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Biomedical subjects

S M Lunte

Publications and source records attributed to S M Lunte.

48 records · Page 3Linked to original sources
PubMed1993

Enzymatic formation and electrochemical characterization of multiply substituted glutathione conjugates of hydroquinone.

1,4-Benzoquinone can undergo redox cycling in the presence of glutathione to produce multiply substituted products. It has previously been shown that the nephrotoxicity of the hydroquinone-glutathione conjugates increases with increasing substitution. However, based on chromatographically-assisted hydrodynamic voltammetry (CA-HDV), the oxidation potential was shown to apparently increase which, should lead to decreased toxicity. From the chemical formation of multiply substituted hydroquinone-glutathione conjugates from benzoquinone and glutathione, it is clear that the thermodynamic oxidation potential must decrease as substitution increases. This was confirmed by cyclic voltammetric (CV) characterization of the isolated conjugates. The discrepancy between the CV and CA-HDV data apparently results from kinetic factors arising from differences in the treatment of the electrode surface between the two experiments. The multiply substituted hydroquinone-glutathione conjugates were also produced in horseradish peroxidase incubations containing hydroquinone and glutathione. These products were identified chromatographically, spectrophotometrically, and electrochemically. The increasing ease of oxidation and the possible enzymatic formation of multiply substituted hydroquinone-glutathione conjugates indicates that this pathway may occur in vivo and contribute to the toxicity of quinones.

Chromatography, Liquid↗
PubMed1992

Monitoring excitatory amino acid release in vivo by microdialysis with capillary electrophoresis-electrochemistry.

Capillary electrophoresis (CE) with electrochemical detection (ED) was used to determine extracellular levels of aspartate, glutamate and alanine in samples from the frontoparietal cortex of the rat which were obtained by microdialysis. The method was used to monitor the effect on the overflow of the excitatory amino acids aspartate and glutamate of an influx of high concentrations of potassium ion. Samples were derivatized with naphthalenedialdehyde-cyanide prior to analysis. Detection limits for aspartate and glutamate were 80 and 100 nM, respectively. CE-ED is extremely useful for the analysis of microdialysis samples because of the very small sample volumes required by this analytical technique. The use of ED provides the requisite sensitivity and allows verification of peak purity by voltammetry.

Amino Acids↗
PubMed1992

Electrochemical characteristics of amino acids and peptides derivatized with naphthalene-2,3-dicarboxaldehyde: pH effects and differences in oxidation potentials.

The electrochemical oxidations of the cyanobenz[f]isoindole (CBI) derivatives of 18 amino acids and 15 peptides, including enkephalins and several enkephalin fragments, were studied. Cyclic voltammetry indicated that the oxidation potentials of derivatized amino acids were virtually independent of pH. The utility of this pH-independence was demonstrated by controlling, through varying the pH, the selectivity with which CBI derivatives could be detected in the presence of phenolic compounds. In addition, hydrodynamic voltammograms of derivatized amino acids and peptides were constructed from chromatographic data and compared. The E1/2 values among the derivatized amino acids covered a range of 215 mV, with the derivatives of the basic amino acids being the easiest to oxidize and those of the acidic amino acids being the most difficult to oxidize. The E1/2 values of the derivatized peptides examined varied by 270 mV, with minor variations in structure capable of producing marked changes in oxidation potential. These results indicate that voltammetry can aid in identification or selective detection of CBI derivatives of amino acids or peptides.

Amino Acid Sequence↗
PubMed1990

Detection of N-acetylcysteine, cysteine and their disulfides in urine by liquid chromatography with a dual-electrode amperometric detector.

A method for the determination of N-acetylcysteine, cysteine and their disulfides in urine is described. The thiols and disulfides are separated by reversed-phase ion-pair chromatography with octyl sodium sulfate as the ion-pairing reagent and detected with a dual-electrode amperometric detector using Au/Hg amalgam electrodes. Both the thiols and disulfides are detected with this system. In addition, dimers and mixed disulfides can be detected individually.

Acetylcysteine↗
PubMed1990

Electroanalytical approaches to understanding benzene metabolism.

Electrochemical techniques are ideally suited to the study of the metabolism of aromatic xenobiotics because the metabolites are frequently easier to oxidize than the parent compounds. In many cases, the trace metabolites have the lowest oxidation potentials and hence electrochemical methods have the greatest selectivity for these compounds. The sensitivity of dual-electrode liquid chromatography-electrochemistry for the detection and identification of trace metabolites was demonstrated by the detection of the secondary metabolite, hydroquinone, in a microsomal incubation containing benzene and ascorbic acid. The use of an electrochemical detector in a series configuration provides increased selectivity for chemically reversible metabolites such as hydroquinone. Electrochemical methods can also be used to generate metabolites. The products of the electrochemical oxidation of phenol and biphenol are compared with those generated in a peroxidase incubation.

Animals↗
PubMed1989

Determination of desmosine, isodesmosine, and other amino acids by liquid chromatography with electrochemical detection following precolumn derivatization with naphthalenedialdehyde/cyanide.

Naphthalenedialdehyde (NDA) in the presence of cyanide (CN) reacts with primary amines to produce fluorescent cyano[f]benzoisoindole (CBI) derivatives. These derivatives have been shown to be substantially more stable than the corresponding o-phthalaldehyde derivatives. However, one drawback of this method is that compounds derivatized at more than one site exhibit quenching, precluding the use of fluorescence detection. The CBI derivatives have been found to be electroactive and are oxidized at a modest oxidation potential (+750 mV). Electrochemical detection is especially useful for the analysis of compounds containing more than one primary amine site because the response is not attenuated as it is in fluorescence detection. Desmosine and isodesmosine were of particular interest because of their importance in elastic fiber and the lack of highly sensitive HPLC methods for the determination of these compounds. Both of these compounds react with NDA/CN to produce electrochemically active derivatives. The combination of derivatization with NDA/CN and electrochemical detection was found to be linear over three orders of magnitude. Detection limits for CBI-lysine and CBI-desmosine were 100 fmol at a S/N of 2. Amino acids in elastin were quantitated using this method. The results correlate well with what has been reported previously in the literature. A significant advantage of the use of liquid chromatography with electrochemical detection with precolumn derivatization with NDA/CN for the analysis of desmosine and isodesmosine is that they can be separated and quantitated individually using this method. In addition, the unique voltammetry of multiderivatized CBI-amino acids can be used to verify peak purity.

Aldehydes↗
PubMed1987

Structural classification of flavonoids in beverages by liquid chromatography with ultraviolet-visible and electrochemical detection.

Liquid chromatography with both UV-VIS and electrochemical detection is used to structurally classify flavonoid compounds in wine and grape juice without isolation of the pure compound. Compounds are classified as flavonols, proanthocyanidins, or anthocyanidins, based on their absorption maxima. Catechol substituted compounds are identified using a dual electrode detector.

Beverages↗
PubMed1984

The use of liquid chromatography with dual-electrode electrochemical detection in the investigation of glutathione oxidation during benzene metabolism.

Liquid chromatography with electrochemical detection provides a powerful tool for the study of glutathione oxidation during benzene metabolism. The use of two dual-electrode detectors allows for the detection of oxidized and reduced glutathione and of phenol and quinone metabolites of benzene. The role of glutathione as a reductant is explored in this paper. Results indicate that hydrogen peroxide is the oxidizing agent primarily responsible for glutathione oxidation during benzene, phenol and hydroquinone metabolism.

Animals↗
PubMed1983

Detection and identification of sulfhydryl conjugates of rho-benzoquinone in microsomal incubations of benzene and phenol.

The glutathione and cysteine conjugates of rho-benzoquinone are detected and conclusively identified in microsomal incubations of benzene and phenol using liquid chromatography/electrochemistry (LCEC). Identification of the compounds is based on retention time, electrochemical behavior and acid hydrolysis. The fact that both of these compounds can be detected easily in a benzene incubation provides further evidence that rho-benzoquinone or the corresponding semiquinone is a product of benzene metabolism in vivo. The conjugation of rho-benzoquinone with glutathione is predominantly a non-enzymatic process. This is illustrated by the fact that the addition of cytosolic glutathione-S-transferases do not significantly increase the amount of glutathione conjugate produced in a phenol incubation containing glutathione. The kinetic constants for phenol metabolism to hydroquinone by microsomal protein are calculated. As suspected, the rate of metabolism of phenol is significantly higher than the rate of benzene metabolism. The Vmax for phenol metabolism was calculated to be 7.1 nmol/min/mg protein and the KM was found to be 0.38 mM. The further oxidation of hydroquinone to rho-benzoquinone appears to be primarily an enzymatic process. Incubations of just hydroquinone with glutathione at 37 degrees C produced only a small amount of the glutathione conjugate. The addition of cytosolic protein increases the amount of rho-benzoquinone produced about 10-fold. This could be due to the peroxidases found in that medium. The addition of microsomal protein and NADPH increases the amount of glutathione conjugate produced to over 100-fold of that produced non-enzymatically. This indicates that a microsomal enzyme is responsible for the oxidation of hydroquinone to rho-benzoquinone in vitro and the subsequent covalent binding to macromolecules.

Animals↗
PubMedDate not supplied

Fluorescence and chemiluminescence detection of oxazole-labelled amines and thiols.

Fluorescence and chemiluminescence analyses of amino acids and thiols derivatized with 2-fluoro-4,5-diphenyloxazole (DIFOX) and 2-chloro-4,5-bis(p-N,N-dimethylaminosulphonylphenyl)oxazole (SAOX-Cl) were investigated. Thirteen diphenyloxazole (DIOX)-derivatized amino acids were separated within 38 min by a linear gradient elution from 100% A [0.05 M phosphate (pH 7.0): CH3CN (75:25)] to 100% B [0.05 M phosphate (pH 7.0):CH3CN (1:1)] over 30 min and an isocratic elution of 100% B for 30 min. The detection limits (S/N = 2) with fluorescence detection were in the range of 19-64 fmol. Thiols derivatized with SAOX-Cl were separated by an isocratic elution using 0.1 M H3PO4:CH3CN (65:35) and detected fluorimetrically. The detection limits (S/N = 2) of reduced glutathione, N-acetylcysteine, 2-mercaptopropionylglycine, cysteine, homocysteine and captopril were 1.2, 1.5, 1.9, 5.7, 6.4 and 7.9 fmol, respectively. Peroxyoxalate chemiluminescence (CL) intensities of sulphonyl-5-N,N-dimethylaminonaphthalene (DNS), SAOX and DIOX derivatives were compared using three different oxalate esters (DFPO, TCPO and TDPO) by flow injection analysis. The relative chemiluminescence intensity (RCL) of SAOX-proline and DIOX-proline were 76-80% and 19-25% of DNS-proline (100%), respectively. Other SAOX and DIOX derivatives showed lower CL intensities (< 12%). Extremely low CL intensities were obtained for the fluorescent tagging reagents (< 0.11%) and their hydrolysis products (< 0.80%). Secondary amino acids and peptides, derivatized with DIFOX in aqueous media at room temperature for 1 h, were detected using DFPO/H2O2. TCPO/H2O2 and TDPO/H2O2 after separation by high performance liquid chromatography.(ABSTRACT TRUNCATED AT 250 WORDS)

Amines↗