Liquid chromatographic determination of azidothymidine in human plasma using on-line dialysis and preconcentration on a silver(I)-thiol stationary phase

An automated method is described for the routine determination of 3′-azido-2′,3′-dideoxythymidine (AZT), the best known drug against acquired immunodeficiency syndrome (AIDS). The method is based on on-line dialysis to remove matrix macromolecules, followed by selective preconcentration and clean-up with a silver(I)-thiol stationary phase. After desorption of the solute with a small plug of perchloric acid, chromatography is applied using an octadecyl-modified silica column. Using UV absorbance detection at 269 nm, the minimum detectable concentration in plasma is 20 ng ml^?1 (600-μl sample). The within-day reproducibility at the 20 ng ml^?1 level is 4.4% and at least 128 samples can be analysed unattendedly without exchanging the dialysis membrane, the precolumn or the analytical column.     

Simultaneous determination of inulin and p-aminohippuric acid (PAH) in human plasma and urine by high-performance liquid chromatography

Inulin and p-aminohippuric acid (PAH) clearances are used for the estimation of glomerular filtration rate (GFR) and effective renal plasma flow (ERPF). A simple and rapid high-performance liquid chromatography (HPLC) method with UV detection is described for the simultaneous determination of inulin and PAH in the same chromatogram in the plasma and urine of humans. Plasma and urine samples were hydrolyzed with perchloric acid (0.7%) in boiling water. The mobile phase consisted of 0.01 M potassium dihydrogenphosphate with 0.02 M tetramethylammonium chloride and o-phosphoric acid (pH 3)-acetonitrile (94:6, v/v), pumped at a rate of 1.2 ml min-1 on a C8 reversed-phase column. Tannic acid was used as the internal standard and UV detection at 285 nm was employed. The calibration curves were linear over the concentration range of 12.5-100 mg l-1 for inulin and 6.25-50 mg l-1 for PAH with determination coefficients greater than 0.997. The method is accurate (bias < 13%) and reproducible (intra- and inter-day relative standard deviation less than 11%), with a limit of quantitation of 12.5 mg l-1 and 6.25 mg l-1 for inulin and PAH, respectively. Analytical recoveries from urine and plasma were ranged from 81 to 108% for both compounds. This fully validated method, which allows the simultaneous determination of inulin and PAH clearances, is simple, rapid (total run time < 10 min) and requires only a 200 microliters plasma or urine sample.     

Simple high-performance liquid chromatographic method for the determination of captopril in biological fluids.

A rapid, simple and sensitive column-switching high-performance liquid chromatographic procedure for the determination of captopril in plasma and urine had been developed. p-Bromophenacyl bromide was used as a derivatizing reagent to react with captopril to form a product that showed ultraviolet-absorbing properties. For plasma samples the protein was removed with 6% perchloric acid before injection. The urine samples were directly injected into the chromatograph. The column-switching system was equipped with a pre-column (5 cm x 0.5 cm I.D.) packed with muBondapak C18 (37-50 microns) and an analytical column (15 cm x 0.5 cm I.D.) packed with YWG-C18, 10 microns. Impurities were washed from the pre-column with 0.2% acetic acid and the retained substances were eluted into the analytical column with acetonitrile-water-acetic acid (35:65:0.4, v/v). Captopril was detected at 260 nm. The calibration curve was linear in the range 20-1000 ng/ml for plasma and 10-200 micrograms/ml for urine. The recoveries averaged 103.2 and 99.5% for plasma and urine, respectively. The coefficients of variation were all less than 10%.     

Quantitation of agmatine by liquid chromatography with laser-induced fluorescence detection

A high performance liquid chromatography (HPLC) method is described for the determination of agmatine, an endogenous neuromodulator. The method involves pre-column derivatization of the sample with a fluorescent tagging reagent, 7-fluoro-4-nitrobenzoxadiazole (NBD-F). The resulting agmatine derivative is stable and can be readily extracted into ethyl acetate at pH 8.5. The extraction enhances the quantification of low level agmatine because it eliminates chromatographic peaks caused by endogenous amino acids. The HPLC separation is carried out on a C₈ reversed phase column and completed in less than 10 min. With laser-induced fluorescence (LIF) detection, the detection limit is 5×10⁻⁹ M agmatine. Method precision (coefficient of variation) is 5% for agmatine in human plasma at the sub-μM level. This method has been validated by determination of agmatine in biological samples including human plasma and rat brain and stomach tissues.     

Simultaneous capillary electrophoretic separation and detection of P(V) and As(V) As heteropoly-blue complexes

A capillary electrophoretic (CE) method was developed for the simultaneous determination of P(V) and As(V). A Mo(VI)-ascorbic acid reagent reacted with a mixture of trace amounts of P(V) and As(V) to form the corresponding heteropoly-blue complexes in 0.05 M acetate buffer (pH 3.5). When 0.05 M malonate buffer was used as a migration buffer, the peaks due to their migrations were well separated in the electropherogram, and the pre-column complex-formation reaction was applied to the simultaneous CE determination of P(V) and As(V) with direct UV detection at 220 nm. With the proposed method, the calibration curves were linear in the concentration range of 5 x 10(-7) - 1 x 10(-4) M, with a detection limit of 1 x 10(-7) M (a signal-to-noise ratio of 3). Interference from foreign ions was also discussed.     

High-performance liquid chromatographic determination of morphine and its 3- and 6-glucuronide metabolites: improvements to the method and application to stability studies

Improvements to previously reported methods for the determination of morphine, morphine-3-glucuronide (M3G) and morphine-6-glucuronide (M6G) in human plasma are described. The improved methods involve the use of a solid-phase extraction cartridge and a chromatographic system which uses paired-ion reversed-phase high-performance liquid chromatography with a radially compressed column. Only one cartridge is used to prepare each sample for chromatography and each cartridge may be used for at least fourteen 1-ml plasma samples. The recovery is greater than 85%. The improvements to the method of sample pretreatment and in the chromatographic conditions have allowed determination of morphine, M3G and M6G in human plasma down to 13.3 nmol/l (coefficient of variation = 9.3%), 108 nmol/l (6.6%) and 41 nmol/l (6.7%), respectively, using ultraviolet detection alone. It was shown that all three compounds were stable in plasma for up to 101 weeks when stored at -20 degrees C.     

Determination of endogenous 3-hydroxykynurenine in plasma

A method is described which permits determination of 3-hydroxykynurenine (l-3-(2-amino-3-hydroxybenzoyl)-alanine), a metabolite of tryptophan, in plasma with a limit of detection of less than 0.1 pmol. This compound can be extracted by means of adsorption onto a strong anion-exchange resin at pH 9.5 in the presence of an antioxidant. Desorption is achieved by shaking with 2 M perchloric acid. Plasma samples are deproteinized and prepurified by column anion-exchange chromatography in order to eliminate the uric acid present. The compound is then determined by reverse-phase HPLC with amperometric detection. Recovery determinations may be made by addition of an internal standard. Concentrations in rat plasma were found to be of the order 85 nM, which appears to be a value much lower than reported earlier.     

Determination of Azidothymidine – an Antiproliferative and Virostatic Drug by Square‐Wave Voltammetry

The 3′‐azido‐3′‐deoxythymidine (AZT, Zidovudine) is an antiproliferative and virostatic drug widely used in human immunodeficiency virus type 1 (HIV‐1) infection treatment. With respect to side effects of high doses and a short half‐life of AZT, a fast and simple detection method for this agent could be helpful. The aim of our study was to determine AZT levels in natural samples (urine, serum, whole blood, and cell cultures, such as the HaCaT line of keratinocytes) without their mineralization and/or purification, by means of electrochemical methods using hanging mercury drop electrode (HMDE). On this electrode, AZT undergoes irreversible reduction at the peak potential near E_p?1.1 V (vs. Ag/AgCl/3 M KCl). Reduction AZT signals were measured by cyclic voltammetry (CV), differential pulse voltammetry (DPV), square‐wave voltammetry (SWV), and constant current chronopotentiometric stripping analysis (CPSA). In phosphate buffer (pH 8) the SWV yielded the best AZT signal with the detection limit of 1 nM. The determination of AZT concentration in biological materials is affected by electroactive components, such as proteins and DNA. For monitoring the influence of these compounds, AZT reduction was performed in the presence of 10 μg/mL calf thymus ssDNA and/or 100 μg/mL bovine serum albumin. In these cases, the detection limit increased to 0.25 μM. Also studied was the AZT concentration in keratinocyte cells (HaCaT line) during cell cultivation. It has been shown that the SWV may be considered as a useful tool for the determination of AZT concentration in cell cultures, and for monitoring AZT pharmacokinetics.     

Direct determination of mitoxantrone and its mono- and dicarboxylic metabolites in plasma and urine by high-performance liquid chromatography

The simultaneous isolation and determination of mitoxantrone (Novantrone) and its two known metabolites (the mono- and dicarboxylic metabolites) were carried out using a high-performance liquid chromatographic (HPLC) system equipped with an automatic pre-column-switching system that permits drug analysis by direct injection of biological samples. Plasma or urine samples were injected directly on to an enrichment pre-column flushed with methanol-water (5:95, v/v) as the mobile phase. The maximum amount of endogenous water-soluble components was removed from biological samples within 9 min. Drugs specifically adsorbed on the pre-column were back-flushed on to an analytical column (Nucleosil C18, 250 X 4.6 mm I.D.) with 1.6 M ammonium formate buffer (pH 4.0) (2.5% formic acid) containing 20% acetonitrile. Detection was effected at 655 nm. Chromatographic analysis was performed within 12 min. The detection limit of the method was about 4 ng/ml for urine and 10 ng/ml for plasma samples. The precision ranged from 3 to 11% depending on the amount of compound studied. This technique was applied to the monitoring of mitoxantrone in plasma and to the quantification of the unchanged compound and its two metabolites in urine from patients receiving 14 mg/m2 of mitoxantrone by intravenous infusion for 10 min.     

Kinetic spectrophotometric determination of josamycin in formulations and spiked human plasma using 3-methylbenzothiazolin-2-one hydrazone/Fe+3 system

A simple kinetic spectrophotometric method was developed for the determination of josamycin in its dosage forms and spiked human plasma. The method is based on reaction of the drug with 3-methylbenzothiazolin-2-one hydrazone/ferric chloride system for a fixed time of 20 min at 70 degrees C and measuring the produced color at 665 nm. The absorbance-concentration plot is rectilinear over the range of 5.0-30.0 microg/mL with detection limit of 1.0 microg/mL (1.2 x 10(-6) M). The determination of josamycin by the fixed concentration and the rate-constant methods is also feasible with the calibration equations obtained, but the fixed-time method proved to be more applicable. The procedure was successfully applied to commercial tablets. The results obtained were favorably compared with those given by reference methods. The method was further extended to the in vitro determination of josamycin in spiked human plasma. The recovery (n = 8) was 100.76 +/- 3.43%. The stoichiometry of the reaction between the drug and the reagent was studied by adopting the limiting logarithmic method, and a proposal of the reaction pathway was presented.     

土壤中铅的分析方法比对

对不同的样品消解方法及电感耦合等离子体质谱、电感耦合等离子体原子发射光谱、石墨炉原子吸收光谱法测定土壤中铅的测定结果进行比对。采用电热板、微波及水浴3种加热方式,选择硝酸、氢氟酸、双氧水、王水、高氯酸、盐酸的不同组合进行土壤样品消解,通过分析测定值的精密度和准确度,考察消解体系对电感耦合等离子体质谱、电感耦合等离子体发射光谱、石墨炉原子吸收光谱法测定结果的影响。结果表明采用电感耦合等离子体质谱法测定土壤中的铅,最适宜的消解体系是硝酸-氢氟酸-高氯酸(微波加热),采用电感耦合等离子体原子发射光谱法测定最适宜的消解体系是硝酸(电热板加热),采用石墨炉原子吸收光谱法测定最适宜的消解体系是硝酸-盐酸-高氯酸(微波加热)。电感耦合等离子体质谱法的精密度和准确度优于另外两种方法。     

Determination of amifloxacin and two of its principal metabolites in plasma and urine by high-performance liquid chromatography using automated column switching

The automated determination of amifloxacin and two of its principal metabolites in human plasma and urine by column-switching high-performance liquid chromatography is described. Plasma or urine samples, diluted 1:1 with 0.5 M sodium citrate buffer pH 2.5, were directly injected onto a cation-exchange pre-column. Following a 2.0-min wash of the pre-column with water at a flow-rate of 1.1 ml/min, the effluent from the pre-column was directed to the analytical column by a column-switching device. The precision of the plasma and urine methods ranged from a +/- 1.9 to +/- 3.6% for all compounds. The accuracies of the methods were within a range of -3.3% to 6.4% of the nominal values for all compounds. Linear responses were observed for all the standards in the range 0.10-5.0 micrograms/ml for plasma and 0.50-100 micrograms/ml for urine for all three compounds. The minimum quantifiable levels were 0.10 and 0.50 micrograms/ml for plasma and urine, respectively. The analytical methods may be used to quantify amifloxacin and the piperazinyl-N-desmethyl and piperazinyl-N-oxide metabolites in plasma and urine samples obtained from humans, monkeys, dogs and rats.     

Determination of aluminum in environmental and biological samples by reversed-phase high-performance liquid chromatography via pre-column complexation with morin

Morin was used as a pre-column reagent for the determination of aluminum by RP-HPLC with fluorescence detection. This method has been successfully applied to direct determination of trace Al in environmental and biological samples. The response was linear from 6 x 10(-9) to 6 x 10(-5) M with a detection limit of 2 x 10(-9) M. In addition, the different Al complexes with morin were separated by the proposed HPLC procedure and their coordination ratios were depicted by molar-ratio method. The results showed that 1:1 and 2:1 Al-morin complexes formed.     

快速柱前衍生HPLC法检测曲格列汀中(R)-3-氨基哌啶

以邻苯二甲醛(OPA)和3-巯基丙酸为衍生试剂,建立了柱前衍生高效液相色谱(HPLC)测定曲格列汀中(R)-3-氨基哌啶含量的分析方法.(R)-3-氨基哌啶与衍生剂在碱性(pH 10.5)条件下于室温反应30s,进行柱前衍生,并利用高效液相色谱-质谱对衍生产物进行定性分析.采用YMC-Triart C18色谱柱(150...     

Simultaneous Determination of Platinum and Palladium by Second Derivative Spectrophotometry Using 3-(2'-Thiazolylazo)-2, 6-Diaminopyridine as Chromophore Ligand

ABSTRACT

A second derivative spectrophotometric method has been developed for the determination of palladium and platinum in mixtures. The method is based on the formation of the platinum and palladium complexes with 3-(2-thiazolylazo)-2, 6-diaminopyridine, (2, 6-TADAP), in the presence of 1.7 M perchloric acid solution, upon heating at 90° C for 30 min and on the subsequent direct derivative spectrophotometric measurement. The zero-crossing approach and the graphic method were used for determination of platinum and palladium, respectively. Each analyte was determinated in the presence of one another in the ranges 8.9×10^-7 -3.1×10^-5 M for platinum and 4.6×10^-7 - 6.8×10^-5 M, for palladium. The detection limits achieved (3a) were found to be 2.7×10^-7 M of platinum and 1.4×10^-7 M of palladium. The relative standard deviations were in all instances less than 1.0%. In this work is included a study of effect of interferents and the application of the proposed method in synthetic mixtures.     

Direct photometric titration of tellurium

A direct photometric titration method has been developed for the determination of tellurium. Tellurium(IV) is titrated with potassium dichromate in 2-6M nitric acid (or 1-4M perchloric acid) at 380-430 nm (blue-violet filter). Selenium(IV) does not interfere.     

N-(1-Naphthyl)ethylenediamine, a New UV Labeling Reagent Used for LC Determination of Valproic Acid in Human Plasma

A high performance liquid chromatography method is presented for the determination of valproic acid levels in human plasma. The method was based on pre-column derivatization using N-(1-naphthyl)ethylenediamine as a new labeling agent. The calibration curve was linear in the investigated concentration range between 0.1 and 100 μg mL^?1 and showed good accuracy and reproducibility. The assay provided a limit of quantification of 0.1 μg mL^?1 for valproic acid and a limit of detection of 10 ng mL^?1, respectively. The presented method was successfully applied to the determination of valproic acid levels in plasma after oral administration of 600 or 800 mg of sodium valproate.     

Direct determination of trace aluminum with quercetin by reversed-phase high performance liquid chromatography

The determination of trace levels of aluminum by high performance liquid chromatography (HPLC) with spectrophotometric detection using quercetin, a bioactive substance as a pre-column reagent, is developed in this paper. The Al-quercetin chelate was separated on a reversed-phase ODS column with a mobile phase consisting of 70% water (pH 1.0 with perchloric acid) and 30% methanol, and detected at its maximum of 415 nm. The response was linear over the 1.0×10⁻⁷ to 8.0×10⁻⁵ M concentration range with a detection limit of 5.0×10⁻⁸ M and a relative standard deviation of 1.0% at the 5×10⁻⁶ M level. The analysis was free from common ions except iron, which could be successfully screened by 1,10-phenanthroline. This method has been employed to the determination of Al in environmental and biological samples. Moreover, direct speciation of labile monomeric Al, the toxic form of Al, in natural water by the present technique was explored. The coordination ratio of Al complex with quercetin was also elucidated by HPLC combined with molar ratio method. Only a 1:1 complex was formed. Because quercetin exists in body, a preliminary thinking of in vivo determination of Al is provided in this study.     

Thallimetric oxidations-V Titrimetric and spectrophotometric determination of hydrogen peroxide

Titrimetric and spectrophotometric methods have been developed for the determination of hydrogen peroxide at mmole and mumole levels respectively. In these methods thallium(III) is used as the oxidant and the reduced thallium(I) is determined oxidimetrically with potassium bromate in the titrimetric method and by measuring the absorbance of thallium(III) at 260 nm in the presence of 0.1M hydrochloric acid and 1M perchloric acid in the spectrophotometric method. Photochemical redox methods for the estimation of hydrogen peroxide in the presence of a number of diverse ions are described.     

Determination of vigabatrin in human plasma and urine by high-performance liquid chromatography with UV-Vis detection

A simple and reliable high-performance liquid chromatographic (HPLC) method with UV-Vis detection has been developed and validated for the determination of vigabatrin (VG) in human plasma and urine. The samples were pre-column derivatizated with 1,2-naphthoquinone-4-sulphonic acid sodium salt (NQS). A good chromatographic separation was achieved on a C18 column with a mobile phase consisting of acetonitrile and 10 mM orthophosphoric acid (pH 2.5) gradient elution. Tranexamic acid was used as an internal standard (I.S.). The method was linear over the concentration range of 0.8-30.0 microg/ml for both samples. The method is precise (relative standard deviation (R.S.D.) <9.13%) and accurate (relative mean error (RME) <-8.75%); analytical recoveries were 81.07% for plasma and 83.05% for urine. The assay was applied to pharmacokinetic study in a healthy volunteer after a single oral administration of 1 g of vigabatrin.