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1.
Gallo P Nasi A Vinci F Fiori M Brambilla G Serpe L 《Analytical and bioanalytical chemistry》2008,391(4):1153-1162
Bovine α1-acid glycoprotein (bAAG) and bovine serum albumin (BSA) are plasmatic acceptors working as carriers by the specific and reversible
binding of several drugs in vivo. We synthesized affinity columns by coupling bAAG and BSA to an activated chromatographic
support through their carbohydrate moieties, to preserve protein tertiary structure and, consequently, to improve the biological
activity in vitro. The bAAG and BSA affinity columns were used to study the binding of acidic and basic drugs. Moreover, a
purification strategy was developed for the cleanup of drug residues from biological matrices and foods, prior to screening
and/or confirmatory analysis, on the basis of the specific molecular recognition between the protein and the drug. The aim
of this work was to test the potency of bAAG- and BSA-based affinity chromatography to bind some veterinary drugs and purify
them in the context of the official control of animal products. The efficiency of these homemade affinity columns in minimising
matrix interference and in selective cleanup of different classes of substances was reported and discussed.
Figure “Coupling strategies in synthesizing plasmatic acceptor affinity columns: the covalent coupling of bAAG and BSA through their
carbohydrate moiety allows one to preserve the gross tertiary structure of the protein and thus its biological activity, whereas
coupling through the ε-amine group of lysine residues can reduce the interactions with the binding sites of the plasmatic
acceptor.” 相似文献
2.
New fast methods for the determination of pharmacokinetic behaviour of potential drug candidates are receiving increasing
interest. We present a new homogeneous method for the determination of drug binding and drug competition for human serum albumin
and α1-acid glycoprotein that is amenable to high-throughput-screening. It is based on selective fluorescent probes and the measurement
of fluorescence polarization. This leads to decreased interference with fluorescent drugs as compared with previously published
methods based on similar probes and the measurement of fluorescence intensity. The binding of highly fluorescent drugs that
still interfere with the probes can be measured by simply titrating the drugs in a two-component system with the serum protein.
The assay may also be used to discover strongly binding protein ligands that are interesting for drug-targeting strategies.
Additionally, binding data could be obtained from larger libraries of compounds for in silico predictive pharmacokinetics.
Figure Fluorescence polarization displacement titration of dansylsarcosine (3D-structure as insert) bound to human serum albumin
(HSA) by naproxene
Electronic supplementary material The online version of this article (doi:) contains supplementary material, which is available to authorized users. 相似文献
3.
The effect of ethanol on human serum albumin stability in aqueous solution
was studied by use of differential scanning calorimetry. A deconvolution of
DSC traces in 2-state model with ΔC
p=0
and ΔC
p≠0
was performed and analysed to obtain information on the interaction of ethanol
with different parts of albumin molecule both fatty acid containing and fatty
acid free. The differences in ethanol binding affinity for both kinds of albumin
were found. At very low concentrations ethanol was observed to be a stabilizer
of the folded state of albumin contrary to the higher concentration where
its binding to the unfolded protein predominates. 相似文献
4.
Sabela MI Gumede NJ Escuder-Gilabert L Martín-Biosca Y Bisetty K Medina-Hernández MJ Sagrado S 《Analytical and bioanalytical chemistry》2012,402(5):1899-1909
The stereoselective binding of the frequently ingested nutraceutical (±)-catechin, with demonstrated differential biological
activity between enantiomers, to human serum albumin (HSA), with the largest complexation and enantioselectivity potential
among the plasmatic proteins, is studied by combining simulations to optimize the experimental design, robust in vitro electrokinetic
chromatographic data, and molecular docking–chiral recognition estimates. Methodological and mathematical drawbacks in previous
reports on (±)-catechin–HSA are detected and eliminated. Recent and novel direct equations extracted from the classical interaction
model allows advantageous univariate mathematical data treatment, providing the first evidence of quantitative (±)-catechin–HSA
enantioselectivity. Also, the binding site in HSA of the enantiomers is approached, and both the experimental enantioselectivity
and the main binding site information are contrasted with a molecular docking approach. 相似文献
5.
Palacios O Ruiz Encinar J Schaumlöffel D Lobinski R 《Analytical and bioanalytical chemistry》2006,384(6):1276-1283
Immunoaffinity chromatography has been investigated for fractionation of serum into selenoalbumin and true selenoproteins.
Among several albumin-depletion kits tested, a multiaffinity column specifically binding albumin and five other major serum
proteins provided the best results. It extracted ca 95% of both albumin and selenoalbumin, which enabled interference-free
determination of glutathione peroxidase, selenoprotein P, and selenoalbumin by size-exclusion chromatography combined with
inductively coupled plasma mass spectrometry (SEC–ICPMS). The efficiency of the multiaffinity column did not vary over a period
of 18 months. The purity of fractions separated by immunoaffinity LC was confirmed by elution-volume matching with standards
in SEC–ICPMS and by selenopeptide mapping in capillary HPLC–ICPMS. Quantification of the selenium distribution among the different
proteins in human serum from a control group and from a person on a selenium-rich diet revealed that 67% of the supplemented
selenium was incorporated into albumin, 30% into glutathione peroxidase, and 3% into selenoprotein P. 相似文献
6.
The application of near-infrared (NIR) dyes (λ
em > 750 nm) to the analysis of biological samples shows much promise, because the long emission wavelengths of such dyes allow
interferences from biomolecule matrices to be minimized. In this paper, a novel NIR dye, 5,5′-dicarboxy-1,1′-disulfobutyl-3,3,3′,3′-tetramethylindotricarbocyanine
(DCDSTCY) has been developed for the spectrophotometric determination of total protein in serum. Under acidic conditions,
the binding of DCDSTCY to proteins caused a new peak at 878 nm, the height of which was proportional to the concentration
of protein. The linear range of the method was found to be 0.04–0.5 μg mL−1 for bovine serum albumin (BSA) and human serum albumin (HSA), and detection limits of 5 ng mL−1 were obtained for these substances. The maximum binding number of BSA with DCDSTCY was measured to be 133. The method proposed
here has been applied to the quantitation of total protein in serum, and recoveries of 96.6–104% were achieved.
Figure Near-infrared probe for protein determination 相似文献
7.
《Analytical letters》2012,45(9):1143-1155
Abstract An antibody IgG-coated polystyrene ball was incubated with an antigen and then with affinity-purified Fab'-enzyme conjugate in the presence of normal IgG, F(ab')2, Fab' or Fab'-bovine serum albumin conjugate. After washing by incubation at 30[ddot]C for 10 min with shaking, the enzyme activity bound to the polystyrene ball was assayed. The non-specific binding of the Fab'-enzyme conjugate to the polystyrene ball considerably decreased in the presence of normal IgG and the other related proteins, while the specific binding decreased only slightly. As a result, the detection limit of hCG, human IgE and human α-fetoprotein was improved 3 to 10-fold. 相似文献
8.
A. Kathiravan R. Renganathan 《Colloids and surfaces. A, Physicochemical and engineering aspects》2008,324(1-3):176-180
The interaction between colloidal TiO2 and bovine serum albumin (BSA) was studied by using absorption and fluorescence spectroscopic methods. The quenching of the intrinsic protein fluorescence in the presence of different concentrations of colloidal TiO2 was analyzed and number of binding sites (n) and apparent binding constant (K) were measured. The quenching mechanism of albumin by colloidal TiO2 is discussed. The energy transfer efficiency (E) and critical transfer distance (R0) were determined. 相似文献
9.
Capillary electrophoresis frontal analysis was applied to 12 low molecular weight compounds including 8 drug substances displaying a range of different properties with respect to binding affinity, binding location, structure, lipophilicity, charge at physiological pH, and electrophoretic mobility. It was found that capillary electrophoresis frontal analysis can be used as a general method to study and quantify drug-human serum albumin interactions. The binding parameters obtained were consistent with literature values. Dextran was in some cases added to the run buffer to improve separation of the drug and human serum albumin plateau peaks. Results indicate that mobility differences between free and complexed human serum albumin give rise to only minor errors. Capillary electrophoresis frontal analysis was also found applicable to the study of human serum albumin drug displacement reactions. Low sensitivity of the UV-detection system was found to be the major limitation of capillary electrophoresis frontal analysis. The method is simple, and minimal effort has to be put into method development, which makes it well suited for screening in early drug development. 相似文献
10.
Dan Wu Guiying Xu Yujun Feng Yajing Wang Yanyan Zhu 《Colloid and polymer science》2009,287(2):225-230
The interaction between bovine serum albumin (BSA) with N, N′-bis(dimethylalkyl) ethylammonium dibromide (C12C2Cm, m = 8, 12) was investigated by spectral methods. It can be seen that C12C2C8 and C12C2C12 mainly interact with tryptophan residues of BSA from synchronous fluorescence spectra. Fluorescence, far-UV, and near-UV
circular dichrosim spectra of BSA are changed by addition of dissymmetric and symmetric gemini surfactant. For surfactant
solution, the polarity of the microenvironment surrounding pyrene is lower while the fluorescence lifetime of it is longer
and the microviscosity is higher in the presence of BSA than those in the absence of BSA. But compared with C12C2C12, C12C2C8 has lower binding ability with BSA due to the shorter hydrophobic tail and lower symmetry. 相似文献
11.
Xie R Johnson W Rodriguez L Gounder M Hall GS Buckley B 《Analytical and bioanalytical chemistry》2007,387(8):2815-2822
To study the carboplatin–protein interaction, a sensitive method using size exclusion chromatography coupled to inductively
coupled plasma mass spectrometry (SEC–ICP–MS) was developed. The complexes formed between plasma proteins and carboplatin
were monitored and identified with this method. Composite blood plasma samples from patients who were undergoing chemotherapy
were analyzed, and carboplatin was found to bind plasma proteins. In addition, blank plasma samples were spiked with carboplatin
and were analyzed as a time course study, and the results confirmed that carboplatin formed complexes with plasma proteins,
primarily albumin and γ-globulin. To further substantiate the study, these two proteins were incubated with carboplatin. The
binding between carboplatin and these proteins was then characterized qualitatively and quantitatively. In addition to a one-to-one
binding of Pt to protein, protein aggregation was observed. The kinetics of the binding process of carboplatin to albumin
and γ-globulin was also studied. The initial reaction rate constant of carboplatin binding to albumin was determined to be
0.74 M−1 min−1, while that for γ-globulin was 1.01 M−1 min−1, which are both lower than the rate constant of the cisplatin–albumin reaction previously reported. 相似文献
12.
Quality testing of human albumin by capillary electrophoresis using thermally cross‐linked poly(vinyl pyrrolidone)‐coated fused‐silica capillary 下载免费PDF全文
To detect the quality of medicinal human albumin by capillary electrophoresis, we produced a fused‐silica capillary coated with thermally cross‐linked poly(vinyl pyrrolidone) to prohibit protein adsorption. This type of capillary was easily obtained by injecting an aqueous poly(vinyl pyrrolidone) solution into a fused‐silica capillary and thermally annealing it at 200°C. Notably, stable and low electro‐osmotic flow was obtained in the poly(vinyl pyrrolidone)‐coated capillary at pH 2.20–9.00, and the separation of a mixture of four basic proteins indicated that the poly(vinyl pyrrolidone)‐coated capillary exhibits excellent repeatability and separation efficiency; moreover, the separation of these four basic proteins could even be achieved at pH 7.00. The protein recovery percentage of human serum albumin in a single‐protein solution and a mixed blood proteins solution was determined to be 97.03 and 95.40% in the poly(vinyl pyrrolidone)50–3 (representing the concentration of the capillary‐injected poly(vinyl pyrrolidone) aqueous solution, 50 mg/mL, and thermal annealing time, 3 h) capillary, respectively. Based on these results, we used the poly(vinyl pyrrolidone)50–3‐coated capillary to quantify the protein content of human albumin, and the results obtained from run to run, day to day and capillary to capillary demonstrated that the coated capillary could be used for quality testing commercially available human albumin. 相似文献
13.
Determination of trace proteins with pyronine Y and SDS by resonance light scattering 总被引:1,自引:0,他引:1
A new resonance light scattering (RLS) probe for determining proteins is presented. The weak RLS of pyronine Y–SDS can be
enhanced substantially by adding proteins in the presence of H2SO4, resulting in a strong and wide RLS band in the region 310–425 nm. The interaction of pyronine Y–SDS with proteins was studied
on the basis of this behavior and a new quantitative method was developed for determining proteins. The enhanced RLS intensity
is proportional to the concentration of proteins in the range 0.15–3.6 μg mL−1 for bovine serum albumin (BSA) and 0.06–4.8 μg mL−1 for human serum albumin (HSA), with detection limits of 21.0 and 12.0 ng mL−1, respectively. This method is characterized by high sensitivity, rapidity of reaction, and simplicity. Four synthetic samples
were determined satisfactorily and recovery was 99.5–101.5%. Results for human serum and urine samples were in agreement with
those obtained by the Bradford method, with relative standard deviations (RSD) of 1.5–3.1%. 相似文献
14.
Summary Separation of the enantiomers of a variety of basic drugs by affinity capillary electrophoresis has been investigated using
α1-acid glycoprotein (α1-AGP) as chiral selector. In order to use a high concentration of α1-AGP without causing low detection sensitivity, the partial filling technique was employed. Enantiomer separations were performed
under conditions (a running buffer at pH 5.0 or 6.0) causing the protein to migrate toward the injection end. Twenty nine
basic racemates were successfully separated by optimizing the protein concentration, buffer pH and organic modifier. α1-AGP obtained from three different suppliers was used to investigate differences among the proteins from different sources.
Although most of the racemates were similarly separated with any of the three types of α1-AGP, some racemates, e.g. acebutolol behaved differently with the three types. The reasons for the different enantioselectivities
of the three types of α1-AGP has not yet been clarified. The method was used to test the optical purity of commercial sulpiride enantiomers and it
was found that the method was suitable and applicable for the purpose. 相似文献
15.
Summary The interaction thermodynamics of heptacarboxylporphyrin (HCP) and protoporhyrin (PP) with human serum albumin (HSA) was studied
by affinity capillary electrophoresis (ACE) over the temperature range of 25–50°C, where HCP and PP bound to HSAvia 1:1 molecular association. The binding equilibrium constants (pH 7.4, phosphate buffer) for the binding of HCP with HSA were
found to decrease with an increase in temperature, whereas the binding constants of the PP/HSA system appeared to be independent
of temperature changes over the range studied. The van’t Hoff relationship (25–50°C) was found to be linear for the interaction
of either HCP or PP with HSA. However, the interaction thermodynamics for both of these porphyrins with HSA were found to
be quite different. In particular, the interaction of HCP (a hydrophilic porphyrin) with HSA appeared to be based on an enthalpy-driven
process, whereas the binding between PP (a hydrophobic porphyrin) and HSA driven by a favorable change in entropy. The ability
of using ACE to evaluate the interaction thermodynamics of serum proteins (e.g., HSA) with ligands (e.g., porphyrins and related
compounds) should aid in the development of new and more effective photosensitizers in the photodynamic therapy of cancer. 相似文献
16.
Summary Affinity chromatography with α1-acid glycoprotein (AGP) and human serum albumin (HSA) stationary phases was applied to screen and analyze the biologically
active components ofRhizoma chuanxiong. Five major peaks and a number of small peaks were resolved based on their affinity for AGP and HSA, respectively, and three
of them, identified as ferulic acid, chuanxiongzine and ligustilide via standard compounds, are regarded as effective components.
The effects of acetonitrile concentration, pH, inorganic salt concentration and temperature on the retention behaviors of
five major components on the two stationary phases were also investigated. It was observed that hydrophobicity is the major
contributor to retention on both stationary phases, and ferulic acid has a weak electrostatic interaction with HSA. It demonstrated
that the chromatograms ofRhizoma chuanxiong on the two stationary phases have concise and different fingerprinting characteristics. The amount of ferulic acid, chuanxiongzine
and ligustilide inRhizoma chuanxiong determined using the AGP column are as much as 0.064%, 0.021% and 2.00% by weight. 相似文献
17.
Maciążek-Jurczyk M Sułkowska A Bojko B Równicka-Zubik J Sułkowski WW 《Spectrochimica acta. Part A, Molecular and biomolecular spectroscopy》2011,82(1):181-190
Interaction of phenylbutazone (PBZ) and aspirin (ASA), two drugs recommended in rheumatoid diseases (RDs), when binding to human (HSA) and bovine (BSA) serum albumins, has been studied by quenching of fluorescence and proton nuclear magnetic resonance (1HNMR) techniques.On the basis of spectrofluorescence measurements high affinity binding sites of PBZ and ASA on albumin as well as their interaction within the binding sites were described. A low affinity binding site has been studied by proton nuclear magnetic resonance spectroscopy.Using fluorescence spectroscopy the location of binding site in serum albumin (SA) for PBZ and ASA was found. Association constants Ka were determined for binary (i.e. PBZ–SA and ASA–SA) and ternary complexes (i.e. PBZ–[ASA]–SA and ASA–[PBZ]–SA).PBZ and ASA change the affinity of each other to the binding site in serum albumin (SA). The presence of ASA causes the increase of association constants KaI of PBZ–SA complex. Similarly, PBZ influences KaI of ASA–SA complex. This phenomenon shows that the strength of binding and the stability of the complexes increase in the presence of the second drug. The decrease of KaII values suggests that the competition between PBZ and ASA in binding to serum albumin in the second class of binding sites occurs. The analysis of 1HNMR spectral parameters i.e. changes of chemical shifts and relaxation times of the drug indicate that the presence of ASA weakens the interaction of PBZ with albumin. Similarly PBZ weakens the interaction of ASA with albumin. This conclusion points to the necessity of using a monitoring therapy owning to the possible increase of uncontrolled toxic effects. 相似文献
18.
Enantiomer separation of drugs by capillary electrophoresis using proteins as chiral selectors 总被引:6,自引:0,他引:6
Haginaka J 《Journal of chromatography. A》2000,875(1-2):235-254
The separation of drug enantiomers using proteins as the chiral selectors in capillary electrophoresis (CE) is considered in this review. The proteins used include albumins such as bovine serum albumin, human serum albumin and serum albumins from other species, glycoproteins such as alpha1-acid glycoprotein, crude ovomucoid, ovoglycoprotein, avidin and riboflavin binding protein, enzymes such as fungal cellulase, cellobiohydrolase I, pepsin and lysozyme and other proteins such as casein, human serum transferrin and ovotransferrin. Protein-based CE is carried out in two modes: in one proteins are immobilized or adsorbed within the capillary, or protein-immobilized silica gels are packed into the capillary (affinity capillary electrochromatography mode), and in the other proteins are dissolved in the running buffer (affinity CE mode). Furthermore, the advantages and limitations of the two modes and the factors affecting the chiral separations of various drugs by protein-based CE are discussed. 相似文献
19.
Margaret Galazka Michelle Tang Vincent A. DeBari Joseph D. Kohles Jung K. Lee Lynn B. Keil Matthew Petersheim 《Applied biochemistry and biotechnology》1999,76(1):1-13
Autoantibodies from patients with antiphospholipid syndrome (APS) recognize an epitope on β2glycoprotein I (β2GPI) only when native β2GPI is adsorbed on surfaces composed of anionic phospholipids or oxidized polystyrene, β2GPI was modified with the crosslinking agent, glutardialdehyde (GDA), which induced exposure of the anti-β2GPI epitope at GDA: β2GPI mol ratios in the range of 500–2000. A second crosslinking agent, dimethyl-suberimidate (DMS), did not expose the epitope,
which may be a consequence of its having less tendency than GDA to form intermolecular links. SDS-PAGE experiments demonstrate
that GDA does promote extensive intermolecular crosslinking of β2GPI, and DMS does not. Formaldehyde also reacts with the lysine residues of β2GPI, but does not expose the epitope. The circular dichroism spectra of native and modified β2GPI confirm that GDA induces changes in conformation that are qualitatively different from those caused by formaldehyde. These
data provide evidence that binding of lysine residues is not a sufficient condition for exposure of the autoepitope, and also
support the likelihood that β2GPI antibodies bind only to aggregates of the protein. Thus, by synthesizing an active holoantigen of β2GPI, conditions were defined that are necessary for binding of human autoantibodies. The authors also suggest that treatment
of phospholipid-binding proteins with chemical agents might provide a strategy to modify their structure and permit exposure
of epitopes, resulting in synthetic antigens for therapeutic and diagnostic use. 相似文献
20.
Antony F. McDonagh Giovanni Agati David A. Lightner 《Monatshefte für Chemie / Chemical Monthly》1998,129(6-7):649-660
Summary. Quantum yields for Z⇄E photoisomerization of the natural bi-chromophore bilirubin IXα in ammoniacal methanol solution were found to vary with excitation
wavelength, whereas no variation (within experimental error) was observed for xanthobilirubic acid and for symmetrically substituted
bilirubins (bilirubin IIIα, bilirubin XIIIα, and mesobilirubin XIIIα) in the same solvent. The quantum yield for Z⇄E photoisomerization of xanthobilirubic acid bound to human serum albumin was also invariant with excitation wavelength. In
contrast, quantum yields for Z⇄E photoisomerization of the symmetrically-substituted bilirubins did show marked excitation wavelength dependency when they
were bound to human serum albumin. These results show that quantum yields for Z⇄E photoisomerization of bilirubins are markedly sensitive to protein binding and to the nature of pyrrole ring β-substituents.
They also demonstrate that wavelength-dependent photochemistry is characteristic of bilirubins with non-identical pyrromethenone
chromophores, as expected from exciton coupling theory, and that complexation with albumin induces wavelength-dependent photochemistry
in symmetrically substituted bilirubins.
Received January 13, 1998. Accepted January 27, 1998 相似文献