High-performance liquid chromatographic determination of phenylephrine and its conjugates in human plasma using solid-phase extraction and electrochemical detection

An HPLC method for the determination of phenylephrine and its conjugates in human plasma was developed and validated. The method for quantitation involved extraction of diluted plasma (subject to hydrolysis with β-glucuronidase for 30 min with 500 units of enzyme per 0.1 ml of plasma at 37°C for the conjugates) on solid-phase weak cation-exchange cartridges followed by elution of the analyte and the internal standard (ethylnorphenylephrine) with 5% triethylamine in methanol. Analysis was carried out on a 15 cm ODS stationary phase using ion-pair reversed-phase chromatography. An electrochemical detector operated at + 1.15 V vs. Ag/AgCl was employed for detection. The standard curves were linear in the range 1.0–50.0 ng ml⁻¹ for phenylephrine and 25.0–500.0 ng ml⁻¹ for phenylephrine obtained from its conjugates. The limit of quantitation was 2.0 ng ml⁻¹ (RSD = 17%) and 25.0 ng ml⁻¹ (RSD = 18%), respectively. Acceptable accuracy and precision were obtained during intra- and inter-batch analyses for both the assays.     

High-performance liquid chromatographic determination of plasma propylthiouracil

A reversed-phase high-performance liquid chromatographic analysis was developed for propylthiouracil in plasma (1 ml). After protein precipitation with acetonitrile, the solution was diluted with water and injected into a liquid chromatograph equipped with C-18 and C-8 columns in series. The peak area was linear over the 0.25-10-mg/liter range, and the recovery was 101 +/- 4.5%. This assay has the advantages of specificity, simplicity, and speed over previously published methods and requires smaller sample volumes. None of 19 drugs tested interfered with the assay.     

High-performance liquid chromatographic determination of methotrexate in plasma

A procedure for the determination of methotrexate in human plasma is reported. The analysis involved extraction of methotrexate as an ion pair in ethyl acetate. Reconstituted residue was analyzed using reverse phase C-18 column and a mobile phase consisting of acetate buffer (87%), methanol (6.5%), and acetonitrile (6.5%). The methotrexate recovery range was 95-97%. Theophylline was used as internal standard with a recovery of 96%. The intraday coefficient of variation for the assay ranged from 1.8-3.0%, while interday variation coefficient range was 3.5-3.7%. The method is selective, reproducible, and covers a wide range of methotrexate concentrations in patient's plasma.     

High-performance liquid chromatographic determination of ethosuximide in plasma

A HPLC method for a rapid determination of ethosuximide in plasma is described. A sample is extracted with ethyl ether from acidic medium with glutethimide as internal standard and injected onto analytical RP-18 column with spectrophotometric monitoring at 244 nm.     

反相高效液相色谱法测定人血浆中萘普生含量

目的：建立ＲＰ－ＨＰＬＣ测定人血浆中萘普生含量的方法，研究该药在人体内药物动力学。方法：血浆在酸性介质中以氯仿提取，Ｃ１８色谱柱，２３０ｎｍ波长紫外检测，以甲基炔诺酮为内标，流动相为甲醇∶水（６５∶３５，预先用磷酸调至ｐＨ３．０～３．１）；所得药时数据采用ｍｉｎｉｍ３．０８程序拟合计算药动学参数。结果：萘普生血药浓度在０．１～２００μｇ·ｍｌ－１范围内与色谱峰高比呈线性关系，ｒ＝０．９９９９（ｎ＝８）。４名健康受试者单剂量口服５００ｍｇ萘普生片后，药时曲线呈一室模型。Ｃｍａｘ＝７３．６±８．３μｇ·ｍｌ－１，Ｔｍａｘ＝２．３±０．５ｈ，Ｔ１／２Ｋｅ＝９．７±３．５ｈ，Ｋｅ＝０．０８±０．０４ｈ－１，ＡＵＣ０→２４＝８２０．０±８５．７μｇ·ｈ·ｍｌ－１。结论：本法简单、快捷、灵敏，能满足药动学研究的需要     

High-performance liquid chromatographic determination of ciprofloxacin in plasma samples

A new bioanalytical high-performance liquid chromatographic (HPLC) method for the determination of ciprofloxacin with norfloxacin as an internal standard was developed and validated for plasma samples. Norfloxacin is structural homologue of ciprofloxacin and exhibits similar retention properties. The quality of respective peak separation is strongly influenced by amphoteric character of ciprofloxacin and norfloxacin as well. In previously published HPLC methods on conventional C18 reversed-phase [F. Belal, A.A. Al-Majed, A.M. Al-Obaid, Talanta 50 (1999) 765–786; G. Carlucci, J. Chromatogr. A 812 (1998) 343–367], ion pair reagents were added into the mobile phase to suppress peak tailing. In comparison with endcapped and high purity silica reversed-phase sorbent (Purospher RP-18e, Merck), which yielded symmetrical peaks, separation efficiency was further enhanced in our method. Gradient elution mode using acetonitril and phosphate buffer pH 3 on the pentafluorophenylpropyl stationary phase (250–4.6 mm Discovery^® HS F5, 5 μm, Supelco) was carried out. The resolution of 4.1 for ciprofloxacin–norfloxacin peaks was achieved. Sample preparation by SPE C18 (Supelclean) with recovery 72% was performed. Fluorescence detection with λ_excit = 280 nm, λ_emis = 446 nm was used. After the validation, the bioanalytical HPLC method was applied to pharmacokinetic studies.     

高效液相色谱法测定血浆中伊立替康的含量

目的建立血浆中伊立替康含量的测定方法。方法以C18色谱柱、0．05mol／LNa2HPO4(pH=3．0,内含0．05mol／L辛烷基磺酸钠)-乙腈=68：32为流动相分离、荧光检测伊立替康,外标法定量。结果伊立替康浓度在25．0-1000μg／L内,浓度与峰面积之间有良好的线性关系(C=0．00249A-8．15,r=0．9994)。最小检出质量浓度为2．0μg／L。50,100,500μg／L伊立替康的相对回收率(％)分别为96．4、98．3和100．4。三个浓度的平均日内RSD为2．87％,平均日间RSD为4．30％。结论本方法快速、简便、准确,可用于科研和临床工作中伊立替康血药浓度的快速检测。     

High-performance liquid chromatographic assay of clonazepam in human plasma using a non-porous silica column

A new HPLC method has been developed for measuring clonazepam (CZP) in plasma, using a reversed-phase non-porous silica column packed with 2 microm particles. CZP in plasma was first purified with a column extraction technique and injected onto a non-porous silica column. The calibration curve was linear from 5-200 ng/ml. The recoveries of CZP added to plasma were more than 94.0%, with a coefficient of variation in the range of 5.1-13.8%. We developed a rapid routine method using a non-porous silica column that was accurate and improved solvent consumption in the measurement of CZP.     

High-performance liquid chromatographic determination of sulfinpyrazone and its metabolites in plasma

A specific sensitive reversed-phase high-performance liquid chromatographic method for simultaneous determination of sulfinpyrazone (Anturane) and five of its metabolites (p-hydroxy-sulfinpyrazone, p-hydroxy-sulfone, sulfone, p-hydroxy-sulfide, and sulfide) in plasma is described. The compounds were extracted from plasma, back-extracted, re-extracted and separated on a 10-micron muBondapak C18 column using 10 mmol/l orthophosphoric acid-acetonitrile-ethanol as the mobile phase. Phenylbutazone was used as internal standard, which was totally separated from all other compounds. The lower limit of sensitivity for sulfinpyrazone, sulfone, p-hydroxy-sulfide and sulfide is 30 ng/ml, and 100 ng/ml for p-hydroxy-sulfinpyrazone and p-hydroxy-sulfone. Concentrations of sulfinpyrazone between 0.5 and 25 micrograms/ml were measured with an average coefficient of variation of 5.1%. Examples of application of this assay are given.     

High-performance liquid chromatographic method for the determination of tiflamizole in plasma

A sensitive, specific, high-performance liquid chromatographic procedure was developed for the measurement of plasma tiflamizole levels. Acidic plasma samples were extracted with three volumes of ether. The ether extracts were combined and evaporated to dryness. The residue was dissolved in acetonitrile, washed with hexane, and the acetonitrile was evaporated to dryness. The residue was dissolved in 0.5 mL of mobile phase consisting of acetonitrile and 0.007 M pH 3 sodium phosphate buffer (70:30, v/v) and then chromatographed on a octadecylsilane bonded microparticulate silica column. The assay is specific, precise, accurate, and can measure 10 ng of tiflamizole in 5 mL of plasma. The method was applied to human pharmacokinetic studies.     

High-performance liquid chromatographic analysis of nitrendipine in human plasma using ultraviolet detection and single-step solid-phase sample preparation

A high-performance liquid chromatographic (HPLC) method was developed for the determination of nitrendipine in human plasma using solid-phase extraction (SPE) and ultraviolet detection. A 30-microl aliquot of methanol (containing 2 microg/ml of the internal standard, nimodipine) was added to a 1-ml aliquot of biological sample. After vortex-mixing, the mixture was loaded on C(18) SPE cartridge which was conditioned with methanol and distilled water. After washing with distilled water, the SPE cartridge was eluted with 1-ml aliquot of diethyl ether. The organic phase was collected and evaporated under nitrogen gas. The residue was then reconstituted with a 100 microl aliquot of mobile phase, and a 50 microl aliquot was injected onto the C(18) reverse-phased column. The mobile phase, 10 mM phosphate buffer (pH 4.5):acetonitrile (50:50, v/v), was run at a flow rate of 1.2 ml/min. The column effluent was monitored using ultraviolet detector at 238 nm. The retention times for nitrendipine and the internal standard were approximately 10.1 and 12.6 min, respectively. The detection limit of nitrendipine in human plasma was 2.0 ng/ml. The coefficients of variation (CV) of the assay were below 16.5% for human plasma, and no interferences from endogenous substances were found. This specific, accurate and precise assay was useful in the study for the pharmacokinetic characteristics of nitrendipine.     

High-performance liquid chromatographic determination of doxorubicin in tissues after solid phase extraction

A method is described for the determination of doxorubicin in tissues. The drug was selectively extracted from the biological matrix by solid phase extraction using 1-ml octadecyl silane extraction columns. Prior to extraction, tissue samples were digested by an enzymic digestion procedure. Daunorubicin was used as an internal standard. Quantitation was by high-performance liquid chromatography (HPLC) using ion-pair chromatography on a reversed-phase column. Detection was by fluorescence.

Recovery of doxorubicin from tissue was 81.7 ± 2.1% mean ±SD. Doxorubicin concentrations as low as 0.01 mg kg⁻¹ could be determined. A typical value of the relative standard error of measurement was 3.7% at 2.1 mg kg⁻¹.     

External-standard high-performance liquid chromatographic method for quantitative determination of furosemide in plasma by using solid-phase extraction and on-line elution.

A rapid and simple external-standard high-performance liquid chromatographic (HPLC) method has been developed for the determination of the concentration of furosemide in plasma. The analyte is extracted with a C-2 ethyl sorbent. On-line elution of the analyte into the HPLC system is accomplished with an advanced automated sample processor (Varian). Furosemide is quantified by fluorescence detection within a linear range of 25 to 1000 ng/mL (average correlation coefficient, 0.9998), with a limit of detection of 1.8 ng/mL. Both internal- and external-standard procedures were evaluated, and the external-standard procedure demonstrated superior characteristics. The external-standard procedure was precise to within a relative standard deviation of 8% and accurate with less than 3% error throughout the concentration range studied. The external-standard HPLC method was used to analyze the concentration of furosemide in greater than 1000 plasma samples obtained from patients with either normal kidney function or renal failure who had received furosemide either orally or intravenously in an experimental setting.     

Determination of acyclovir in plasma by solid-phase extraction and column liquid chromatography

After oral administration, valacyclovir, the L-valyl ester of acyclovir, converts to the antiherpes virus drug, acyclovir. The bioavailability of acyclovir after valacyclovir administration is between 3- to 4.5-fold higher than that achieved after oral acyclovir administration. Therefore, despite the drug's short terminal half-life (3 hours), acyclovir plasma concentrations obtained after oral administration of the prodrug offer a more convenient dosage regimen in patients with herpes zoster than that required after acyclovir administration. Acyclovir is also used for viral infection prophylaxis in patients with hematologic disorders and in those who have undergone solid organ transplantation. We have described a simple and selective liquid chromatographic method for the determination of acyclovir in plasma using a new polymeric reversed-phase sorbent for solid-phase extraction. A mean acyclovir absolute recovery of 90% was found after elution of the drug from the cartridge with the mobile phase. This procedure allowed us to measure 62.5 ng/mL of acyclovir with an acceptable precision using a plasma volume of 250 microL, and no drug was found to interfere with the assay. This method is suitable for the therapeutic monitoring of acyclovir in patients who have been given a wide variety of coadministered drugs.     

High-performance liquid chromatographic determination of cis-dichlorodiammineplatinum(II) in plasma ultrafiltrate

A reproducible, simple and sensitive high-performance liquid chromatographic method was described for the quantitative analysis of cis-diamminedichloroplatinum(II) (CDDP) in ultrafiltrate plasma in the presence of nickel chloride as internal standard. CDDP and the internal standard were chelated by exchange with diethyldithiocarbamate. After derivatization, the mixture was directly injected into the column. Chromatography was performed on an Ultrasphere column and the eluent measured spectrophotometrically at 260 nm for CDDP and at 250 nm for the internal standard. The peak area ratio of CDDP to the internal standard varied linearly with concentration over the range 0.05–10 μg ml⁻¹. Precision and reproducibility were both excellent and the limit of quantification was 0.03 μg ml⁻¹ using only 0.5 ml of ultrafiltrate. The present method, without extraction, should be entirely automated. This assay may be suitable for therapeutic drug monitoring in patients receiving CDDP.     

High-performance liquid chromatographic assay for the determination of nilotinib in human plasma

A precise and convenient high-performance liquid chromatography (HPLC) method has been established to assay nilotinib in human plasma. Chromatographic separation of nilotinib was performed on a LiChrosphere(?)100 RP-18(e) column (250 mm×4.0 mm, 5 μm) using a mixture of acetonitrile and 0.01 M phosphate buffer (pH 3.0) (42 : 58, v/v) under isocratic conditions at a flow rate of 1.0 ml/min with ultraviolet (UV) detection at 266 nm. The calibration curve showed linearity at concentrations between 250 ng/ml and 5000 ng/ml (r(2)>0.999). The mean±S.D. absolute recovery of nilotinib from plasma was 99.2±3.3%. The coefficients of variation of both intra- and inter-day precision were below 9.1%. These results indicate that this new HPLC-based quantification may be useful for therapeutic drug monitoring of nilotinib to help manage treatment in patients with chronic myeloid leukemia in clinical practice.     

Validation of an automated liquid chromatographic method for omeprazole in human plasma using on-line solid-phase extraction

An automated system using on-line solid-phase extraction and HPLC with UV detection has been validated in order to determine omeprazole in human plasma. The extraction was carried out using C18 cartridges. After washing, omeprazole was eluted from the cartridge with mobile phase onto an Inertsil ODS-2 column. The developed method was selective and linear for drug concentrations ranging between 5 and 500 ng ml(-1). The recovery of omeprazole ranged from 88.1 to 101.5%, and the limit of quantitation (LOQ) was 5 ng ml(-1). The intraday accuracy ranged from 93.1 to 106.2% and the interday accuracy varied from 95.4 to 105.1%. For the LOQ, good values of precision (8.7 and 17.5% for intraday and interday, respectively) were also obtained. This automated system has been applied to determine omeprazole in human plasma samples from bioequivalence studies.     

High-performance liquid chromatographic determination of cephacetrile.

A rapid and accurate quantitative determination of cephacetrile in finished bulk and dosage forms is reported. The high-performance liquid chromatographic method is free of interference by acetyl hydrolysis products and synthesis by-products. The assay can be performed in about 15 min, affording less than 0.7% coefficients of variation within and between days. The chromatographic results are in good agreement with the microbiological assay requested by the "Code of Federal Regulations" for certification of cephacetrile sodium.