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Biomedical subjects

W Voelter

Publications and source records attributed to W Voelter.

At least 145 records · Page 8Linked to original sources

Usefulness of Ficoll in electric field-mediated cell fusion.

Electrofusion is a technique that enables the production of new cell types with desired properties to be done. Ehrlich ascites tumor cells are fused by means of an electric field. Under too harsh external field strength or pulse length conditions, however, membrane breakdown leads to a loss of cellular cytoplasm. Addition of the high polymer Ficoll to the fusion medium increases its density and osmotic pressure, thus preventing cytoplasm from running out and, therefore, maintaining cell viability. Fusion between cells of different sizes is made possible or facilitated by means of Ficoll, as big cells do not lose their cytoplasm under conditions required for enhancing membrane permeability of the small cells. In presence of the proteolytic enzyme pronase, addition of Ficoll to the fusion medium further raises the fusion percentage compared to the exclusive addition of pronase. The fusion of cells of different densities is also greatly facilitated and its percentage increased by addition of Ficoll, thus obviating the necessity to perform electrofusion under conditions of microgravity.

Animals↗

Isolation purification and properties of a site-specific proteolytic enzyme "valyl-proteinase" from Candida tropicalis.

A highly specific proteolytic enzyme cleaving at the carboxyl group of valine has been isolated from Candida tropicalis. Its specificity has been determined by digesting beta-lactoglobulin and a number of synthetic peptides. The enzyme a glycoprotein, has a molecular mass of 40 +/- 7 kDa on the basis of sodium dodecyl sulphate polyacrylamide gel electrophoresis. Its optimum activity occurs at 37 degrees C at a pH between 8-9. It has been named "Valyl-proteinase" because of its selective cleavage.

Amino Acids↗

High-performance liquid chromatographic comparison of in vivo and in vitro drug metabolism.

Using 2-ethyl-3-(4- hydroxybenzoyl )benzofuran ( EHBB ) as an example, biotransformation in rabbits and rats and by rat hepatocyte suspensions was studied by high-performance liquid chromatography (HPLC) and mass spectrometry. The biotransformation of N-alkyl-substituted piperidines by rat hepatocytes gives valuable information about the pharmacodynamics of this series of compounds. It is demonstrated that a simple reversed-phase HPLC pre-column technique is much superior to the classical sample purification via extraction. The utility of hepatocytes for the investigation of drug metabolism is demonstrated.

Animals↗

[A new peptide alkaloid from Discaria febrifuga Mart].

On the basis of spectroscopic investigations (1H-NMR, 13C-NMR, UV, IR and MS spectroscopy), elemental analysis and degradation reactions the structure of a new alkaloid discarine G from the bark extract of Discaria febrifuga Mart. is elucidated.

Acetylation↗

Quantitative determination of O-(beta-hydroxyethyl)-rutosides in serum by high-performance liquid chromatography.

A procedure for the quantitative determination of O-hydroxyethylated rutosides by high-performance liquid chromatography is described, which can be used for the detection of these modified flavonoids in human serum. Serum samples are processed by the addition of acetone, which removes most of the proteins. After passing the supernatant through a microcolumn of Amberlite XAD-2 and washing with water, the hydroxyethylated rutosides are eluted with methanol. The eluate is concentrated in vacuo. The methanolic solution of the residue is chromatographed on RP-8 columns using UV and fluorescence detectors. The mono- to tetrahydroxyethylated constituents and their corresponding aglycones could be separated with a step gradient, starting with a solvent system of water-methanol-acetic acid (70:30:6) followed by a mixture of water-ethanol-acetic acid (70:30:6). Alternatively, the rutosides can be separated by a linear gradient of water-acetonitrile. An almost linear calibration curve and about 80% recovery are obtained. A detection limit of 1 mg/l is achieved. Pharmacokinetic studies in human volunteers are described.

Chromatography, High Pressure Liquid↗

Radioimmunoassay of 5 alpha-pregnane-3,20-dione. A metabolite of placental progesterone.

A radioimmunoassay (RIA) procedure has been developed for the measurement of 5 alpha-pregnane-3,20-dione (DHP) in human plasma after ether extraction of the plasma-samples, followed by column chromatography. The antiserum was generated in rabbits with 6 alpha-carboxyethylmer-captoprogesterone-BSA conjugate. The high affinity (Ka = 3.72 X 10(9) l/mol) antiserum binds 40% of 40 picograms of tritiated 5 alpha-pregnane-3,20-dione at working dilutions of 1 : 2800. Negligible cross-reactivity of the antiserum was detected with 5 beta-pregnane-3,20-dione (1.8%). Other hydroxylated pregnanes showed minor cross-reactivity (5-40%). The cross-reacting steroids were all separated from pregnanedione by one chromatographic step. The plasma levels of 5 alpha-pregnane-3,20-dione were measured from 22nd to 42nd week of normal pregnancy and compared to published data.

Female↗

[Peptide alkaloids from Discaria febrifuga Mart].

From the bark extract of Discaria febrifuga Mart. the known alkaloids frangufoline (I), adouétine (II), scutianine B (III), franganine (IV) and two new ones, discarine C (V) and D (VI), were isolated. Their structures are elucidated by degradation reactions and spectroscopic investigations.

Alkaloids↗

A simple high-performance liquid chromatographic pre-column technique for investigation of drug metabolism in biological fluids.

A simple reversed-phase high-performance liquid chromatographic pre-column technique for investigation of drug metabolism is described. This rapid method circumvents the "classical" sample purification via extraction by direct purification and enrichment of the sample on the pre-column. Almost 100% recovery of a drug (aminopyrine) and its metabolites from biological fluids is achieved. This is in strong contrast to "classical" sample preparation which allows a recovery of 30-100% depending on the polarity of the investigated compound. The procedure described has been successfully applied to the investigation of the metabolic pattern of aminopyrine in rat plasma and cell incubation media.

Aminopyrine↗

Fluorescence labelling in trace analysis of biological samples: simultaneous determination of free fatty acids and related carboxylic compounds.

Free fatty acids in serum, together with representatives of a new class of oxirane carboxylic acids having blood-glucose-lowering activity, were determined simultaneously by fluorescence labelling with 4-bromomethyl-7-methoxycoumarin (Br-Mmc) followed by separation by high performance liquid chromatography. Esterification kinetics of selected compounds were studied, and separation conditions for analysis of oxirane carboxylic acids, related compounds, and a series of saturated and unsaturated fatty acids as their Mmc estes were established. Three compounds (3, 6 and 7) were determined from serum quantitatively by extraction, derivatization, and analysis by high-performance liquid chromatography. Detection limits from serum (1ml) were 50 ng/ml. Two hours after intravenous bolus injection of 10 mg/kg of compound 6 in dog, a serum level of 1.1 mg/1 of administered compound was determined.

Animals↗

Determination of selected pyrimidines, purines and their metabolites in serum and urine by reversed-phase ion-pair chromatography.

The qualitative and quantitative determination of selected pyrimidines, purines and azapurines and their metabolites by reversed-phase ion-pair high-performance liquid chromatography, using tetrabutylammonium hydroxide as pairing ion and isocratic chromatrographic conditions, is described. The method provides a selective and sensitive assay for these classes of compounds examined in complex biological fluids.

Allopurinol↗

High-performance liquid chromatographic investigation of the amino acid, amino sugar and neutral sugar content in glycoproteins.

A methods for the simultaneous separation and determination of amino acids, amino sugars and neutral carbohydrates is described. Stepwise elution systems with sodium citrate and borate buffers have developed for the ion-exchange liquid chromatographic separation of amino acids and sugars, using 8-micrometer particle size resins and the Stein and Moore and orcinol colorimetric method for detection. With the aid of this system, the direct quantitative comparison of sugars and amino acids by liquid chromatography becomes possible for the first time.

Amino Acids↗

High-performance liquid chromatographic analysis of amino acids and peptide-hormone hydrolysates in the picomole range.

A high-pressure liquid chromatographic system for the separation of amino acids and peptide-hormone hydrolysates is described. A modified high-pressure liquid chromatography (Hewlett-Packard, model 1010B) equipped with an automatic gradient-generating system, and the applications of fluorescamine and ion-exchange resins of particle size 8 +/- 2 mum permit the detection of amino acids in the picomole range with good resolution. Compared with conventional amino acid analyzers, the times for separation are reduced by a factor of 3--4.

Amino Acids↗

The initial binding of Cu(II) to some amino acids and dipeptides: a 13C nuclear-magnetic-resonance study.

The initial binding of Cu2+ ot L-lysine, L-histidine, glycyl-histidine and histidyl-glycine in aqueous solutions was examined by 13C nuclear magnetic resonance spectroscopy. The measurements were carried out in a substantially improved way employing the pulse Fourier transform technique. Spectra of both high quality and resolution were obtained. Cu2+ complex formation with L-lysine occurred with the alpha-amino and carboxyl group attributable to the well expressed broadening effect of the 13C signals of the alpha-carbon atom and the carboxyl atom. The epsilon-amino group was not involved. Measurements of the Cu chelates using L-histidine and glycyl-histidine and histidyl-glycine confirmed the ambidentate nature of the histidine residue. It was concluded that an equilibrium exists between two Cu-complex species designated as histamine-like and histamine-like/glycine-like species. In the homogeneous histamine-like Cu complex, the Cu2+ is exclusively bound with 4 nitrogens, while in the other species one oxygen of the glycyl carboxyl group is involved in the Cu2+ binding. Blocking of this carboxyl groups by peptide bonding as found in histidyl-glycine favoured the formation of a Cu complex where the imidazole carbons of the histidyl residue were the most influenced species.

Amino Acids↗