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Biomedical subjects

C K Lim

Publications and source records attributed to C K Lim.

At least 109 records · Page 6Linked to original sources
PubMed1987

High-performance liquid chromatographic analysis of phytoestrogens in soy protein preparations with ultraviolet, electrochemical and thermospray mass spectrometric detection.

The phytoestrogens daidzein, genistein, coumestrol, formononetin, and Biochanin A are separated on a C18 reversed-phase column (Hypersil ODS) with methanol-0.1 M ammonium acetate buffer, pH 4.6 (60:40, v/v) as eluent. The retention and resolution are affected by buffer concentrations, pH type, and proportion of organic solvent in the mobile phase. Detection in the (low pg range) is achieved with an electrochemical detector, and the compounds are positively identified by high-performance liquid chromatography-thermospray mass spectrometry. Daidzein and genistein were found in high concentrations in all soy protein preparations analyzed.

Chromatography, High Pressure Liquid↗
PubMed1987

Rapid high-performance liquid chromatography assay for salivary and serum caffeine following an oral load. An indicator of liver function.

A rapid isocratic reversed-phase high-performance liquid chromatography (HPLC) system for the quantitative measurement of serum and salivary caffeine is described. The best separation of caffeine from other methylxanthines was achieved by chromatography on an ODS-Hypersil column using a solvent system of 0.1 M ammonium acetate pH 4.6-acetonitrile (85:15, v/v). The effluent was monitored at 280 nm. Caffeine was extracted from diluted serum and saliva samples (10-500 microliter) by adsorption on a small Bond-Elut C18 cartridge and recovered by elution with methanol. Thermospray HPLC-mass spectrometry conditions were optimized to afford a means of directly identifying caffeine in samples. The positive-ion mass spectrum was characterized by an intense protonated molecular ion, MH+, at m/z 195 and negligible fragmentation. When the mass spectrometer was operated in selected ion monitoring mode, caffeine could be detected in less than 1 microliter of serum and saliva at a concentration of 1 microgram/ml. Caffeine (3.5 mg/kg body wt.) was administered orally to healthy adults, children, and newborn infants, and to patients with liver disease. The clearance rate and half-life were determined as a test of liver function. A prolongation in the elimination of caffeine was observed in patients with liver disease and, although there was some overlap in the values obtained for patients with noncirrhotic liver disease and healthy persons, the oral caffeine load test may usefully serve as a dynamic assessment of liver function in the serial follow-up of patients with liver disease.

Adult↗
PubMed1987

An HPLC assay for rat liver ferrochelatase activity.

A rapid, reliable, sensitive and reproducible HPLC method was developed for the assay of ferrochelatase activity in rat liver. The assay was carried out aerobically with Zn2+ and mesoporphyrin or protoporphyrin IX as substrates. Zn-porphyrins formed were extracted with dimethyl sulphoxide/methanol (30:70, v/v) containing Zn-deuteroporphyrin as the internal standard for separation and quantification by reversed-phase chromatography. The Km for mesoporphyrin was 5.9 microM, for protoporphyrin IX 8.8 microM and for zinc 6.0 microM. The specific activities were 33.1 +/- 5.0 nmol Zn-mesoporphyrin or 13.4 +/- 2.0 nmol Zn-protoporphyrin formed per hour per mg of protein for mitochondria and 12.3 +/- 2.2 nmol Zn-mesoporphyrin or 4.6 +/- 0.9 nmol Zn-protoporphyrin per hour per mg of protein for liver homogenate.

Animals↗
PubMed1986

Assaying erythrocyte haem biosynthetic enzyme activities by high-performance liquid chromatography with the advanced automated sample processor.

The four cytosolic haem biosynthetic enzymes in erythrocytes were assayed with the Varian advanced automated sample processor (AASP) for rapid sample concentration and clean-up with fast and effective high-performance liquid chromatography systems for separation and quantitation. In the assay for 5-aminolaevulinic acid dehydrase, the porphobilinogen (PBG) formed was extracted on a C18 AASP cartridge and separated by reversed-phase ion-pair chromatography with 32% methanol in 0.05 M sodium acetate buffer (pH 3.5), containing 5.4 mM of 1-heptane-sulphonic acid as eluent. PBG was the substrate for the simultaneous assay of hydroxymethylbilane synthase and uroporphyrinogen III synthase. The uroporphyrinogen I and III isomers formed were oxidised to porphyrins, concentrated on a C2 or C8 cartridge, and separated by reversed-phase chromatography with 13% acetonitrile in 1 M ammonium acetate buffer (pH 5.16) as eluent. Uroporphyrinogen decarboxylase was estimated with pentacarboxylic porphyrinogen III as substrate. The coproporphyrinogen formed was extracted on a C2 or C8 cartridge, oxidised to coproporphyrin and separated by reversed-phase chromatography with 30% acetonitrile in 1 M ammonium acetate buffer (pH 5.16) as mobile phase.

Ammonia-Lyases↗
PubMed1986

Assay of blood acetaldehyde by HPLC with fluorescence detection of its 2-diphenylacetyl-1,3-indandione-1-azine derivative.

A simple sensitive method for the assay of blood acetaldehyde is described. Samples are collected into an organic solution of 2-diphenylacetyl-1,3-indandione-1-hydrazone. Proteins are precipitated immediately and carbonyl compounds react to form fluorescent azine derivatives. Methanol in the reagent mixture prevents the oxidation of any ethanol present, precipitates protein and releases any absorbed acetaldehyde. After protein removal, the reaction is completed by the addition of an acid catalyst and a portion of the reaction mixture is subjected to quantitative analysis by high performance liquid chromatography.

Acetaldehyde↗
PubMed1986

A high-performance-liquid-chromatographic method for the assay of coproporphyrinogen oxidase activity in rat liver.

An h.p.l.c. method was developed for the assay of coproporphyrinogen oxidase activity in rat liver. The protoporphyrinogen IX formed is completely oxidized to protoporphyrin IX for separation and quantification by reversed-phase chromatography with mesoporphyrin as the internal standard. The Km of coproporphrinogen oxidase is 1.01 +/- 0.23 microM. The activities are 4.07 +/- 0.40 nmol of protoporphyrin IX/h per mg of mitochondrial protein and 224 +/- 19 nmol of protoporphyrin IX/h per g of liver tissue homogenate. The method is sensitive enough for measuring enzyme activity in small amounts of human tissue from needle biopsy.

Animals↗
PubMed1986

High-performance liquid chromatography of uroporphyrinogen and coproporphyrinogen isomers with amperometric detection.

A reversed-phase h.p.l.c. system, with an ODS-Hypersil column with acetonitrile or methanol in ammonium acetate buffer as mobile phase, is described for the separation of uro-and copro-porphyrinogen isomers. The porphyrinogens are detected amperometrically with sensitivity comparable with that of the fluorescent detection of porphyrins. The effects of pH, buffer concentration and organic modifiers on retention and resolution were studied. The method is suitable for both analytical and preparative separation of porphyrinogens.

Acetates↗
PubMed1986

Reversed-phase high-performance liquid chromatography of conjugated and unconjugated bilirubins in body fluids.

A novel high-performance liquid chromatography (HPLC) system is described for the separation of bilirubin and its conjugates in body fluids. Biliary bilirubin mono- and diglucuronide can be analysed directly on a C18 reversed-phase column with acetonitrile-dimethyl sulphoxide-0.1 M ammonium acetate (pH 5.16) (50:50:85, v/v/v) as mobile phase. However, the simultaneous determination of conjugated and unconjugated bilirubins in plasma required conversion of the conjugates into their methyl esters by alkaline methanolysis before HPLC separation of the C18 column eluted with acetonitrile-dimethyl sulphoxide-0.50 M ammonium acetate (pH 4.6) (50:50:40, v/v/v). The method is superior to, and more flexible than, previously described reversed-phase systems by allowing precise control of retention times by adjustment of pH, buffer concentration and the relative proportion of organic modifier in the mobile phase.

Bile↗
PubMed1986

Analysis of tumour-localizing haematoporphyrin derivative by high-performance liquid chromatography and fast-atom bombardment mass spectrometry.

Reversed-phase chromatography using a MOS-Hypersil (C8) column with methanol-1 M ammonium acetate buffer (pH 4.6) (60:40) as mobile phase has been developed for the isolation of tumour-localizing haematoporphyrin derivative (HPD). The system effectively resolved the diastereoisomers of haematoporphyrin and its acetyl derivatives. The chromatography peaks were identified by fast-atom bombardment mass spectrometry and were confirmed by chemical synthesis. The main components of HPD before alkaline hydrolysis were diacetylhaematoporphyrin, 8-(1-acetoxyethyl)haematoporphyrin and 3-(1-acetoxyethyl)haematoporphyrin with small amounts of haematoporphyrin, 8-(1-hydroxyethyl)-3-vinyldeuteroporphyrin, 3-(1-hydroxyethyl)-8-vinyldeuteroporphyrin, 8-(1-acetoxyethyl)-3-vinyldeuteroporphyrin, 3-(1-acetoxyethyl)-8-vinyldeuteroporphyrin and protoporphyrin. After hydrolysis with 0.1 M sodium hydroxide, the main components were haematoporphyrin, hydroxyethylvinyldeuteroporphyrins, and protoporphyrin.

Acetylation↗
PubMed1986

Separation of ribonucleotides, deoxynucleotides, cyclic nucleotides and deoxycyclic nucleotides by reversed-phase high-performance liquid chromatography.

A reversed-phase gradient elution system with methanol-triethylammonium phosphate buffer (83.3 mM, pH 6.0) as eluent on C(18)-bonded silica is described for the separation of 38 ribonucleotides, deoxynucleotides, cyclic nucleotides and deoxycyclic nucleotides in less than 33 min. The retention of the nucleotides can be precisely controlled by adjusting the pH, buffer concentration and methanol content in the mobile phase. The system is especially useful for the analysis of low levels of cyclic nucleotides in cells and tissues.

Journal Article↗
PubMed1985

The effects of ethanol and anticonvulsants on erythroid delta-aminolaevulinic acid synthase.

The activity of delta-aminolaevulinic acid (ALA) synthase was measured in bone marrow from 13 control subjects, 12 chronic alcoholic patients and 9 patients on long term anticonvulsant therapy. The majority of patients in both groups had macrocytic red cells in the absence of megaloblastic changes in the marrow. There was a significant increase in ALA synthase activity in the alcoholic patients but no significant increase in enzyme activity in the patients on anticonvulsants and overall there was no correlation of activity with MCV. The macrocytosis associated with these drugs does not therefore appear to result from accelerated erythroid haem synthesis.

5-Aminolevulinate Synthetase↗
PubMed1984

High-performance liquid chromatography of dicarboxylic porphyrins and metalloporphyrins: retention behaviour and biomedical applications.

The retention behaviour of the main dicarboxylic porphyrins, haemato-, deutero-, meso- and protoporphyrins, together with the Fe, Co, Cu and Zn complexes of meso- and protoporphyrins have been systematically studied. Hydrophobic chromatography with methanol-ammonium acetate buffer systems on reversed-phase columns provided the best selectivity, efficiency and resolution. The retention of the porphyrins is controlled by the relative hydrophobicity of the porphyrin side-chain substituents. The insertion of a metal ion into the porphyrin macrocycle, however, completely alters the electronic environment around the central nitrogen atoms of the porphyrins. The retention is then greatly influenced by the species of inserted metal ion, to accept axial ligands from the mobile phase, although hydrophobic interaction of the side-chain substituents with the stationary phase surface is still an important factor. The retention behaviour can be precisely controlled by adjusting the pH, buffer concentrations and types and proportions of organic solvents in the mobile phase. The analysis of porphyrins and metalloporphyrins in blood, and the determination of ferrochelatase in bone marrow are examples of biomedical applications.

Acetates↗
PubMed1984

Ammonium acetate: a general purpose buffer for clinical applications of high-performance liquid chromatography.

The chromatographic properties of ammonium acetate have been studied, and its use as a general purpose buffer for reversed-phase chromatography has been investigated. Simple ammonium acetate systems can often replace complicated and expensive buffers, with or without ion-pairing agents, with improved column selectivity and efficiency. A wide range of clinically important compounds have been successfully separated by ammonium acetate buffer systems. These include creatinine, bilirubin, verapamil and its metabolites, etoposide and teniposide, vitamin A, biogenic amines and porphyrins.

Acetates↗