.