scholarly journals One-electron oxidation pathway of thiols by peroxynitrite in biological fluids: bicarbonate and ascorbate promote the formation of albumin disulphide dimers in human blood plasma

1998 ◽  
Vol 329 (2) ◽  
pp. 405-413 ◽  
Author(s):  
Giuseppe SCORZA ◽  
Maurizio MINETTI
Keyword(s):  
Blood Plasma ◽  
Peptide Mapping ◽  
Biological Fluids ◽  
Protein S ◽  
Human Blood Plasma ◽  
Thiol Groups ◽  
Chain Reactions ◽  
Thiyl Radicals ◽  
Protein Thiols ◽  
The One

Recent studies have shown that peroxynitrite oxidizes thiol groups through competing one- and two-electron pathways. The two-electron pathway is mediated by the peroxynitrite anion and prevails quantitatively over the one-electron pathway, which is mediated by peroxynitrous acid or a reactive species derived from it. In CO2-containing fluids the oxidation of thiols might follow a different mechanism owing to the rapid formation of a different oxidant, the nitrosoperoxycarbonate anion (ONOOCO2-). Here we present evidence that in blood plasma peroxynitrite induces the formation of a disulphide cross-linked protein identified by immunological (anti-albumin antibodies) and biochemical criteria (peptide mapping) as a dimer of serum albumin. The albumin dimer did not form in plasma devoid of CO2 and its formation was enhanced by ascorbate. However, analysis of thiol groups showed that reconstituting dialysed plasma with NaHCO3 protected protein thiols against the oxidation mediated by peroxynitrite and that the simultaneouspresence of ascorbate provided further protection. Ascorbate alone did not protect thiol groups from peroxynitrite-mediated oxidation. ESR spin-trapping studies with N-t-butyl-α-phenylnitrone (PBN) revealed that peroxynitrite induced the formation of protein thiyl radicals and their intensity was markedly decreased by plasma dialysis and restored by reconstitution with NaHCO3. PBN completely inhibited the formation of albumin dimer. Moreover, the addition of iron-diethyldithiocarbamate to plasma demonstrated that peroxynitrite induced the formation of protein S-nitrosothiols and/or S-nitrothiols. Our results are consistent with the hypothesis that NaHCO3 favours the one-electron oxidation of thiols by peroxynitrite with formation of thiyl radicals, •NO2, and RSNOx. Thiyl radicals, in turn, are involved in chain reactions by which thiols are oxidized to disulphides.

scholarly journals Gas phase oxidants of cigarette smoke induce lipid peroxidation and changes in lipoprotein properties in human blood plasma. Protective effects of ascorbic acid

1991 ◽  
Vol 277 (1) ◽  
pp. 133-138 ◽  
Author(s):  
B Frei ◽  
T M Forte ◽  
B N Ames ◽  
C E Cross
Keyword(s):  
Lipid Peroxidation ◽  
Ascorbic Acid ◽  
Blood Plasma ◽  
Gas Phase ◽  
Cigarette Smoke ◽  
Human Blood ◽  
Biological Fluids ◽  
Human Blood Plasma ◽  
Lipid Hydroperoxides ◽  
Protein Thiols

Cigarette smoke (CS) is known to contain a large number of oxidants. In order to assess the oxidative effects of CS on biological fluids, we exposed human blood plasma to filtered (gas phase) and unfiltered (whole) CS, and determined the rate of utilization of endogenous antioxidants in relation to the appearance of lipid hydroperoxides. Lipid peroxidation was measured with a specific and sensitive assay that can detect lipid hydroperoxides at plasma levels as low as 10 nM. We found that exposure of plasma to the gas phase of CS, but not to whole CS, induces lipid peroxidation once endogenous ascorbic acid has been oxidized completely. In addition, CS exposure caused oxidation of plasma protein thiols and albumin-bound bilirubin, whereas uric acid and alpha-tocopherol were not consumed at significant rates. In plasma exposed to the gas phase of CS, low-density lipoprotein exhibited slightly increased electrophoretic mobility, but there was no apparent degradation of apolipoprotein B. Our results support the concept of an increased vitamin C utilization in smokers, and suggest that lipid peroxidation induced by oxidants present in the gas phase of CS leads to potentially atherogenic changes in lipoproteins.

scholarly journals One-electron oxidation pathway of peroxynitrite decomposition in human blood plasma: evidence for the formation of protein tryptophan-centred radicals

1997 ◽  
Vol 321 (3) ◽  
pp. 743-750 ◽  
Author(s):  
Donatella PIETRAFORTE ◽  
Maurizio MINETTI
Keyword(s):  
Blood Plasma ◽  
Human Blood ◽  
Structural Information ◽  
Biological Fluids ◽  
Nitroxide Radical ◽  
Amide Bond ◽  
Human Blood Plasma ◽  
Oxidation Pathway ◽  
Protein Adduct ◽  
Radical Adduct

Exposure of human blood plasma to peroxynitrite in the presence of 3,5-dibromo-4-nitrosobenzenesulphonic acid (DBNBS) resulted in the trapping of a strongly immobilized nitroxide radical adduct. The adduct was due to protein-centred radicals derived not only from serum albumin but also from other major plasma proteins (fibrinogen, IgG, α1-antitrypsin and transferrin). Urate significantly protected plasma from the peroxynitrite-induced DBNBSŐplasma protein adduct, whereas ascorbate and glutathione were protective at concentrations exceeding those usually found in plasma. Alkylation of plasma ŐSH groups did not affect the intensity of DBNBSŐplasma protein adduct, whereas bicarbonate increased its formation, thus showing a pro-oxidant effect. The DBNBSŐplasma protein adduct provided little structural information, but subsequent non-specific-protease treatment resulted in the detection of an isotropic three-line spectrum, indicating the trapping of radicals centred on a tertiary carbon. The nitrogen hyperfine coupling constant of this adduct and its superhyperfine structure were similar to those of DBNBSŐtryptophan peptides with the α-amino group of tryptophan linked in the amide bond, consistent with a radical adduct formed at C-3 of the indole ring of tryptophan-containing peptides. DBNBS was unable to trap radicals derived from peroxynitrite-treated tyrosine or tyrosine-containing peptides. Methionine treated with peroxynitrite resulted in the trapping of at least two DBNBSŐmethionine adducts with hyperfine structures different from that of protease-treated DBNBSŐplasma proteins. These results demonstrate that peroxynitrite induced in blood plasma the formation of protein radicals centred on tryptophan residues and underline the relevance of the one-electron oxidation pathway of peroxynitrite decomposition in biological fluids.

scholarly journals Determination of felodipine in biological fluids

2019 ◽  
Vol 100 (4) ◽  
pp. 650-656
Author(s):  
L L Kvachakhiya ◽  
V K Shormanov ◽  
N S Kononenko
Keyword(s):  
Blood Plasma ◽  
Biological Object ◽  
Biological Fluid ◽  
Biological Fluids ◽  
Human Blood Plasma ◽  
Gas Liquid Chromatography ◽  
Optimal Conditions ◽  
Mass Ratio ◽  
Layer Chromatography

Aim. Development of methods for the determination of felodipine in blood and plasma. Methods. The study object was felodipine [3-ethyl-5-methyl-4-(2,3-dichlorophenyl)-2,6-dimethyl-1,4-dihydropyridine-3,5-dicarboxylate]. The experiments were carried out on model mixtures of felodipine with blood and human blood plasma. Acetone was proposed as an isolating agent for the extraction of felodipine from biological fluids. To identify and quantify felodipine in extracts from blood and plasma, the methods of thin-layer chromatography, spectrophotometry and gas-liquid chromatography in combination with mass spectrometry were proposed. Results. The possibility of using acetone as an isolating agent to extract felodipine from biological fluids is demonstrated. The optimal conditions for the extraction of felodipine with acetone were found to be achieved already at 2-fold infusion of a biological object with an isolating agent, if the mass ratio of isolating liquid and biological material at each infusion stage is at least 2:1, and the infusion time is at least 30 minutes. Optimal felodipine purification conditions were achieved in a macrocolumn (15×1 cm) of Silasorb S-18 sorbent of 30 μm with elution of the substance with the polar eluent acetonitrile-water (7:3). The methods of determining felodipine in the blood and plasma were developed. With the content of felodipine of 25 mg in 25 g of biological fluid, the developed methods allow determining 86.01–87.86% in blood and 95.64–96.18% of the substance in blood plasma. The values of the detection limit of felodipine in the blood and plasma by the developed methods are 200 μg/100 g and 150 μg/100 g, respectively. Conclusion. Methods for the determination of felodipine in biological fluids were developed based on isolating with acetone and purification in the Silasorb S-18 sorbent column; use of these methods allows determining up to 87.86% of the analyte in the blood and up to 96.18% in the blood plasma.

Validation of Simvastatin Analysis Methods in Human Blood Plasma (In Vitro) Using HPLC-UV Detector

2018 ◽  
Vol 8 (2) ◽  
Author(s):  
Iyan Sopyan ◽  
Cynthia Jaya ◽  
Driyanti Rahayu
Keyword(s):  
Blood Plasma ◽  
Human Plasma ◽  
Disease Risk ◽  
Protein Precipitation ◽  
Precipitation Method ◽  
Human Blood Plasma ◽  
Analysis Method ◽  
Uv Detector ◽  
Number Of Patients ◽  
System Suitability

The use of simvastatin (SV) increases along with the increasing number of patients with hyperlipidemia and cardiovascular disease risk factors. Consequently, this condition leads to the increasing need of analytical determination of SV in blood plasma. Analysis of SV in human plasma using protein precipitation method and HPLC with UV detector has not been reported. This research was purpose to find out the rapid, accurate, and valid of SV analysis method in human plasma. In this research plasma samples were treated with protein precipitation method. The analyte was then analyzed using HPLC with C18 column 250x4 mm and 5 µm of particle size, the mobile phase contained of phosphate buffer 0.01 M (pH 4.0) and acetonitrile 30:70 v/v with flow rate 1 mL/minute, and detected at 239 nm. The analysis method was validated based on some parameters, such as selectivity, accuracy, precision, repeatability, linearity, LOD, LOQ, and system suitability. The result showed selectivity represented by Rs was 2.870, repeatability by its CV less than 2%, and linearity by its coefficient correlation (r) 0.9992 for concentration range 0.08-0.32 ppm. Based on chromatogram peak area, LOD and LOQ were 0.0132 and 0.0440 ppm respectively, accuracy and precision were 86.25-89.36% and 0.66-1.81% were obtained. The result of system suitability test from retention time and chromatogram peak area showed by its CV less than 2%. The analysis method was proved to be valid for SV analysis in human plasma

Dual Layer Mixed Matrix Hollow Fiber Membranes for Outside-In Filtration of Human Blood Plasma

2020 ◽  
Author(s):  
O.E.M. ter Beek ◽  
M.K. van Gelder ◽  
C. Lokhorst ◽  
D.H.M. Hazenbrink ◽  
B.H. Lentferink ◽  
...  
Keyword(s):  
Blood Plasma ◽  
Human Blood ◽  
Hollow Fiber ◽  
Human Blood Plasma ◽  
Hollow Fiber Membranes ◽  
Mixed Matrix ◽  
Fiber Membranes

Detectability of Plasma Proteins in SRM Measurements

2018 ◽  
Vol 16 (1) ◽  
pp. 74-81 ◽  
Author(s):  
Olga I. Kiseleva ◽  
Elena A. Ponomarenko ◽  
Yulia A. Romashova ◽  
Ekaterina V. Poverennaya ◽  
Andrey V. Lisitsa
Keyword(s):  
Blood Plasma ◽  
Human Plasma ◽  
Limit Of Detection ◽  
Human Blood Plasma ◽  
Analytical Sensitivity ◽  
Plasma Proteome ◽  
Protein Targets ◽  
Blood Plasma Sample ◽  
Specificity And Sensitivity ◽  
Human Plasma Proteome

Background: Liquid chromatography coupled with targeted mass spectrometry underwent rapid technical evolution during last years and has become widely used technology in clinical laboratories. It offers confident specificity and sensitivity superior to those of traditional immunoassays. However, due to controversial reports on reproducibility of SRM measurements, the prospects of clinical appliance of the method are worth discussing. </P><P> Objective: The study was aimed at assessment of capabilities of SRM to achieve a thorough assembly of the human plasma proteome. </P><P> Method: We examined set of 19 human blood plasma samples to measure 100 proteins, including FDA-approved biomarkers, via SRM-assay. </P><P> Results: Out of 100 target proteins 43 proteins were confidently detected in at least two blood plasma sample runs, 36 and 21 proteins were either not detected in any run or inconsistently detected, respectively. Empiric dependences on protein detectability were derived to predict the number of biological samples required to detect with certainty a diagnostically relevant quantum of the human plasma proteome. </P><P> Conclusion: The number of samples exponentially increases with an increase in the number of protein targets, while proportionally decreasing to the logarithm of the limit of detection. Analytical sensitivity and enormous proteome heterogeneity are major bottlenecks of the human proteome exploration.

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