Simultaneous Ion-Pairing Liquid Chromatographic Determination of the Major Metabolites of Styrene and Carbamazepine,and of Unchanged Carbamazepine in Urine

Abstract

A high-performance liquid chromatographic method for a simultaneous quantitative determination of carbamazepine (CBZ) and of the major metabolites of CBZ (trans-10,11-dihydroxy-10,11-dihydrocarbamazepine, TDC; carbamazepine-10,11-epoxide, CBZ-E) and those of styrene (S) (hippuric acid, HA; mandelic acid, MA; phenylglyoxylic acid, PA) in the rat urine is described. Separation is achieved on a Nova-Pak reverse-phase column by isocratic elution. Excellent resolution was obtained by adding to the acetonitrile-water mobile phase, tetrabutylammonium chloride (0.005 M) and methanol (1%). Detection is effected by UV absorption at 230 nm with a total analysis time of less than 18 min. An aliquot of diluted urine is injected directly onto the liquid chromatographic column. The limits of sensitivity of CBZ, CBZ-E, TDC, HA, MA, and PA are 3.3, 2.0, 1.8, 3.1, 1.2, and 3.1 μg/ml of diluted rat urine, respectively. Precision and accuracy of the method are found to be acceptable. The method can be used for studying the interaction between these two xenobiotics. Preliminary studies have shown its potential application to human investigations.     

Rapid Quantitative Method for the Determination of Dextrose,Mannitol, and Sorbitol in Meat Products by Liquid Chromatography

Abstract

A liquid chromatographic (LC) method for the determination of dextrose, mannitol, and sorbitol in meat products was developed. Dextrose, mannitol, and sorbitol were extracted from comminuted meat products with 52% ethanol. After filtration, the extracts were purified by passing through a C₁₈ Sep-Pak cartridge and two ion exchange resin Econo-Columns in series. After concentration and filtration, extracts were analyzed by liquid chromatography using a cation exchange analytical column and a differential refractometer detector. Homogeneously ground samples of cooked and fresh sausages and ground beef were fortified with dextrose, mannitol, and sorbitol at four different concentrations. Average overall recovery for all three compounds at all four levels of fortification was greater than 80% with coefficients of variation less than 10%.     

Simultaneous Determination of Carbamazepine,Phenobarbital and Their Major Metabolites in Serum,Brain Tissue and Urine by High-Performane Liquid Chromatographya

Abstract

A high-performance liquid chromatographic method is described for the simultaneous measurement of carbamazepine, phenobarbital and their major metabolites in small samples of serum, brain tissue and urine. This involves solvent extraction, reversed-phase chromatography and ultraviolet detection at 195 nm. Glucuronides in urine are hydrolyzed by enzymatic cleavage with β-glucuranidase. Quantitation is based on the peak-height ratio of the analyte to its internal standard (10, 11-dihydrocarbamaze-pine or p-methylphenobarbital). The results obtained show that the method is precise and reproducible.     

A Sensitive Flow Injection Fluorimetry for the Determination of Carbamazepine in Human Plasma

Abstract

A simple, sensitive, and specific flow injection fluorimetric method has been developed for the determination of carbamazepine (CBZ). The proposed method is based on use of a solid‐phase reactor containing lead dioxide for on‐line oxidization of CBZ into a strongly fluorescent compound in a medium of phosphoric acid. The product has a green‐yellow fluorescence at a maximum excitation wavelength of 355 nm and an emission wavelength of 478 nm. Under the optimum conditions, the fluorescence intensity is proportional to the concentration of CBZ ranging from 0.0005 to 4.000 µg mL^?1. The detection limit is 5.7×10^?5 µg mL^?1 (2.4×10^?10 mol L^?1) and the relative standard deviation is 1.4% at the sampling rate of 45 h^?1. The proposed method has been applied to clinical estimation of CBZ in real patients' plasma samples with the results compared with those obtained by HPLC method.     

高效液相色谱法同时测定血清中茶碱、苯妥英钠、苯巴比妥及卡马西平的药物浓度

报道了用高效液相色谱法（HPLC）同时测定血清中茶减、苯妥莫纳、苯巴比妥及卡马西平的药物浓度。实验条件:Nova-PahC18柱，流动相为甲醇-水（1:1，V／V），检测波长210nm，流速为1mL／min，萃取液为级访-异丙醇（95:5，V／V）。方法具有灵敏（10-9）、准确（回收率在97％～105％之间）、快速（7min）等特点，对临床血药浓度监测有实际应用价值。     

Serum Injection of the HPLC Column for Carbamazepine Assay

Abstract

Serum was injected directly on an HPLC column packed with C₁, 6.5 μm particle size, 300 Å pore-packing material for carbamazepine determination. The use of a wide-pore column with low hydrophobicity eliminated excessive pressure buildup in the column from protein precipitation which is usually caused by the high concentration of organic solvents in the mobile phase. This simple approach can be utilized for the determination of other drugs and endogenous compounds in serum.     

Application of PMR Spectrometry in Pharmaceutical Analysis III. Assay of Carbamazepine

Abstract

Carbamazepine, a tricyclic antidepressant, has been assayed in its tablets (Tegretol ®) by the application of PMR spectrometry. The proposed method involves comparing the integral of the aromatic and olefinic protons of carbamazepine, in the range 6.60 -7.60 ppm, to that of the singlet of a known amount of hexamethylcyclotrisilazane (at 0.00 ppm) used as internal standard. The method is simple, rapid and accurate. The average percent recovery, of carbamazepine from its tablets, is 96.88 ± 0.93. In addition, the PMR spectrum obtained helps in checking the identity and purity of the drug extracted from its pharmaceutical preparation.     

Gas Chromatographic Determination of Trimethylsilyl Derivatives of Free Amino Acids from a Botanical Source

Abstract

The application of gas chromatography for the separation of TMS-amino acids from a botanical source was demonstrated. The trimethyl-silyl derivatives of the extracts from germ free tobacco tissue cultures were prepared by reacting amino acid extracts with bis(trimethylsilyl)-trifluoroacetamide (BSTFA) using acetonitrile as a reaction solvent following preliminary separation of the free acids by ion exchange chromatography. Gas chromatographic separation was accomplished with a 10% OV-11 glass column and temperature programming. The findings compare favorably with other chromatographic methods. Structures of the TMS-amino acids were confirmed by gas chromatography-mass spectrometry combination. Mass spectral data for each derivative is presented for the principal protein amino acids observed as well as γ-aminobutyric acid, asparagine, α-aminobutyric acid and β-alanine.     

Extraction of Anthracyclines from Biological Fluids for HPLC Evaluation

Abstract

A new technique for the extraction of anthracyclines and their metabolites from plasma or serum is described, which gives suitable extracts for HPLC analysis of these drugs in patients. This technique consists in a very rapid chromatographic step on the bonded silica contained in small open columns (C-18 Sep-Paks, Waters Associates). This extraction gives quantitative recoveries of all the anthracyclines and metabolites that have been tested; 0.2 to 3 ml of plasma can be processed, with concentrations up to 5,000 ng/ml. The advantages of this technique over classical techniques using an organic extraction with a partition between two phases are: speed, simplicity, efficacy, reproducibility and similarity of recoveries for various anthracyclines.     

High Performance Liquid Chromatographic Determination of Triazolam in Human Serum

Abstract

A high performance liquid chromatographic method which utilizes UV-detection has been developed for the sensitive and specific determination of triazolam in human serum. Using 8-chloro-6-phenyl-l-ethoxymethyl-4H-s-triazolo[4, 3-a][1, 4]benzodiazepine as an internal standard, serum samples were buffered with 2 ml of 4M NaOH and extracted twice with 5 ml aliquots of toluene. The combined toluene extracts were evaporated to dryness and the residue dissolved in the chromatographic mobile phase. The samples were chromatography on a microparticulate reverse-phase column using a 0.06M acetic acid:acetonitrile (61:39) mobile phase. Known metabolites of triazolam did not interfere in the analysis. A linear relationship between peak height ratios and concentrations was observed, with the lower limit of detection being approximately 1 ng of triazolam. The utility of the method was demonstrated by administering therapeutic doses of the drug to human volunteers and monitoring serum triazolam concentrations as a function of time.     

Electron Capture Gas Chromatographic Assay for Miconazole and Clotrimazole In Skin Samples

Abstract

Electron capture gas chromatographic assay procedures were developed for quantitating the antifungal agents clotrimazole and miconazole. The procedures were specifically developed for analysis of the drugs in superficial samples of human skin. The analytical methods were sensitive to 5 ng of miconazole and 10 ng clotrimazole per tissue sample.     

High Performance Liquid Chromatography and Proteolytic Enzyme Characterization of Peptides in Tooth Pulp Extracts

Abstract

Metabolic profiles are obtained for peptides contained in tooth pulp extracts. To determine which high performance liquid chromatographic peaks are due to peptides, a series of proteolytic enzymes (chymotrypsin, trypsin, and carboxypeptidase A) are utilized. Results from treatment of extracts with immobilized enzymes demonstrate that virtually all peaks in this reverse phase system are due to peptides. This current study is a necessary component in a larger research program focusing on quantification of enkephalin-and endorphin-related peptides in biologic extracts including brain and tooth pulp tissue.     

High-Performance Liquid Chromatographic Assay for Chlorquine and its Two Major Metabolites,Desethylchloroquine and Bidesethylchloroquine in Biological Fluids

Abstract

A high-performance liquid chromatography (HPLC) method for the determination of chloroquine and its two major metabolites in biological fluids is described. Hydroxychloroquine is used as an internal standard (I.S.). Drug, metabolites and I.S. were extracted as bases with diethyl ether by a single step procedure. After drying and evaporation of the organic phase, the residue was dissolved into the mobile phase and injected into the chromatographic system. Separation was performed using a normal phase column (Inertsil sill with mixture of acetonitrile, methanol and ammonia as mobile phase. The detection was carried out by fluorescence measurement : excitation wavelength was set at 325 nm and emission at 380 nm. The limit of detection was near 3.7 ng ml^?1 for chloroquine and metabolites. No chromatographic interference could be detected by endogenous compounds or other antimalarial drugs. Because of the good accuracy of the method, concentrations were determinated with a relative standard deviation lower than 7% at the 25 ng ml^?l level for all substances.

An excellent precision was obtained over the range of concentrations tested, 25–1000ng ml^?l. This method can be applied to therapeutic, pharmacokinetic and epidemial studies.     

Pre-Column Switching Techniques for the Determination of Drugs and Metabolites in Body Fluids in Research and Routine Analysis

Abstract

The use of a column switching system for direct injection of samples and of a sample clean-up on reversed phase pre-columns is described. The pre-columns were filled with spherical C-18 silica gel of particle size 30 μm.

Two applications are reported on: (1) the direct injection of serum samples for the simultaneous analysis of nine antiepileptic drugs and metabolites and (2) the determination of phenytoin and of carbamazepine in serum ultra-filtrates.

The purge liquid for the sample clean-up was diluted phosphoric acid, and the eluent mixture for the chromatographic separation was water/acetonitrile. The analytical column (length 12.5 cm) was filled with C-18 silica gel of particle size 5 μm. A gradient elution was chosen for the first application, while the second application was carried out using isocratic chromatographic conditions.     

Simultaneous Determination of Five Antibacterials in Swine Muscle by High Performance Liquid Chromatography

Abstract

A high performance liquid chromatographic method (HPLC) for the determination of olaquindox, morantel, furazolidone, nitrofurazone and carbadox residues in swine muscles was developed. The drugs were extracted from muscles with acetonitrile and cleaned up by alumina column. HPLC analysis was carried out on an Inertsil C8 column with a mobile phase of acetonitrile-water-acetic acid (3:97:1), and the drugs were detected at 340 nm. The average recoveries of all drugs added to muscles at 0.1 ppm level were more than 75% and the detection limit of each drug was 0.03 ppm in muscles.     

Thin-layer chromatographic fingerprinting of the nonvolatile fraction extracted from the medicinal herb Cistus incanus L.

ABSTRACT

The aim of this study was to provide the first from-start-to-end thin-layer chromatographic method of fingerprinting the Cistus incanus L. raw herbal material, with a purpose to further use it for rapid screening, authentication, and quality control of the traded C. incanus L. herbs. To this effect, 12 different C. incanus L. samples purchased as herbal teas from a local market were extracted by means of the accelerated solvent extraction (ASE) with chemometrically optimized solvent extraction mixture and temperature (methanol–water, 27:73, v/v; 130°C), to derive the polar fraction from the plant samples. Then, the extracts were developed in two thin-layer chromatographic systems, both using the commercially precoated silica gel 60 chromatographic plates, yet two different mobile phases (mobile phase 1, ethyl acetate–formic acid–acetic acid–water, 100:11:11:13, v/v/v/v, and mobile phase 2, ethyl acetate–dichloromethane–formic acid–acetic acid–water, 100:10:10:10:11, v/v/v/v/v). The chromatograms were densitometrically scanned in the reflectance mode at the wavelength λ?=?366?nm to obtain fingerprints of the extracts derived from individual C. incanus L. samples. Mobile phase 2 performed slightly better, because with its use, the maximum number of 11 peaks could be seen in the respective fingerprints, whereas with mobile phase 1, the maximum number of 10 peaks only. Then an antioxidant potential of the investigated herbal extracts was assessed, making use of mobile phase 2 and the 0.20% methanol solution of 2,2-diphenyl picrylhydrazyl as a visualizing reagent. The resulting chromatograms were densitometrically scanned in the extinction mode at the wavelength λ?=?550?nm to obtain biological fingerprints of the extracts. Finally, chromatographic and biological fingerprints underwent a semiquantitative evaluation in terms of the contents of the extracted polar fraction and an overall antioxidant potential of the individual plant species.     

Thermal Properties of Binary Mixtures of β-Cyclodextrin with Carbamazepine Polymorphs

The thermal behaviour of binary mixtures between β-cyclodextrin (β-CD) and either carbamazepine polymorphic Form I (CBZ I), Form III (CBZ III) or dihydrate was investigated in order to assess possible interactions of CBZ solid phases with β-CD. Physical mixtures and kneaded binaries of β-CD and different CBZ crystal forms were studied by differential scanning calorimetry, thermogravimetric analysis and hot stage microscopy. The pattern of transition of CBZ Form III into Form I is strongly influenced by β-CD. The liquid-solid transition is practically absent when anhydrous CBZ/β-CD mixes are tested, as a consequence of an interaction between β-CD and liquid CBZ that hinders CBZ recrystallisation as Form I occurring after CBZ Form III melting. Water loss on heating of CBZ dihydrate in the presence of β-CD leads in all cases to the formation of CBZ Form I. This revised version was published online in August 2006 with corrections to the Cover Date.     

High Performance Liquid Chromatographic Analysis of Tolmetin,Indomethacin and Sulindac in Plasma

Abstract

Isocratic and gradient reversed phase high-performance liquid chromatographic (HPLC) methods for the quantitation of tolmetin, indomethacin, and sulindac and their respective metabolites in plasma were developed. Only the determination of the parent drugs was possible using the isocratic technique. Specific, simultaneous determination of each drug and its respective metabolites was achieved using the gradient technique. The effect of pH and ionic concentration of the mobile phase on retention time was studied. Statistical analysis demonstrated excellent precision and linearity over the following ranges: 1–40, 0.1–3, and 0.1–3 ug/ml plasma for tolmetin, indomethacin, and sulindac respectively. Both methods have been applied to the analysis of patient samples.     

High Performance Liquid Chromatographic Determination of Trimethoprim and Sulphadiazine in Medicated Fish Feedstock

Abstract

A high performance liquid chromatographic (HPLC) method for the determination of sulphadiazine and trimethoprim in medicated fish feedstock is described. It is based on the extraction of the drugs into methanol followed by separation on a μ-Bondapak column using an aqueous acetonitrile mobile phase of pH 3.0. The method was validated by determining intra- and inter-assay precision and the coefficient of variation was found to be less than 8% for both drugs. Callbration curves were linear over the range 160–4000 pg drug per g of feedstock and the method was applied to the determination of a commercial feedstock formulation.     

LC–MS3 Strategy for Quantification of Carbamazepine in Human Plasma and Its Application in Therapeutic Drug Monitoring

This study developed a detection method based on the strategy of HPLC/MS³ and verified its suitability by quantifying carbamazepine in human plasma. The high-performance liquid chromatography–tandem mass spectrometry (HPLC/MS³) system was performed using a Shimadzu UFLC XR liquid chromatography and a SCIEX QTRAP^® 5500 linear ion trap triple quadrupole mass spectrometer. The specific operation was as follows: the sample protein was firstly precipitated using methanol, then carbamazepine and carbamazepine-D₂N¹⁵ were separated on an ACQUITY UPLC HSS T3 column using the gradient elution with solvent A (0.1% formic acid) and solvent B (0.1% formic acid in acetonitrile) at a flow rate of 0.25 mL/min. Each sample was run for 7 min. This method was validated for various parameters including accuracy, precision, selectivity, linearity, LLOQ, etc. Only 5 μL of sample plasma could obtain the result of LLOD 0.5 µg/mL. The intra-day and inter-day precision was <8.23%, and accuracy was between −1.74% and 2.92%. This method was successfully used for monitoring the blood concentration of epilepsy patients after carbamazepine treatment.