Simple and rapid RP-HPLC method for simultaneous determination of acyclovir and zidovudine in human plasma

Combination therapy with acyclovir and zidovudine is used for the treatment of herpes-infected immunocompromised patients. In the view of the optimal drug concentrations (minimum effective concentrations) for viral suppression and avoidance of drug toxicity, monitoring of drug levels has been considered essential to determine drug concentrations in plasma after administration of a dose of acyclovir and zidovudine. A simple, precise, and rapid RP-HPLC method has been developed for this purpose. Chromatographic separation was performed using methanol-water (50 + 50, v/v), pH 2.5 adjusted with orthophosphoric acid, as an isocratic mobile phase at a flow rate of 0.8 mL/min with an Inertsil ODS (C18) column (5 microm particle size, 250 x 4.60 mm id). Detection was carried out using a UV photo diode array detector at 258 nm. The plasma samples were prepared by a protein precipitation method. The retention time for acyclovir and zidovudine was 3.5 +/- 0.2 and 6.2 +/- 0.3 min, respectively. The method was linear in the range of 200-1800 and 400-3600 ng/mL with LOQ of 200 ng (SD = +/-1.4) and 400 ng (SD = +/-0.9) for zidovudine and acyclovir, respectively, in plasma. The mean accuracy was 98.0 and 96.4%, with average extraction recovery of 64.8 +/- 2.1 and 77.5 +/- 1.7% for lower nominal concentrations of acyclovir and zidovudine, respectively.     

Determination of ng/mL levetiracetam using ultra-high-performance liquid chromatography-photodiode absorbance

This paper demonstrates the analysis of levetiracetam, a new chiral antiepileptic drug, at ng/mL levels using an ultra-high-performance liquid chromatography (UHPLC)-photodiode absorbance (PDA) method. Three different sample preparation methods, liquid-liquid extraction with Extrelut, solid phase extraction (SPE) with Oasis HLB and Oasis MAX SPE cartridges, and protein precipitation with organic solvents were carried out. The last preparatory method is the simplest and provides the best recoveries: between 97.1% and 100.4% with RSD value below 5%. The column for separation is BEH C18 column (1.7 μm particle size and 100 × 2.1 mm i.d.) and acetonitrile-phosphate buffer (pH = 6.6; 0.01 M) (10/90 v/v) is the mobile phase. The results obtained are compared to analysis conducted by the HPLC method. The UHPLC method was validated in the range of 2-100 μg/mL levetiracetam concentration (R(2) = 0.9997). LOD and LOQ are 10 ng/mL and 33 ng/mL, respectively. The developed UHPLC method was applied to plasma samples of patient with epilepsy.     

Validation of high-performance liquid chromatographic-mass spectrometric method for the analysis of lidocaine in human plasma

A sensitive and simple liquid chromatography-tandem mass spectrometry method is developed and validated for the determination of lidocaine in human plasma. Bupivacaine is used as an internal standard, and the plasma extraction is performed by a simple liquid-liquid extraction. The limit of quantitation (LOQ) is 0.5 ng/mL with a signal-to-noise ratio greater than 5. The calibration curve is linear from 0.5 to 250 ng/mL with an r2 greater than 0.99. The coefficients of variation for within- and between-assay imprecision, including LOQ, are < or = 13% and < or = 8%, respectively. The percentage of inaccuracy for within- and between-assay, including LOQ, are < or = 9% and < or = 5%, respectively. The absolute recovery of lidocaine and bupivacaine are greater than 84% and 82%, respectively. The higher sensitivity and accuracy of this method allow the measurement of low concentrations of lidocaine in plasma from a clinical study of topically applied lidocaine in healthy subjects.     

Fast quantification of cyclophosfamide and its N‐dechloroethylated metabolite 2‐dechloroethylcyclophosphamide in human plasma by UHPLC‐MS/MS

A rapid, novel and reliable UHPLC‐MS/MS method was developed and validated for simultaneous determination of cyclophosphamide (CP) and its dechloroethylated metabolite, 2‐dechloroethylcyclosphamide (2‐DCECP) in human plasma. The plasma samples were conducted by protein precipitation with 3‐fold acetonitrile, containing 0.1% formic acid. Mass spectrometric detection was performed using electrospray positive ionization with multiple reaction monitoring mode, using tinidazole as internal standard (IS). Chromatographic separation was performed on an Agilent poroshell 120 SB‐C₁₈ column (2.1 × 75 mm, 2.7 µm) using gradient elution of acetonitrile and 0.1% formic acid at a flow rate of 0.5 mL/min, the total run time was 2.5 min. The limit of quantification (LOQ) was 20 ng/mL for both CP and 2‐DCECP. Accuracies and precisions were <15% at LOQ and below 10% at quality control concentration levels. This UHPLC‐MS/MS method was successfully applied for the estimation of CP and 2‐DCECP in human plasma, which was also useful for clinical toxicology studies and therapeutic drug monitoring of CP. Copyright © 2014 John Wiley & Sons, Ltd.     

Validation of a high-performance chromatographic method for determination of cefotaxime in biological samples

An analytical method for detecting and quantifying cefotaxime in plasma and several tissues is described. The method was developed and validated using plasma and tissues of rats. The samples were analyzed by reversed phase liquid chromatography (HPLC) with UV detection (254 nm). Calibration graphs showed a linear correlation (r > 0.999) over the concentration ranges of 0.5–200 μg/mL and 1.25–25 μg/g for plasma and tissues, respectively. The recovery of cefotaxime from plasma standards prepared at the concentrations of 25 μg/mL and 100 μg/mL was 98.5 ± 3.5% and 101.8 ± 2.2%, respectively. The recovery of cefotaxime from tissue standards of liver, fat and muscle, prepared at the concentration of 10 μg/g was: 89.8 ± 1.2% (liver), 103.9 ± 6.5% (fat) and 97.8 ± 2.1% (muscle). The detection (LOD) and quantitation (LOQ) limits for plasma samples were established at 0.11 μg/mL and 0.49 μg/mL, respectively. The values of these limits for tissues samples were approximately 2.5 times higher: 0.3 μg/g (LOD) and 1.25 μg/g (LOQ). For plasma samples, the deviation of the observed concentration from the nominal concentration was less than 5% and the coefficient of variation for within-day and between-day assays was less than 6% and 12%, respectively. The method was used in a pharmacokinetic study of cefotaxime in the rat and the mean values of the pharmacokinetic parameters are given.     

Determination of atorvastatin and its metabolite ortho-hydroxyatorvastatin in human plasma by on-line anion-exchange solid-phase extraction and liquid chromatography tandem mass spectrometry

A rapid, sensitive, and specific method was developed and validated using liquid chromatography-tandem mass spectrometry for the simultaneous quantitation of atorvastatin (ATV) and its major metabolite ortho-hydroxyatorvastatin (o-HATV) in human plasma. The sample preparation involved a liquid–liquid extraction without chlorinated solvents and an on-line solid-phase extraction exploring the possibilities that anion exchange offers. The analytical method presented intraday and day-to-day variation below 10%; intraday and day-to-day accuracy stood between 94% and 105%; the limit of quantification was 0.1 ng/mL for ATV and 0.5 ng/mL for o-HATV; and the recovery was above 75% for both molecules. This method was applied successfully to quantitate ATV and o-HATV concentrations in an unstudied renal transplant recipient cohort treated with an immunosuppressive regime of tacrolimus and mycophenolic acid and a cohort of hypercholesterolemic patients included in the study as a control group. It can be used to evaluate patient adherence, drug–drug interactions, and pharmacokinetic/pharmacodynamic relationships. The results in our study showed that ATV and o-HATV levels in the renal transplant group were significantly increased (p < 0.001), compared to the hypercholesterolemic group.     

Quantification of doxazosin in human plasma using hydrophilic interaction liquid chromatography with tandem mass spectrometry

Hydrophilic interaction LC with MS/MS (HILIC-MS/MS) was described as a rapid, sensitive, and selective method for the quantification of doxazosin in human plasma. Doxazosin and cisapride (internal standard) were extracted from human plasma with ethyl acetate at alkaline pH and analyzed on an Atlantis HILIC Silica column with the mobile phase of ACN/ammonium formate (100 mM, pH 4.5) (93:7 v/v). The analytes were detected using an ESI MS/MS in the selective-reaction-monitoring mode. The standard curve was linear (r = 0.9994) over the concentration range of 0.2-50 ng/mL. The LOQ for doxazosin was 0.2 ng/mL using 100 microL plasma sample. The CV and relative error for intra- and interassay at four QC levels were 3.7-8.7% and 0.0-9.8%, respectively. The matrix effect for doxazosin and cisapride were practically absent. The recoveries of doxazosin and cisapride were 67.4 and 61.7%, respectively. This method was successfully applied to the pharmacokinetic study of doxazosin in humans.     

Separation,determination of six impurities in methotrexate drug substance using ultra-performance liquid chromatography

Methotrexate(MTX) is an antineoplastic therapeutic medicine as antimetabolite of folic acid. In this paper, a sensitive and rapid ultra-performance liquid chromatographic(UPLC) method was developed and validated for the separation and determination of impurities in MTX drug substances. The UPLC method was accomplished on an Agilent Zorbax Extend C-18(50 mm 4.6 mm, 1.8 mm) with a gradient elution system composed of sodium dihydrogen phosphate in water(20 mmol/L, pH 3.0) and acetonitrile. The flow rate was 2.2 mL/min. The method was validated. The calibration curves displayed good linearity(r 0.999) within the tested concentration ranges. The limit of detection(LOD) and limit of quantification(LOQ) of the six analytes were all less than 0.774 mg/mL and 1.03 mg/mL. The relative standard deviation(RSD) for intra- and inter-day precision of the six analytes was less than 9.8%, including at the LOQ. The average recovery ranged from 95.2% to 103% except at the LOQ, where recovery ranged from 82.7% to 117%. The validated method was successfully used to determine the relative abundance of six impurities in the MTX drug substances.     

Simultaneous analysis of twenty-one glucocorticoids in equine plasma by liquid chromatography/tandem mass spectrometry

A method for the simultaneous separation, identification, quantification and confirmation of the presence of 21 glucocorticoids (GCC) in equine plasma by liquid chromatography coupled with triple stage quadrupole tandem mass spectrometry (LC/TSQ-MS/MS) is described. Plasma sample augmented with the 21 GCC was extracted with methyl tert-butyl ether (MTBE) and analyzed by positive electrospray ionization. Desoxymetasone or dichlorisone acetate was used as the internal standard (IS). Quantification was performed by IS calibration. For each drug, one major product ion was chosen and used for screening for that drug. Analyte confirmation was performed by using the three most intense product ions formed from the precursor ion and the corresponding mass ratios. The recovery of the 21 GCC when spiked into blank plasma at 5 ng/mL was 45-200% with coefficient of variation (CV) from 0.3-18%. The limit of detection (LOD) and that of quantification (LOQ) for most of the analytes were 50-100 pg/mL and 1 ng/mL, respectively, whereas that of confirmation (LOC) was 100-300 pg/mL depending on the analyte. Intra- and inter-day precisions expressed as CV for quantification of 1 and 10 ng/mL was 1.0-17%, and 0.51-19%, respectively, and the accuracy was from 84-110%. The linear concentration range for quantification was 0.1-100 ng/mL (r(2) > 0.997). Estimated measurement uncertainty was from 11-37%. This study was undertaken to develop a method for simultaneous screening, identification, quantification and confirmation of these agents in post-race equine plasma samples. The method has been successfully applied to screening of a large number of plasma samples obtained from racehorses in competition and in pharmacokinetic studies of dexamethasone in the horse and concurrent changes in endogenous GCC, hydrocortisone and cortisone. The method is simple, sensitive, selective and reliably reproducible.     

Stability-indicating reversed-phase liquid chromatographic method for simultaneous determination of atorvastatin and ezetimibe from their combination drug products

A simple, precise, and rapid stability-indicating reversed-phase column liquid chromatographic (RP-LC) method has been developed and subsequently validated for simultaneous estimation of atorvastatin (ATV) and ezetimibe (EZE) from their combination drug product. The proposed RP-LC method utilizes a LiChrospher 100 C18, 5 microm, 250 x 4.0 mm id column at ambient temperature; the optimum mobile phase consists of acetonitrile-water-methanol (45 + 40 + 15, v/v/v) with apparent pH adjusted to 4.0 +/- 0.1; mobile phase flow rate of 1.0 mL/min; and UV detection at 250 nm. ATV, EZE, and their combination drug product were exposed to thermal, photolytic, hydrolytic, and oxidative stress conditions, and the stressed samples were analyzed by the proposed method. There were no other coeluting, interfering peaks from excipients, impurities, or degradation products due to variable stress conditions, and the method is specific for the estimation of ATV and EZE in the presence of degradation products. The response was linear over the concentration range of 1-80 microg/mL for ATV and EZE. The mean recoveries were 99.27 and 98.5% for ATV and EZE, respectively. The intermediate precision data were obtained under different experimental conditions, and the calculated value of the coefficient of variation was found to be less than the critical value. The proposed method can be useful in the quality control of bulk manufacturing and pharmaceutical dosage forms.     

Validation of a high-performance chromatographic method for determination of cefotaxime in biological samples

An analytical method for detecting and quantifying cefotaxime in plasma and several tissues is described. The method was developed and validated using plasma and tissues of rats. The samples were analyzed by reversed phase liquid chromatography (HPLC) with UV detection (254 nm). Calibration graphs showed a linear correlation (r > 0.999) over the concentration ranges of 0.5–200 μg/mL and 1.25–25 μg/g for plasma and tissues, respectively. The recovery of cefotaxime from plasma standards prepared at the concentrations of 25 μg/mL and 100 μg/mL was 98.5 ± 3.5% and 101.8 ± 2.2%, respectively. The recovery of cefotaxime from tissue standards of liver, fat and muscle, prepared at the concentration of 10 μg/g was: 89.8 ± 1.2% (liver), 103.9 ± 6.5% (fat) and 97.8 ± 2.1% (muscle). The detection (LOD) and quantitation (LOQ) limits for plasma samples were established at 0.11 μg/mL and 0.49 μg/mL, respectively. The values of these limits for tissues samples were approximately 2.5 times higher: 0.3 μg/g (LOD) and 1.25 μg/g (LOQ). For plasma samples, the deviation of the observed concentration from the nominal concentration was less than 5% and the coefficient of variation for within-day and between-day assays was less than 6% and 12%, respectively. The method was used in a pharmacokinetic study of cefotaxime in the rat and the mean values of the pharmacokinetic parameters are given. Received: 25 May 1998 / Revised: 27 July 1998 / Accepted: 1 August 1998     

Comparative analytical quantitation of clenbuterol in biological matrices using GC-MS and EIA

A simple and sensitive procedure utilizing GC-MS for the identification and quantitation of clenbuterol in biofluids and tissues is described. This improved method utilizes trimethylboroxine for the derivatization of clenbuterol, requires only 1 mL/g of biological sample, and most importantly does not require an extra cleaning step for urine specimens prior to extraction. Linear quantitative response curves have been generated for derivatized clenbuterol over a concentration range of 5-200 ng/mL. The extraction efficiency at four representative points of the standard curve exceeded 90% in both specimen types (plasma and urine). Linear regression analyses of the standard curve in both specimen types exhibited correlation coefficients ranging from 0.997 to 1.000. The Limit of detection (LOD) and Limit of quantitation (LOQ) values for plasma specimens were determined to be 0.5 and 1.5 ng/mL respectively. For urine specimens, LOD and LOQ values were 0.2 and 0.7 ng/microL respectively. Percentage recoveries ranged from 91 to 95% for urine and 89 to 101% for plasma. Precision and accuracy (within-run and between-run) studies reflected a high level of reliability and reproducibility of the method. In addition to its reliability, sensitivity and simplicity, this modified procedure is more efficient and cost effective, requiring less time, only 1 mL of sample, and minimal amounts of extraction solvents. The applicability of the method for the detection and quantitation of clenbuterol in biological tissues of rats treated with the drug was demonstrated successfully. For comparative analysis of clenbuterol in plasma and liver samples, both GC-MS and enzyme immunoassay (EIA) methods are found to be suitable. Due to potential antibody-cross reactivity with EIA, the GC-MS method is the method of choice for most samples because of its specificity. However, the EIA method is considered the method of choice for analysis of clenbuterol found in concentrations below the limits of quantitation by GC-MS due to its sensitivity.     

Determination of tianeptine in tablets by high-performance liquid chromatography with fluorescence detection

A sensitive and selective high-performance liquid chromatographic method has been developed for the determination of tianeptine (Tia) in tablets. The method is based on derivatization of Tia with 4-chloro-7-nitrobenzofurazan (NBD-CI). A mobile phase consisting of acetonitrile-10 mM orthophosphoric acid (pH 2.5; 77 + 23) was used at a flow rate of 1 mL/min on a C18 column. The Tia-NBD derivative was monitored using a fluorescence detector, with emission set at 520 nm and excitation at 458 nm. Gabapentin was selected as an internal standard. Linear calibration graphs were obtained in the concentration range of 45-300 ng/mL. The lower limit of detection (LOD) was 10 ng/mL at a signal-to-noise ratio of 4. The lower limit of quantitation (LOQ) was 45 ng/mL. The relative standard values for intra- and interday precision were <0.46 and <0.57%, respectively. The recovery of the drug samples ranged between 98.89 and 99.85%. No chromatographic interference from the tablet excipients was found. The proposed method was validated in terms of precision, robustness, recovery, LOD, and LOQ. All the validation parameters were within the acceptance range. The proposed method was applied for the determination of Tia in commercially available tablets. The results were compared with those obtained by an ultraviolet spectrophotometric method using t- and F-tests.     

Development of an HPLC method for the determination of salidroside in beagle dog plasma after administration of salidroside injection: application to a pharmacokinetics study

A simple RP-HPLC method was established for the determination of salidroside in dog plasma. Salidroside is one of the most active ingredients of Rhodiola L. The method had within-run precision values in the range of +/- 2.3 to +/- 9.1% (n = 5) and between-run precision in the range of +/- 3.2 to +/- 9.8%. A simple protein precipitation for salidroside extraction was processed using ACN at precipitant-to-plasma volume ratio (P-P ratio) of 3:2. The extraction recoveries of salidroside at seven concentrations were higher than 63.2%. There was a linear relationship between chromatographic area and concentration over the range of 0.83-520 microg/mL for salidroside in plasma (R = 0.9926). The LOQ (S/N = 10) of the method was 0.83 microg/mL. The method was applied in a study of the pharmacokinetics of salidroside injection in six beagle dogs. The major pharmacokinetic parameters of C(max), AUC(0-24), AUC(0-infinity), and t(1/2) of salidroside in beagle dogs after i.v. administration of a single 75 mg/kg (5 mL/kg) dose were 96.16 +/- 8.59 microg/mL, 180.3 +/- 30.6 microg h/mL, 189.3 +/- 32.1 microg h/mL, and 2.006 +/- 0.615 h, respectively.     

Determination of aripiprazole in rat plasma and brain using ultra-performance liquid chromatography/electrospray ionization tandem mass spectrometry

Aripiprazole is an important antipsychotic drug. A simple, sensitive and rapid ultra‐performance liquid chromatography/electrospray ionization tandem mass spectrometry (UPLC‐ESI‐MS/MS) method was developed and validated for the simultaneous quantification of this compound in rat plasma and brain homogenate. The analyte was extracted from rat plasma and brain homogenate using a weak cation exchange mixed‐mode resin‐based solid phase extraction. The compound was separated on an Agilent Eclipse Plus C₁₈ (2.1 × 50 mm, 1.8 µm) column using a mobile phase of (A) 0.1% formic acid aqueous and (B) acetonitrile with gradient elution. The analyte was detected in positive ion mode using multiple reaction monitoring. The method was validated and the specificity, linearity, limit of quantitation (LOQ), precision, accuracy, recoveries and stability were determined. The LOQ was 0.5 ng/mL for aripiprazole in plasma and 1.5 ng/g in brain tissue. The MS response was linear over the concentration range 0.5–100 ng/mL for aripiprazole in plasma and 1.5–300 ng/g in brain tissue. The precision and accuracy for intra‐day and inter‐day were better than 14%. The relative and absolute recoveries were above 72% and the matrix effects were low. This validated method was successfully used to quantify the rat plasma and brain tissue concentrations of the analyte following chronic treatment with aripiprazole. Copyright © 2012 John Wiley & Sons, Ltd.     

Determination of picroside II in dog plasma by HPLC and its application in a pharmacokinetics study

A sensitive and simple high-performance liquid chromatography method with UV detection was developed and validated for determining picroside II in dog plasma. Paeoniflorin was employed as internal standard and the sample pre-treatment procedure consists of deproteinization by addition of acetonitrile. Chromatographic separations were performed on a Shimadzu VP-ODS column (250 x 4.6 mm i.d., 5 microm). The mobile phase consisted of acetonitrile-0.1% acetic acid aqueous (v/v), 23:77, v/v, at a rate of 1 mL/min. Detection was carried out at a wavelength of 266 nm. Calibration standards ranged from 0.25 to 500 microg/mL in dog plasma and the mean correlation coefficient of 0.9981 was found for the linear calibration curves (n = 6). The limit of quantification (LOQ) was 0.25 microg/mL. Intra- and inter-assay RSD ranged from 0.70 to 7.5%. Accuracy (%bias) ranged from -6.3 to 6.0%. This method was applied to the pharmacokinetic study of picroside II in dogs. The study demonstrated the plasma picroside II concentration-time curves were fitted to the two-compartment open model and showed linear pharmacokinetics.     

Ultraviolet-photodiode array and high-performance liquid chromatographic/mass spectrometric studies on forced degradation behavior of glibenclamide and development of a validated stability-indicating method

A forced degradation study on glibenclamide was performed under conditions of hydrolysis, oxidation, dry heat, and photolysis and a high-performance column liquid chromatographic-ultraviolet (HPLC-UV) method was developed to study degradation behavior of the drug under the forced conditions. The degradation products formed under different forced conditions were characterized through isolation and subsequent infrared/nuclear magnetic resonance/mass spectral analyses, or through HPLC/mass spectrometric (HPLC/MS) studies. The drug degraded in 0.1 M HCI and water at 85 degrees C to a major degradation product, 5-chloro-2-methoxy-N-2-(4-sulfamoylphenyl)ethyl]benzamide (III), and to a minor product, 1-cyclohexyl-3-[[4-(2-aminoethyl)-phenyl]sulfonyl]urea (IV). Upon prolonged heating in the acid, the minor product IV disappeared, resulting in formation of 5-chloro-2-methoxy-benzoic acid (II) and an unidentified product (I). Heating of the drug in 0.1 M NaOH at 85 degrees C yielded II and IV as the major products and I and III as the minor products. The drug and the degradation products formed under different conditions were optimally resolved on a C18 column using ammonium acetate buffer (0.025 M, pH 3.5)-acetonitrile (45 + 55) mobile phase at a flow rate of 0.6 mL/min, with detection at 230 nm. The method was validated for linearity, precision, accuracy, and specificity. Limit of detection (LOD) and limit of quantitation (LOQ) values were also determined. The method could be successfully applied for simultaneous quantification of glibenclamide and the major product, III. The response of the method was linear in a narrow [0.4-10 micro/mL, correlation coefficient (r2) = 0.9982] and a wide (0.4-500 microg/mL, r2 = 0.9993) concentration range for glibenclamide, and in the concentration range of 0.025-50 microg/mL (r2 = 0.9998) for III. The method proved to be precise and accurate for both glibenclamide and III. It was specific for the drug and also selective for each degradation product, and LOQ values for the drug were 0.1 and 0.4 microg/mL, whereas those for III were 0.010 and 0.025 microg/mL, respectively.     

Rapid determination of gemcitabine in plasma and serum using reversed-phase HPLC

Gemcitabine (2'2'-difluorodeoxycytidine) is a pyrimidine analog used in the treatment of a variety of solid tumors. After intravenous (i.v.) administration, it is rapidly inactivated to 2'-deoxy-2',2'-difluorouridine (dFdU). A sensitive analytical method for the quantitation of gemcitabine is required for the assessment of alternative dosage and treatment schemes. A rapid and robust RP-HPLC assay for analysis of gemcitabine in human and animal plasma and serum was developed and validated using 2'-deoxyuridine (dU) and 5-fluoro-2'-deoxyuridine (5FdU) as internal standards. It is based on protein precipitation, the use of an Atlantis dC18 column of 100 mm length (inner diameter, 4.6 mm; particle size, 3 microm) and isocratic elution using a 10 mM phosphate buffer, pH 3.0, followed by isocratic elution with the same buffer containing 3% of ACN. For gemcitabine, RSD values for intraday and interday precision were < 4.4 and 5.3%, respectively, the LOQ was 20 ng/mL, and the assay was linear in the range of 0.020-20 microg/mL with an accuracy of > or =89%. The recovery for gemcitabine, dU and 5FdU was 86-98%. The assay was applied to determine gemcitabine levels in plasma samples of patients collected during and shortly after conventional infusion of 25-30 mg/kg body mass (levels: 2.0-18.9 microg/mL) and rats that received lower doses (1.5 mg/kg) via i.v., subcutaneous and oral drug administration (levels: 0.20-2.60 microg/mL). It could also be applied to estimate dFdU levels in human plasma.     

UV spectrophotometric determination of bioflavonoids from a semisolid pharmaceutical dosage form containing Trichilia catigua Adr. Juss and Ptychopetalum olacoides Bentham standardized extract: analytical method validation and statistical procedures

A precise, accurate, and sensitive UV spectrophotometric method was developed and validated for routine quantification of total bioflavonoids, expressed as rutin, from a topical oil-in-water pharmaceutical emulsion containing the extract of Trichilia catigua Adr. Juss and Ptychopetalum olacoides Bentham. The method was validated experimentally, and the data were treated rigorously by statistical analysis. The following analytical parameters were assessed: linearity, specificity, intra- and interrun precision measured as relative standard deviation (RSD, %), intra- and interrun accuracy (E, %), recovery (Rec., %), limit of detection (LOD, microg/mL), and limit of quantification (LOQ, microg/mL). The UV spectrophotometric method was linear (r = 0.9995) for standard rutin over the concentration range of 5.0-15.0 microg/mL with specificity for total bioflavonoids (expressed as rutin) at 361.0 nm with an absence of interferents from the complex matrix; RSD of < or = 1.79%, intrarun (E = 97.88 +/- 1.75 to 99.0 +/- 0.33%) and interrun (E = 98.38 +/- 1.12 to 100.79 +/- 1.30%) accuracy; Rec. = 98.64 +/- 0.42 to 100.74 +/- 0.41%; LOD = 0.20 microg/mL; and LOQ = 0.30 microg/mL.     

Determination of Rhein in blood plasma by HPLC with UV detection and its application to the study of bioequivalence

A method is proposed for determining rhein (an active metabolite of diacerein) in human blood plasma by reversed-phase high-performance liquid chromatography with UV detection. The analyte was selectively extracted from blood plasma by liquid-liquid extraction with a 5% solution of tert-butyl methyl ether in ethyl acetate. The recovery of rhein is 68 ± 3%, LOQ 100 ng/mL. The stability of the analytes and linearity, selectivity, sensitivity, reproducibility, and accuracy of the developed method are evaluated. The pharmacokinetics of diacerein in the oral administration of drug formulations by 18 healthy volunteers is studied.