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Abstract  

The paper presents the method of preparation and determination of physicochemical properties of silica gels with a chemically bonded BSA phase as well as studies of the effect of support porosity on the synthesis. Wide-porous Z-300 and narrow-porous Z-100 silica gels were studied. The investigations showed a significant effect of pore size on the synthesis of stationary phases with BSA. Modification with protein results in changes of adsorption properties and porosity of adsorbent samples. Changes of physicochemical properties result in significant changes of geometrical and structural heterogeneity of the support (specific surface area, fractal coefficient) as well as energetic heterogeneity of the samples.

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Abstract  

The kinetics of bovine serum albumin (BSA) denaturation in the absence and the presence of urea was studied by the iso-conversional method and the master plots method using differential scanning calorimetry (DSC). The observed denaturation process was irreversible and approximately conformed to the simple order reaction, and the denaturation did not follow rigorously first-order kinetic model or other integral order reaction models. The denaturation temperature (T m), apparent activation energy (E a), approximate order of reaction (n), and pre-exponential factor (A) all distinctly decreased as the 2 mol L−1 urea was added, which indicated that the urea accelerated the denaturation process of BSA and greatly reduced thermal and kinetic stability of BSA. This study also demonstrated that the iso-conversional method, in combination with the master plots method, provides a valuable and useful approach to the study of the kinetic process of protein denaturation.

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The effects of the mycotoxin patulin on the thermodynamics and kinetics of the transition of bovine serum albumin (BSA) in aqueous solution were studied by Differential Scanning Calorimetry and Photoluminescence methods. Results show that in the presence of patulin, the free enthalpy change during the transition of BSA was decreased by an average of ∼ 46 kJ/mol, the free energy change was decreased by ∼ 4 kJ/mol, and the activation energy fell from ∼ 1546 to ∼ 840 kJ/mol. These results indicate that the bioactivity of patulin is based on the kinetic rather than on the thermodynamic properties of the transition. This is the first evidence of the direct interaction of patulin with the free thiol-containing BSA, a process which could contribute to the adverse cyto- and genotoxic effects induced by patulin.

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Abstract  

With 2,3,5,6-tetrafluorophenyl 3-(nodo-carboranyl) propionate (TCP) as a new potential bi-functional linker, bovine serum albumin (BSA) was conjugated with 211At, and the conjugated model protein (211At-TCP-BSA) was preliminarily evaluated in vitro and in vivo by comparison with 211At-SAB-BSA and 211At-SAPC-BSA, which conjugated with 211At via aryl derivatives ATE (N-succinimidyl-3-(tri-n-butylstannyl) benzoate) or SPC (N-succinimidyl 5-(tributylstannyl)-3-pyridinecarboxylate). The radiolabeled intermediate 211At-TCP was coupled to BSA in yields ranging from 35 to 45% with radiochemical purity of more than 98%. The conjugated 211At-TCP-BSA exhibited considerable stability in vitro in 0.1 mol/L PBS (pH 7.6) at room temperature (RT), similar to 211At-SAPC-BSA and 211At-SAB-BSA. Biodistribution of the 211At conjugated protein was investigated in NIH strain mice by I.V injection. The results showed that 211At-TCP-BSA was constantly stable in vivo as well as in vitro, but more stable than 211At-SAPC-BSA and 211At-SAB-BSA. These results implied that radioastatinated carboranes based on B–At bonds are higher stability than radioastatinated aryl derivatives based on C–At to in vivo deastatination. In other word, TCP should be a promising bi-functional linker for 211At conjugation of proteins or antibodies.

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Abstract  

Monte Carlo simulations were carried out to calculate optimum design parameters of an accelerator based Beam Shaping Assembly (BSA) for Boron Neutron Capture Therapy (BNCT) of Brain Cancer setup. Epithermal beam of neutrons were obtained through moderation fast neutrons from 7Li(p,n) reaction in a high density polyethylene moderator and a graphite reflector. The dimensions of the moderator and the reflector were optimized through optimization of epithermal/fast neutron intensity ratio as a function of geometric parameters of the setup. Results of our calculation showed the capability of our setup to treat the tumor within 4 cm of the head surface. The calculated Peak Therapeutic Ratio for the setup was found to be 2.15. With further improvement in the polyethylene moderator design and brain phantom irradiation-arrangement, the setup capabilities can be improved to reach further deep-seated tumor.

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This paper presents investigations of phase and structural transitions occurring in water adsorbed on the surface of bovine serum albumin (BSA) and on the so-called ‘intelligentrs or ‘smart’ silica gel surface covered with a chemically bonded BSA phase. Cyclic changes of heat flow (HF) were observed in the samples studied during cooling and heating of the measuring cell of the differential scanning calorimetry (DSC) apparatus. These cyclic changes reflect structural transitions occurring in the water adsorbed on the surface at subambient and elevated temperatures. This is connected with cyclic changes (decay and reproduction) of ice-like structures existing in the adsorbed water layers. On the basis of quantitative investigations it appears that, depending on the direction of the cooling or heating process of the samples studied, the number of ice-like water structures in the surface film increases or decreases. It has been stated that the observed fluctuations occur spontaneously and suddenly in the whole volume of adsorbed water in different and not regular temperature ranges, especially at the ‘paradoxical effect’ temperatures.

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Abstract  

Bovine serum albumin (BSA) is a soft globular protein that undergoes conformational changes through several identified transition steps in the pH range 2–13.5. The ability to change conformation makes BSA capable of complexing different ligands from fatty acids to cations or drugs and carries them in the bloodstream. Microcalorimetric titration of BSA with NaOH solution was performed to measure the enthalpy of conformational changes. Two exothermic enthalpy changes were found in the course of the titration between pH 3 and 9.5, which can be identified with the E–F, and the F–N transitions. The enthalpy change at pH 3.5 (transition from the E to the F form of BSA, folding of intra-domain helices in domain I) is independent of the protein concentration. The second transition (F–N, folding of domain III) was observed at pH 4.8 for the 0.1% BSA solution, but it shifted to higher pH values as the protein concentration increased to 0.2% and 0.3%. The tightening of the protein structure with increasing pH was verified measuring intrinsic fluorescence of tryptophan residues. At even higher pH value, pH 10.5, fluorescence measurements revealed protein expansion. The BSA conformational changes were also measured by dynamic light scattering. The hydrodynamic diameter was smaller at the i.e.p. of BSA (5–7 nm at pH ~5) and larger at the two ends of the pH range (17.5 nm at pH 2 and 8.3 nm at pH 10).

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JPC - Journal of Planar Chromatography - Modern TLC
Authors: Mihaela Vlassa, Virginia Coman, Miuţa Filip, Florina Copaciu, Aurora Mocanu, and Maria Tomoaia-Cotişel

The basic of all proteins is constituted by approximately 20 common amino acids that differ only in the structure of their side chain. The purpose of our work was to use the overpressured layer chromatography (OPLC) technique for the separation and identification of the essential and non-essential amino acids from some most common proteins, like hen egg yolk protein, bovine serum albumin (BSA), and Type I Collagen from bovine achilles tendon. In this order, the target proteins were acid-hydrolyzed and derivatized. For the separation of essential amino acids from egg yolk, two successive OPLC developments (normal and overrunning) were performed, followed by direct derivatization with 4-hydroxybenzaldehyde and ninhydrin on the chromatoplate for identification. In the hydrolyzed egg yolk, nine essential amino acids were identified. Between lysine and histidine, a poor separation was obtained. The amino acids from BSA and Type I Collagen were separated and identified as phenylthiohydantoin (PTH) derivatives by OPLC. The supposed identity of the amino acids as PTH derivatives was confirmed by reversed-phase high-performance liquid chromatography (RP-HPLC). Seven essential amino acids (arginine, histidine, threonine, valine, methionine, isoleucine, and leucine) were found in bovine serum albumin (BSA), and six (histidine, threonine, valine, methionine, leucine, and lysine) in Type I Collagen. The same eight non-essential amino acids (cysteine, asparagine, glutamine, glycine, alanine, serine, tyrosine, and proline) were identified in these two proteins. By RP-HPLC, other two non-essential amino acids (aspartic acid and glutamic acid) were found both BSA and Type I Collagen. Cysteic acid was also found in BSA. In our samples, tryptophan was missing due to its destruction during hydrolysis.

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Abstract  

The effect of glucose (0–15 mass%) on the kinetics of bovine serum albumin (BSA) denatured aggregation at high concentration in aqueous solution has been studied by differential scanning calorimetry. The observed denatured aggregation process was irreversible and could be characterized by a denaturation temperature (T m), apparent activation energy (E a), the approximate order of reaction, and pre-exponential factor (A). As the glucose concentration increased from 0 to 15 mass%, T m increased, E a also increased from 514.59409±6.61489 to 548.48611±7.81302 kJ mol−1, and A/s−1 increased from 1.24239E79 to 5.59975E83. The stabilization increased with an increasing concentration of glucose, which was attributed to its ability to alter protein denatured aggregation kinetics. The kinetic analysis was carried out using a composite procedure involving the iso-conversional method and the master plots method. The iso-conversional method indicated that denatured aggregation of BSA in the presence and absence of glucose should conform to single reaction model. The master plots method suggested that the simple order reaction model best describe the process. This study shows the combination of iso-conversional method and the master plots method can be used to quantitatively model the denatured aggregation mechanism of the BSA in the presence and absence of glucose.

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Abstract  

The kinetics of protein thermal transition is of a significant interest from the standpoint of medical treatment. The effect of sucrose (0–15 mass%) on bovine serum albumin denatured aggregation kinetics at high concentration was studied by the iso-conversional method and the master plots method using differential scanning calorimetry. The observed aggregation was irreversible and conformed to the simple order reaction. The denaturation temperature (T m), the kinetic triplets all increased as the sucrose concentration increased, which indicated the remarkable stabilization effect of sucrose. The study purpose is to provide new opportunities in exploring aggregation kinetics mechanisms in the presence of additive.

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