High Perfomance Liquid Chromatography Research Articles

Solid-phase extraction–high-perfomance liquid chromatography determination of verapamil and norverapamil enantiomers in urine

A simple, rapid, sensitive and selective method has been developed for the stereospecific determination of verapamil and its metabolite, norverapamil in urine. For sample preparation we utilized a membrane-based solid-phase extraction (SPE) disk consisting of a thin, particle-loaded membrane inserted in a plastic syringe-like barrel. The particles, which may be C 8 or C 18 bonded phase (C 8 in this work), are embedded within a matrix of PTFE (Teflon) fibrils. Overall analyte recoveries were above 85%, even at low concentration of 3.0 ng/ml with reproducibilities (C.V. values) below 13.1%. This method of extraction has the advantage of speed and considerable reduction in solvent volumes compared to conventional SPE and solvent extraction. The separation of all the enantiomers was achieved using a single chiral stationary phase column, the cellulose-based reversed-phase, Chiralcel OD-R. Analyte concentrations of less than 3.0 ng/ml could be quantitated with C.V. values below 14%. Calibration curves were linear in the range 2.5–300 ng/ml. Intra-day and inter-day reproducibilities were 10.5–14.2% at 3 ng/ml, 4.8–9.3% at 138.5 ng/ml and 7.8–10.1% at 280 ng/ml level, respectively, for all the enantiomers.

Occurrence of Fusaproliferin, Fumonisin B1, and Beauvericin in Maize from Italy

The occurrence of fusaproliferin (FP) was investigated in 22 samples of preharvest maize ears Fusarium rot, collected in 1994 in Italy. The most frequently recovered Fusarium species were Fusarium moniliforme and Fusarium proliferatum. Chemical analysis by high-perfomance liquid chromatography and gas chromatography/mass spectrometry identified FP (up to 500 mg/kg) in nine samples mostly infected by F. proliferatum. FP was often found to be associated with fumonisin B1 (FB1) (up to 300 mg/kg) and beauvericin (BEA) (up to 520 mg/kg). The occurrence of FP as a natural maize contaminant and its co-occurrence with FB1 and BEA is reported here for the first time. Keywords: Fusaproliferin; fumonisin B1; beauvericin; maize; Fusarium; toxins; contamination

Kinetics of hydrolysis of monofunctional and bifunctional monochloro- s-triazine reactive dyes

The kinetics of hydrolysis of one monochloro- s-triazine and one bis(monochloro- s-triazine) reactive dye were investigated at dyeing conditions of pH 11 and temperature 80°C, and, for the monofunctional dye, also at the lower temperature of 20°C. The alkaline hydrolysis was monitored by high perfomance liquid chromatography. The pseudo-first-order rate constants of the monochloro- s-triazine dye, the bis(monochloro- s-triazzine) form and the monochloromonohydroxybis- s-triazine form of the bifunctional dye were determined and compared.

関節炎モデルにおける血中ピリジノリン測定の有用性

Urinary pyridinoline (U-Py) is a useful marker of bone and cartilage loss in rheumatoid arthritis (RA) . The serum pyridinoline (S-Py) concentration instead of U-Py was measured both in rats with adjuvant-induced (AA) and mice with collagen-induced (CIA) arthritis by high perfomance liquid chromatography (HPLC) . On the seventh day after adjuvant innoculation, the AA rats showed a significant increase in S-Py levels. The level of S-Py was gradually increased with the development of AA. Moreover, in CIA mouse, another arthritic model, on sixty days after innoculation of the type II collagen the S-Py level was also significantly higher than that in the normal mice.These findings indicate that the measurement of S-Py instead of U-Py may be useful as marker of bone and cartilage destruction in patients with rheumatoid arthritis.

Determination of Nematicide Aldicarb and Its Metabolites Aldicarb Sulfoxide and Aldicarb Sulfone in Soils and Potatoes by Liquid Chromatography With Photodiode Array Detection

Abstract Solid phase extraction has been combined with High Perfomance Liquid Chromatography using a Photodiode Array Detector to isolate, determine, quantify and recover trace concentrations of aldicarb and its toxic metabolites, aldicarb sulfoxide and aldicarb sulfone, in soils and potatoes. A method is proposed which eliminates matrix interferences. The procedure involved extracting with acetonitrile or dichloromethane, initial separation and clean-up with a Sep-Pack Florisil or Silica cartridges. The extract can be successfully analyzed by HPLC-DAD and recoveries for pesticides were not influence by the use of this extraction method. The results have been applied to the systematic study of these compounds in growns of patatoes. The lowest detectable concentration for each pesticide is 40 μg kg−1 in soil and 15 μg kg−1 in potatoes.

Structure of hydroxycinnamic acid derivatives established by high-perfomance liquid chromatography with photodiode-array detection

Structure of hydroxycinnamic acid derivatives established by high-perfomance liquid chromatography with photodiode-array detection

Carotenoid and Carotenoid Ester Composition in Mango Fruit As Influenced by Processing Method

The predominant carotenoids and carotenol fatty acid esters in extracts from fresh, frozen, and canned mango fruit slices have been separated and identified by high-perfomance liquid chromatography on two «in series» C 18 reversed-phase columns using a diode array detector. The pigment pattern was composed of oxygenated carotenoids (xanthophylls), carotenol mono(fatty acid esters), hydrocarbon carotenoids, and carotenol bis(fatty acid esters). The major identified components were antheraxanthin, violaxanthin, cis-isomers, and epoxy derivatives of lutein and β-carotene. Canned mango fruit slices showed a significantly different pattern with β-carotene the main carotenoid component (90%). Large differences in Hunter color values were observed between fresh or frozen and canned mango slices

Transglycosylation Activity of Mucor Hiemalis Endo-β-N-Acetylglucosaminidase Which Transfers Complex Oligosaccharides to the N-Acetylglucosamine Moieties of Peptides

A novel endo-β-N-acetylglucosaminidase in the culture fluid of Mucor hiemalis isolated from soil was found to have transglycosylation activity. This endo-β-N-acetylglucosaminidase, Endo-M, could liberate the complex type of asparagine-linked oligosaccharides by hydrolysis of diacetylchitobiose linkage from glycoproteins. The treatment of Endo-M with N-acetylglucosamine and asialotransferrin glycopeptide having the complex type of oligosaccharides resulted in the transfer of the released oligosaccharide from the glycopeptide to N-acetylglucosamine. The structure of the product after transfer was deduced to be (GlcNAc)2-Man-(Gal-GlcNAc-Man)2 by a combination method of pyridylamination and high perfomance liquid chromatography, and mass-spectrometry. The enzyme could transfer the complex type of oligosaccharide from asialotransferrin glycopeptide to peptidyl-N-acetylglucosamine prepared from bovine ribonuclease with the high-mannose type of oligosaccharide. This will lead to the construction of neoglycoproteins containing different types of oligosaccharides.

Distribution of unusual long and branched polyamines in thermophilic eubacteria belonging to "Rhodothermus," Thermus and Thermonema.

Polyamines of thermophilic Gram-negative eubacteria, "Rhodothermus marinus" ATCC 43812, Thermus sp. ATCC 43814 and Thermonema lapsum ATCC 43542 were analyzed by high-perfomance liquid chromatography and gas chromatography-mass spectrometry. "R. marinus" contained spermidine, spermine, thermopentamine, a tertiary tetraamine (N4-aminopropylspermidine) and a quaternary pentaamine (N4-bis(aminopropyl) spermidine). Thermus sp. ATCC 43814 contained putrescine, cadaverine, norspermidine, spermidine, homospermidine, norspermine, spermine, thermospermine, aminopropylhomospermidine, caldopentamine, agmatine, two tertiary tetraamines (N4-aminopropylnorspermidine and N4-aminopropylspermidine) and two quaternary pentaamines (N4-bis(aminopropyl)norspermidine and N4-bis(aminopropyl)spermidine). Homospermidine and homospermine were detected in Thermonema lapsum as the major polyamine. These distribution patterns of long and branched polyamines are distinctive in the thermophiles, indicating that unusual polyamine profiles serve to estimate chemotaxonomic and phylogenetic relationships within thermophilic eubacteria.

Genetic nature of low capsaicin content in the variant strains induced by grafting in Capsicum annuum L.

The genetic nature of low capsaicin content in variant strains induced by grafting was elucidated by crossing the G5S23 strain with the two cultivars, ‘Yatsubusa’ (Capsicum annuum L. var. fasciculatum Ilish) and ‘Spanish Paprika’ (C. annuum L. grossum Sendt) originally used for grafting. Decreased capsaicin synthesis was a stable characteristic for at least several generations from G5S16 to G5S23 and was transmitted to the progenies of crosses. This fact shows that the decreased capsaicin synthesis is a genetic trait in the graft-induced variant strains. The genetically dependent characteristic of pungency of the G5S23, ‘Yatsubusa’ and ‘Spanish Paprika’ were analysed by High Perfomance Liquid Chromatography measurement and tasting assay.

グルタチオンの安定性に及ぼす微量元素の影響と安定化

Glutathione (GSH) was combined with trace elements in infusion solution for total parenteral nutrition.The effects of the trace elements (copper, zinc and selenium) on the stability of GSH, and the stabilization of GSH by protamine sulfate (PT) and ascorbic acid (AC) were studied by reversed-phase high-perfomance liquid chromatography using 4-(amino-sulfonyl)-7-fluoro-2, 1, 3-benzoxadiazole (ABD-F).The contents of GSH decreased in the presence of zinc, copper, and selenium, in that order.In particular, 14%of GSH remained after 10 minutes at 25°Cin the presence of 5μM of selenium.The contents of GSH decreased with an increase in the concentration of selenium.GSH was stabilized by PT or AC in the presence of trace element.When 5μM of cupric sulfate was added to the GSH solution after 20 minutes, 39.5% of GSH remained.On the other hand, 70.6% of GSH remained under the same conditions in the presence of 10mg of AC per ml.

Protein a, hydroxyapatite and diethylaminoethyl: Evaluation of three procedures for the preparative purification of monoclonal antibodies by high-perfomance liquid chromatography

Three rapid, reproducible and feasible monoclonal antibody purification procedures by means of high-performance liquid chromatography have been evaluated. The stationary phases employed were high-performance hydroxyapatite, high-performance Protein A and high-performance anion-exchange resin. The purity of the final immunoglobulin preparations was determined by sodium dodecyl sulphate-polyacrylamide gel electrophoresis under reducing conditions and, subsequently, by high-performance gel permeation chromatography. The immunoreactivity of purified monoclonal antibodies was determined by the radioimmunoassay method.

On-line post-column fluorescence derivatization of arginine-containing peptides in high-perfomance liquid chromatography

A selective detection system based on on-line post-column fluorescence derivatization is described for the analysis of arginine-containing peptides by high-performance liquid chromatography. The peptides are automatically converted into fluorescent derivatives with benzoin, a fluoregenic reagent for guarnidino compounds, after separation on a reversed-phase column (TSK gel ODS-120T) and detection in an ultraviolet absorption detector. The system permits fluorescence detection at 435 nm emission with irradiation at 325 nm for arginine-containing peptides in as little as picomole amounts. The chromatogram obtained with fluroscence detection only shows peaks corresponding to arginine-containing peptides. The facile detection of arginine-containing fragments in the tryptic digest of β-melanocyte stimulating hormone as a model compound could be achieved by comparison with a chromatogram obtained with ultraviolet absorption detection at 215 nm.

Evolution of phenolic acids and aldehydes during the different production process of “Fino” Sherry wine

The separation and determination of phenolic acids and aldehydes in “Fino” sherry wine, using high-perfomance liquid chromatography (HPLC), has enabled a study to be carried out of their evolution during the initial process in “fino” sherry wine production. A continuous extraction method with ethyl ether at pH 2 was chosen, by which the greatest reprodubility in the extraction of these compounds is achieved from must as well as from the wine. For stabilization of the must samples, a mixture of ascorbic acid (0.2%) and DMF (15%) was used, preventing the evolution of colour and the beginning of fermentation. These methods have permitted the study of changes in the content of the phenolic acids and aldehydes during the different stages of “fino” wine making.

Determination of the beta-adrenoceptor blocking drug bupranolol in plasma and tissues of the rat by high-perfomance liquid chromatography with ultraviolet detection

Determination of the beta-adrenoceptor blocking drug bupranolol in plasma and tissues of the rat by high-perfomance liquid chromatography with ultraviolet detection

高速液体クロマトグラフィーによる穀物中のLuteoskyrinの定量

高速液体クロマトグラフィーによる穀物中のLuteoskyrinの定量

The detection of leukotrienes in the carrageenin-induced pleurisy of rat.

As an inflammatory model, rat carrageenin-induced pleurisy was used. The pleural exudate was collected 5 hr after the injection of the 2% λ-carrageenin solution to the right pleural cavity. After measurement of the exudate volume, the extraction of leukotriene (LT) and HETE was performed by addition of 4-fold volumes of ethanol. This crude extract caused the very slow contraction of the guinea-pig (GP) ileum, which was similar to that of LTC4 and was inhibited by FPL55712(1 μg/ml). For further puriflication, LTC4 and LTD4 in the sample were separated as 80% ethanol eluate with Sepahdex LH-20 and LTB4, and HETEs were extracted with diethylether. After evaporation, the purified samples dissolved in methanol were applied to high perfomance liquid chromatography. The peaks, coinciding to the retention time of LTC4 and LTD4, respectively, were collected for examination of the contraction on GP ileum. The contraction by the sample was also inhibited by FPL55712. When the ratios of the contractile activity over the peak height on HPLC were compared, the values of the two peaks of the sample were similar to those of LTC4 and LTD4, respectively. LTB4 in the pleural fluid at 5 hr was smaller than 5,12-diHETE. Several mono-HETEs were also found. These results show the presence of LTC4 and LTD4 in the pleural fluid at 5 hr. As LTC4 and LTD4 are potent in plasma exudation, they might play a role in the carrageenin-induced pleurisy at 5 hr.

High-perfomance liquid chromatography of free and bound phenolic acids in the eggplant ( Solonum melongena L.)

High-perfomance liquid chromatography of free and bound phenolic acids in the eggplant ( Solonum melongena L.)

Rapid separation of biologically active syrian hamster lymphotoxin in high yield by size-exclusionl high-performance liquid chromatography

A rapid, simple and reproducible high-perfomance liquid chromatography (HPLC) procedure is described for the partial purification of biologically active Syrian hamster lymphotoxin. Lymphotoxin, a series of lymphocyte derived immunological glycoprotein hormones with molecular weights of 20,000 to 56,000, has been partially purified by a single-step HPLC procedure or by a two-step isoelectric focusing—HPLC procedure. The HPLC separation method uses silica-based protein separation columns eluted with a 0.1 M sodium phosphate, pH 7.4, 0.1% polyethylene glycol 4000 bufer at a flor-rate of 0.5 ml/min ar room temperature. Nearly complete recoveries of biologically active lymphotoxin can be obtained with as much as a 13-fold purification in less than 1 h.

Determination of Zero Retention Times (t0) by Temperature Dependent Reversed Phase High Perfomance Liquid Chromatography

Abstract For a given reversed phase HPLC system the zero retention time (t0) can be determined by measuring brutto retention times at different temperatures. For calculation of t0 value temperature intervals must be equidistant in 1/T. If at least two compounds are separated on the column having almost the same sorption enthalpies a more simple intersecting point method can be practised. In this case only two temperature points are necessary for graphic evaluation. The presented methods for determination of t0 values are proved for noradrenaline and adrenaline under selected experimental conditions using aqueous solutions of alkaline perchlorates as mobile phases. Thus t0 values can be determined with an accuracy and a precision of less than ± 3%.