0000000000073366
AUTHOR
Adela Sevillano-cabeza
Sensitive determination of probenecid in urine samples by reversed-phase liquid chromatography and UV-visible detection using solid-phase extraction techniques for sample clean-up
This study describes a rapid method for the determination of probenecid in human urine by liquid chromatography with UV detection at 254 nm, after clean-up through a C8 solid-phase extraction column. Liquid chromatography was carried out on a C18-bonded phase using an acetonitrile-acetate buffer (pH=4) gradient elution. Ethacrynic acid was used as internal standard. The system has been applied to the determination of probenecid in the 0.10–100.0 μg/ml concentration range; the limit of detection was 5 ng/mL.
New micromethod combining miniaturized matrix solid-phase dispersion and in-tube in-valve solid-phase microextraction for estimating polycyclic aromatic hydrocarbons in bivalves.
Abstract Miniaturized matrix solid-phase dispersion (MSPD) was developed for the extraction of common polycyclic aromatic hydrocarbons (PAHs) from bivalve samples (100 mg, dry weight). Additional clean-up and analyte enrichment was accomplished by in-tube solid-phase microextraction (SPME). For this purpose the extracts collected after MSPD were diluted with water and injected into a capillary column coated with the extractive phase. This capillary column was connected to the analytical column by means of a switching valve. Separation and quantification of the PAHs were carried out using a monolithic LC column and fluorescence detection. Since the in-tube SPME device allowed the processing …
Estimation of diuretic drugs in biological fluids by HPLC
This critical review of different methods proposed for the determination and screening of diuretics is directed mainly, because of its potential application, towards highperformance liquid chromatography.
Determination of theophylline and paraxanthine in urine samples by liquid chromatography using the H-point standard additions method
Abstract The simultaneous determination of theophylline and paraxanthine in urine samples by the H-point standard additions method (HPSAM) is described. Samples are extracted with C 18 solid-phase extraction cartridges and chromatographed using a Hypersil C 18 -ODS column and a mobile phase consisting of acetonirile-phosphate buffer in the gradient elution mode. Under these conditions theophylline and paraxanthine are eluted with short retention times. Although their chromatographic peaks are overlapped and their spectra are very similar, the H-point standard additions method provides excellent results in the determination of both xanthines at therapeutic levels.
o-Phthalaldehyde–N-acetylcysteine polyamine derivatives: formation and stability in solution and in C18 supports
A comparative study of different derivatization procedures has been performed in order to improve the stability of the reaction products o-phthalaldehyde-N-acetylcysteine (OPA-NAC) polyamines. Procedures such as solution derivatization, solution derivatization followed by retention on a packing support, derivatization on different packing supports and on-column derivatization, have been optimized and compared. The degradation rate constant (k) of the derivative was dependent on the procedure used and on the analyte. For the spermine (the most unstable isoindol tested) k was 8 +/- 2 x 10(-2) min(-1) in solution versus 7.7 +/- 1.1 x 10(-4) min(-1) on the (C18) solid support. The results obtai…
A comparison of various calibration techniques applied to the ninhydrin-cefoxitin determination
Abstract This report deals with spectrophotometric determination of cefoxitin from intact antibiotics or from its acid induced degradation product by means of their reaction with ninhydrin in a strong sulphuric acid medium. To obtain the calibration graphs four methods were used: absorbance of reacted cefoxitin against a reagent blank as an analytical signal, first-derivative signal of absorbance against a water blank as an analytical signal, first-derivative signal of absorbance against a reagent blank as an analytical signal and the H-Point Standard Additions Method (HPSAM) by using absorbance increment values as analytical signals. The HPSAM is applied to the determination of cefoxitin i…
On-line trace enrichment for the determination of ethacrynic acid in urine by liquid chromatography and column-switching
Abstract A chromatographic method based on liquid chromatography (LC) using column-switching for the determination of ethacrynic acid in urine samples, is described. The proposed system uses a Hypersil ODS-C18, 30 μm (20 mm × 2.1 mm i.d.) pre-column for the pre-concentration and separation of ethacrynic acid from the biological matrix. Polar urinary compounds are removed by flushing the pre-column with purified water, and the enriched analyte is then switched in back-flush mode onto an HP-LiChrospher RP C18, 5 μm (125 × 4 mm i.d.) analytical column, where it is chromatographed using an acetonitrile-acetate buffer gradient elution. The UV detector was set at 275 nm. The recovery of drug was …
Comparative Study on the Determination of Cephalexin in its Dosage Forms by Spectrophotometry and HPLC with UV-vis Detection
This paper discusses the spectrophotometric determination of cephalexin as the intact cephalexin or as its acid-induced degradation product. Cephalexin can be determined in the range 1 × 10−5−18 × 10−5M with relative standard deviations of 5-1%. The limits of quantitation and detection were 10−5 and 0.3 × 10−5M, respectively. These procedures were compared with reversed-phase HPLC determination. No interference was observed in the presence of common pharmaceutical adjuvants. The H-point standard additions method was applied in order to correct for the possible presence of the cephalexin precursor, 7-aminocephalosporanic acid; this improves the selectivity of the UV-vis spectrophotometric me…
New spectrophotometric procedure for determining cefotaxime based on derivatization with 1,2-naphthoquinone-4-sulphonate into solid-phase extraction cartridges--application to pharmaceutical and urine samples.
Cefotaxime was derivatised with 1,2-naphthoquinone-4-sulphonate (NQS), extracted into solid-phase cartridges (C18) and detected using a UV-visible detection system. Optimum conditions for this new procedure were: hydrogencarbonate-carbonate buffer, pH 10.5, 5-min reaction time at 25 degrees C and an NQS concentration of 7.1x10(-3) mol l(-1). The accuracy and the precision of the liquid-solid procedure were tested. The procedure was used to measure cefotaxime in pharmaceutical and urine samples. The results obtained were contrasted with those reported for a HPLC method for urine samples. The generalized H-point standard additions method was used to measure cefotaxime in urine samples.
Development of the H-point standard additions method for coupled liquid chromatography and UV-visible spectrophotometry
Abstract This work establishes the fundamentals of the H-point standard additions method for liquid chromatography for the simultaneous analysis of binary mixtures with overlapped chromatographic peaks. The method was compared with the deconvolution method of peak suppression and the second derivative of elution profiles. Different mixtures of diuretics were satisfactorily resolved.
Sensitive determination of ethacrynic acid in urine samples by reversed-phase liquid chromatography with ultraviolet detection using solid-phase extraction techniques for sample clean-up
Abstract A rapid method is described for the identification and determination of ethacrynic acid in human urine samples by liquid chromatography with UV detection, the retention time of the analyte being 4.9 min. The samples were previously conditioned using C 18 solid-phase extraction columns and chromatographed on an HP-LiChrospher RP C 18 column (5 μm; 125 mm × 4 mm i.d.) with gradient elution with acetonitrile-acetate buffer (pH 4). The detector was set at 275 nm and furosemide was used as an internal standard. The procedure was applied to the determination of ethacrynic acid at concentrations of 0.01-10 μg ml -1 and the limit of detection was 6 ng ml -1 .
Liquid Chromatographic Analysis of Amphetamine and Related Compounds in Urine Using Solid-Phase Extraction and 3,5-Dinitrobenzoyl Chloride for Derivatization
A chromatographic method for the analysis of amphetamine and related compounds in urine using 3,5-dinitrobenzoyl chloride (3,5-DNB) as a labeling reagent is presented. This assay is based on the employment of solid-phase extraction (SPE) cartridges for sample cleanup and derivatization. Experimental conditions are optimized for the simultaneous derivatization of ephedrine, norephedrine, pseudoephedrine, beta-phenylethylamine, amphetamine, methamphetamine, and 3-phenylpropylamine. The derivatives formed are separated in a LiChrospher 1000 RP18 (125 x 4-mm i.d., 5-microns film thickness) analytical column using a water-acetonitrile gradient elution and detected at 254 nm. Derivatization in C1…
Urine polyamines determination using dansyl chloride derivatization in solid-phase extraction cartridges and HPLC
The derivatization of biogenic amines such as putrescine, cadaverine, spermidine and spermine with dansyl chloride in solid phase extraction cartridges is described. Different types of filling materials were tested in order to have the highest retention of the different analytes. The best results were obtained by using C18 cartridges. The optimal conditions were: amine solution buffered at pH 12, 2 mM dansyl chloride (acetone-bicarbonate solution 20 mM (pH 9-9.5), 2 + 3 v/v) as reagent concentration, room temperature and 30 min reaction time. The developed procedure was applied to the determination of these polyamines in urine samples from healthy controls and cancer patients using HPLC wit…
In-tube solid-phase microextraction coupled by in valve mode to capillary LC-DAD: Improving detectability to multiresidue organic pollutants analysis in several whole waters
Abstract A simple and fast capillary chromatographic method has been developed to identify and quantify organic pollutants at sub-ppb levels in real water samples. The major groups of pesticides (organic halogens, organic phosphorous, and organic nitrogen compounds), some hydrocarbons (polycyclic aromatic hydrocarbons), phthalates and some phenols such as phenol and bisphenol A (endocrine disruptors) were included in this study. The procedure was based on coupling, in-tube solid-phase microextraction (IT-SPME) by using a conventional GC capillary column (95% methyl–5% phenyl substituted backbone, 80 cm × 0.32 mm i.d., 3 μm film thickness) in the injection valve to capillary liquid chromatog…
Improved Solid Phase Extraction Procedure for Assay of Cephalosporins in Human Urine Samples
Abstract Solid phase extraction technique has been evaluated for the treatment of urine samples in the analysis of cephalosporins before injection into an HP-Hypersil ODS-C18 column. Cephalexin, cefotaxime, cefazolin, cefuroxime, and cefoxitin were tested with seven different reversed-phase extraction column cartridges and the obtained urine extracts were not clean. However, 3M Empore extraction disk cartridges packed with octadecyl (C18) bonded silica provided clean extracts with a single extraction. The recoveries of the five cephalosporins ranged from 56 to 60 % in the 1.25–500 g/mL concentration range. The assay was accurate, precise, and adequate for testing the drug content in urine s…
On-Line Derivatization into Precolumns for the Determination of Drugs by Liquid Chromatography and Column Switching: Determination of Amphetamines in Urine
A chromatographic system for the on-line derivatization of drugs using column switching is described. The system uses a 20 mm × 2.1 mm i.d. precolumn packed with a unmodified ODS stationary phase. This column is used for sample cleanup and enrichment of the analytes. Next, the trapped analytes are derivatized by injection of the derivatization reagent into the precolumn. Finally, the derivatives are transferred to the analytical column for their separation under reversed-phase conditions. The influence of several parameters such as the reaction time, the amount of derivatization reagent, or the system design has been studied using some amphetamines as model compounds and three derivatizatio…
Automated pre-column derivatization of amines in biological samples with dansyl chloride and with or without post-column chemiluminescence formation by using TCPO-H2O2.
On-line automation of two different liquid chromatographic procedures, a pre-column derivatization system and a pre- and post-column system, in order to generate chemiluminescence is reported. Dansyl chloride (Dns-Cl) was used as a pre-column reagent to form fluorophores and bis(2,4,6-trichlorophenyl) oxalate (TCPO) and hydrogen peroxide (H2O2) as a post-column reagent to generate chemiluminescence. This procedure is based on the employment of a primary column packed with C18 material inserted in a multi-dimensional assembly for sample clean-up and derivatization with Dns-Cl. The dansyl derivatives formed are transferred and separated in a LiChrospher 100 RP18 analytical column (125 x 4 mm …
Column-switching techniques for screening of diuretics and probenecid in urine samples
A method based on high-performance liquid chromatography using column-switching is described for the screening of diuretics and probenecid in urine samples. The system uses a 20- x 2.1-mm i.d. precolumn, packed with a Hypersil ODS-C18, 30-microns stationary phase, for the on-line sample cleanup and enrichment. Untreated urine samples are directly injected, and the precolumn is flushed for 1 min with water to eliminate polar matrix components. The retained analytes are then back-flushed by means of a six-port switching valve onto a Hypersil ODS-C18 analytical column (5 microns, 250- x 4-mm i.d.), where they are separated using an acetonitrile/phosphate buffer (pH = 3) gradient elution. Under…
Determination of acetazolamide in human urine samples by reversed-phase high-performance liquid chromatography in the presence of xanthines.
A simple, rapid and selective high-performance liquid chromatographic assay for the determination of acetazolamide in urine samples is described. After extraction with ethyl acetate, the drug is chromatographed on an HP-Hypersil ODS-C18 column with a mobile phase of acetonitrile-phosphate buffer (pH 3) and ultraviolet detection at 275 nm. The efficiency of the extraction, the linearity and the reproducibility of the method permit the evaluation of acetazolamide urinary excretion a long time after its administration.
A new derivatization procedure for the determination of cephalexin with 1,2-naphthoquinone 4-sulphonate in pharmaceutical and urine samples using solid-phase extraction cartridges and UV–visible detection
Abstract The present report shows how to derivatize cephalexin with 1,2-naphthoquinone-4-sulphonate (NQS) into solid-phase extraction cartridges (C18) using UV–visible detection. Optimum conditions for this new procedure are: hydrogen carbonate/carbonate buffer pH=10.5, 5 min reaction time at 25°C and NQS concentration of 7.1×10−3 mol l−1. The accuracy and the precision of the method were tested. The procedure was used to measure cephalexin in pharmaceutical and urine samples. The results obtained were contrasted with those reported by UV-spectrophotometric and HPLC methods for pharmaceutical samples and with HPLC method for urine samples. The H-point standard additions method was used to m…
Solid-Phase Extraction and Clean-Up Procedures in Pharmaceutical Analysis
Solid-phase extraction (SPE) using small cartridges filled with sorbents of a small particle size has rapidly established itself as an important sample clean-up technique. It has prospered at the expense of liquid-liquid extraction (LLE), which is considered labor intensive and frequently plagued by problems, such as emulsion formation and use of large volumes of hazardous solvents. A remarkable characteristic of SPE is its easy adaptation to on-line mode by column-switching techniques; switching can be effected manually or by automated controllers. The same analyte/sorbent interactions that are exploited in liquid chromatography (LC) are of use in SPE, but particle sizes employed are great…
Derivatization of amines in solid-phase extraction supports with 9-fluorenylmethyl chloroformate for liquid chromatography
Abstract A new method based on reaction in solid-phase extraction cartridges with 9-fluorenylmethyl chloroformate is described for improved derivatization of amines prior to liquid chromatography. The effects of the reaction conditions, the kinetics of the reaction and the kind of packing material on the derivatization have been evaluated. The reliability of the described approach has been tested by derivatizing propylamine, diethylamine, diethanolamine, glycine, glutamic acid, p-aminobenzoic acid and β-phenylethylamine. Reaction yields compared with those of the analogous solution derivatizations were in the 44–169% range, and satisfactory linearity was achieved at concentrations of the am…
Amphetamine and Methamphetamine Determinations in Biological Samples by High Performance Liquid Chromatography. A Review
Abstract This critical review shows the different high performance liquid chromatography methods proposed for amphetamine and methamphetamine determinations. It is directed mainly towards sample clean up and derivatizations steps, because of their significance in such determinations.
Column-switching techniques for high-performance liquid chromatography of drugs in biological samples.
In recent years, an increasing number of publications have demonstrated the potential of column-switching techniques for the chromatographic separation, determination and preparative isolation of analytes from biological matrices. Column-switching systems greatly facilitate drug analysis, by on-line sample clean-up and trace enrichment, or by improving the analytical separative process. In this paper, the main applications of column-switching techniques to drug analysis in biological samples, are reviewed.
Solid-Phase Extraction Techniques for Assay of Diuretics in Human Urine Samples
Abstract Solid-phase extraction techniques were evaluated for the treatment of urine samples in the analysis of diuretics before injection into an HP-Hypersyl ODS-C18 column. Six different reversed-phase extraction columns were tested, and the results obtained are compared with those obtained in a classical liquid-liquid extraction with ethyl acetate. The solid-phase extraction procedures are the best overall choice for all the diuretics tested, due to their versatility, the minor time-consuming, and the good recovery percentages obtained. C18 and C8 packings give the highest recoveries for a majority of the diuretics studied. However, CH or PH columns, due to their greater selectivity, can…
Determination of triamterene in urine by HPLC using fluorescence detection and column-switching
A liquid chromatographic method incorporating column-switching and fluorimetric detection for the determination of triamterene in untreated urine, is described. The urine samples (5 μL) were directly introduced onto an Hypersil ODS-C18, 30 μm (20 mm×2.1 mm I.D.) pre-column. Polar urinary compounds were removed by flushing the pre-column with water for 1 min, and the analyte was then switched onto an HP-LiChrospher RP C18,5 μm (125 mm×4mm ID) analytical column using an acetonitrile/phosphate buffer gradient elution. Fluorescence detection was performed at 230 nm excitation and 430 nm emission wavelengths. The recovery of drug was 102±2% in the 0.10–20.0 μg/mL concentration range, the limit o…
Extractive-spectrophotometric determination of furosemide with sodium 1,2-naphthoquinone-4-sulphonate in pharmaceutical formulations
ABSTRACT An extractive-spectrophotometric study based on the reaction of furosemide with 1,2-naphthoquinone-4-sulphonate (NQS) is described. Optimal conditions are: NaH2PO4-Na2HPO4 pH 7.5, NQS 7.7×...