摘要:

AbstractIn order to improve the urinary neopterin measurement, the reversed‐phase HPLC method has been reevaluated. The parameters which influence the chromatographic behavior of 12 pteridines were studied: nature of buffer, pH,ionic strength, addition of organic modifier to the mobile phase. Accordingly, an isocratic HPLC method is described which offers a good compromise between specificity and analysis time. This method is well‐suited to automation in routine clinical laboratory use. Using this HPLC method, urinary neopterin related to creatinine was determined in lung diseases (neoplasm, sarcoïdosis and bronchial asthma) and in kidney allografts. This method was shown to be useful in the diagnosis and in the monitoring of treatment of rejection epis

ISSN:0269-3879    

DOI:10.1002/bmc.1130020502

出版商:John Wiley&Sons, Ltd.    

年代:1987

数据来源: WILEY

摘要:

AbstractWe have developed a high performance thin layer chromatography (HPTLC) system for quantitative determination of androgens, corticosteroids, mineralocorticoids and gestagens on silicagel KG‐60 HPTLC‐plates with different solvent systems. A complete separation of androgens, gestagens and metabolites was achieved with dichlormethane/cyclohexane/acetone (70:25:5). Corticosteroids, mineralocorticoids and their derivatives were completely separated with diethylether/isooctane/isopropanol (70:25:5). The quantitativein situfluorescence determination was carried out after post‐chromatographic derivatization with cinnamic aldehyde, 4‐dimethylaminobenzaldehyde and sulfuric acid. The sensitivity of detection was found between 500 pg and 1 ng per spot.The steroid metabolism as catalysed by rat liver microsomal oxidoreductases was measured by these procedures, and was compared with determination of steroids by gas chromatography (GC). According to HPTLC, steroids were reduced by NADPH‐5α‐reductase (EC 1.3.1.4) in the order progesterone>testosterone>aldosterone>cortisol>corticosterone. The enzyme activities as measured by HPTLC agree well with those obtained by GC (r= 0.94). When turnover of enzyme assays, speed of determination, detection limit, application to labile steroids and costs of steroid determination are considered, all points speak in fav

ISSN:0269-3879    

DOI:10.1002/bmc.1130020503

出版商:John Wiley&Sons, Ltd.    

年代:1987

数据来源: WILEY

摘要:

AbstractA rapid and specific method has been developed for the determination of orphenadrine concentration in plasma. It involves a one‐step sample preparation usingn‐hexane/isopropyl alcohol (98:2) extraction, and analysis by gas chromatography on a wide bore capillary column using nitrogen/phosphorus detection. This procedure considerably simplifies previously reported assays and is specific and sensitive enough for the determination of orphenadrine in plasma of patients on chronic ther

ISSN:0269-3879    

DOI:10.1002/bmc.1130020504

出版商:John Wiley&Sons, Ltd.    

年代:1987

数据来源: WILEY

摘要:

AbstractTissue concentrations of the amino acids tryptophan, methionine, valine, phenylalanine, isoleucine and leucine were determined in rat liver, skeletal muscle and brain tissue and human plasma. The effect of storage at −20°C for up to 21 days was examined. Significantly elevated levels after 7 and 21 days of storage were observed in liver and brain tissue but no changes were found in muscle and plasma. Minimal changes occurred in liver tissue stored up to 6 hours at −4°C. This study emphasizes the need for rapid deporteinization of tissues to avoid changes in the free pool of amino

ISSN:0269-3879    

DOI:10.1002/bmc.1130020505

出版商:John Wiley&Sons, Ltd.    

年代:1987

数据来源: WILEY

摘要:

AbstractNew methods are presented for measuring total vitamin C and the ascorbic acid/dehydroascorbic acid ratio in gastric juice. Extracts are prepared from a gastric juice which are suitable for direct injection onto a Waters Nova‐pak C18Radial‐pak cartridge for high performance liquid chromatography (HPLC) using ultraviolet absorbance at 270 nm for detection. Both enable removal of interfering mucus and mucopolysaccharide breakdown products in a novel way. The first uses mini‐columns of Sephadex G‐50, run in acidic conditions to remove large molecular weight material while maintaining the ascorbic acid/dehydroascorbic acid ratio as it was in the fresh sample. Addition of dithiothreitol converts the dehydroascorbic acid quantitatively to ascorbic acid, thus enabling measurement of both components. The second method converts all the dehydroascorbic acid to ascorbic acid at the outset. A perchloric acid extract is neutralized and passed through a Sep‐Pak C18. A new internal standard, reductic acid, is introduced for ascorbic acid analysis which behaves identically on Sep‐Pak C18. Samples are analysed by ion‐pair chromatography using 0.02 M NH4H2PO4buffer (pH 7.1): methanol (80:20 v/v) containing 0.62 g/L tetrapentylammonium bromide. The detection limit was 1 ng ascorbic acid, and chromatography was completed in 5 min. The values obtained by the two independent HPLC methods were in good agreement with each other and with those obtained by the 2,4‐dinitrophenylhydrazine colo

ISSN:0269-3879    

DOI:10.1002/bmc.1130020506

出版商:John Wiley&Sons, Ltd.    

年代:1987

数据来源: WILEY

摘要:

AbstractMethods for the determination of CS‐570, a chemically stable prostacyclin analogue, in plasma were developed by using an immobilized antibody column followed by fluorescence HPLC and GC/MS. The CS‐570 antibody, obtained from rabbit plasma by giving CS‐570–BSA for a few months, was coupled to Sepharose 4B and used as extraction phase for sample clean‐up and extraction of the drug. A plasma sample was applied to this column, washed with water and the drug was eluted with 90% acetonitrile. 0.02% (w/v) 9‐anthryldiazomethane (ADAM) was added to the extract to form a fluorescent derivative. The CS‐570–ADAM adduct exhibited high sensitivity when applied to HPLC with fluorescence detection and column switching. The detection limit was 1 ng/mL when 1 mL of plasma was available. Additionally, a pentafluorobenzyltrimethylsilyl derivative of CS‐570 showed excellent sensitivity when determined by capillary GC interfaced to negative ion chemical ionization MS using a stable isotope labelled analogue as an

ISSN:0269-3879    

DOI:10.1002/bmc.1130020507

出版商:John Wiley&Sons, Ltd.    

年代:1987

数据来源: WILEY

摘要:

AbstractThree different direct HPLC methods for the determination of 3‐methoxy‐4‐hydroxymandelic acid (VMA, vanillylmandelic acid), 5‐hydroxyindoleacetic acid (5‐HIAA) and 3‐methoxy‐4‐hydroxyphenylacetic acid (HVA, homovanillic acid) in urine were compared: two spectrofluorometric methods, applying discontinuous gradients, and one serial coulometric linear gradient method. The imprecision study (n= 6) revealed comparable coefficients of variation (CV), intra‐assay ranging 1.4–11.1%, and inter‐assay ranging 5.9–11.8% for physiological and moderately elevated levels of VMA, 5‐HIAA and HVA. All methods showed good linearities up to 100 μmol/L for each of the three compounds studied. Analytical recoveries were 97–114% for VMA, 87–103% for 5‐HIAA, and 80–95% for HVA. Recoveries were not dependent on urinary relative densities in the range 1.010–1.030 kg/L or on protein content (prior to acidification) in the range 0.1–3 g/L, or on the pH of conservation in the range 2–5 or on storage temperature in the range −20–+22°C for three weeks.The distributed‐sample comparison revealed acceptable correlations and clinically unimportant accuracy differences between the methods. It is concluded that direct fluoro netric and electrochemical HPLC methods can be used in the determination of major catecholamine and serotonin metabolites in human urine for clinical diagnosis and

ISSN:0269-3879    

DOI:10.1002/bmc.1130020508

出版商:John Wiley&Sons, Ltd.    

年代:1987

数据来源: WILEY

摘要:

AbstractCysteamine, an amino thiol, was separated by rapid isocratic cation exchange chromatography and detected by electrochemical oxidation at a platinum electrode maintained at +0.45 V relative to an Ag/AgCl reference electrode. Eluent pH and electrode working potentials were optimized and the effects of alternative buffers and organic modifiers have been examined. On column sensitivity for cysteamine was 1.5 pmol at a signal‐to‐noise ratio of 5. Although the specificity was good, plasma samples required maximal sensitivity whereas urine samples required greater selectivity, which was achieved by use of lower working potentials. Cysteamine concentrations were determined in serial samples of plasma and urine from volunteers who had received a single oral dose of 200 mg of the drug. Cysteamine was rapidlyoxidized in vivo, and detection required prior reduction with dithiothreitol before analy

ISSN:0269-3879    

DOI:10.1002/bmc.1130020509

出版商:John Wiley&Sons, Ltd.    

年代:1987

数据来源: WILEY

摘要:

AbstractA one‐step HPLC method was developed for the purification of protein G, a cell wall molecule from group C and G streptococci with immunoglobulin G‐ and albumin‐binding properties. LysedEscherichia colibacteria infected with lambda‐phages containing the protein G gene from group G streptococci were used as a starting material for the preparations. The lysate was applied to a column with immobilized human immunoglobulin G or human serum albumin. Protein G was selectively bound and eluted at pH 2.0. A 750‐fold purification was achieved. Sodium dodecylsulfate + polyacrylamide gel electrophoresis showed that the highly purified protein G consisted of three sets of doublets with the apparent molecular weight of 64 and 67, 56 and 58, and 45 and 47 kilodaltons, respectively. A specific method for quantitation of small amounts of protein G was developed and used for specific tracing of the protein after the affinity chromatography. Goat polyclonal antibodies were bound to an antigen coated to the plastic walls of microtiter plates, causing the Fc‐region of the immunoglobulins to be directed outwards. Unknown samples of protein G were then allowed to compete with radio‐iodinated protein G (solid phase radioassay) or protein G coupled to alkaline phosphatase (enzyme linked sorbent assay) for t

ISSN:0269-3879    

DOI:10.1002/bmc.1130020510

出版商:John Wiley&Sons, Ltd.    

年代:1987

数据来源: WILEY

摘要:

AbstractA rapid method for the determination of 4‐methylpyrazole in plasma is described. Internal standard (3‐methylpyrazole) is added to the plasma which is subsequently applied to a BondElut SCX column. After washing the column the 3‐ and 4‐methylpyrazoles are eluted with a phosphate buffer and analyzed by isocratic reversed‐phase high performance liquid chromatography with UV detection. The concentration of 4‐methylpyrazole is calculated from the 4‐methylpyrazole/3‐methylpyrazole peak high ratio. The method is linear between 2.5 and 100 μmol/L with a within‐day precision (CV) of 2.2% (n= 10) and a day‐to‐day precision of 2.8% (n= 30). The sensitivity is sufficient for analysis of plasma levels in th

ISSN:0269-3879    

DOI:10.1002/bmc.1130020511

出版商:John Wiley&Sons, Ltd.    

年代:1987

数据来源: WILEY