The influence of Al3+ion on porcine pepsin activity in vitro

The in vitro effect of Al3+ ions in the concentration range 1.710-6M-8.710-3M on pepsin activity at pH 2, via kinetic parameters and its electrophoretic mobility was evaluated. Kinetic study demonstrated the existence of an activation effect of Al3+ at pH 2 on pepsin molecule. Kinetic analysis with respect to concentrations of haemoglobin showed that Al3+ ions increase the maximal velocity (Vmax) and kcat values rather than apparent affinity for substrate (KS) implying the non-competitive nature of activation which indicated that aluminium was a non-essential activator of partial non-competitive type. The values of the equilibrium constants KS and KmA for dissociation of corresponding complexes were evaluated as 0.9040.083mM and 8.560.51M, respectively. Dissociation constant KA, of activator from enzyme-activator complex calculated via kinetic and direct measurement of Al3+ binding data, as well as activation constant A50, the activator concentration that gives a rate equal to half at a saturating concentration of activator, were found to be 8.820.90M, 8.390.76M, and 8.050.48M respectively. Native PAGE electrophoresis shows the decrease in electrophoretic mobility of pepsin and confirms modification of the electric charge and conformational changes of pepsin caused by bound Al3+ on the pepsin molecule. Al3+ induced conformational changes of pepsin were verified by UV-VIS and IR spectra. Moreover, the absence of conformational changes in the haemoglobin molecule in the presence of Al3+ ions confirms that the obtained activation is a consequence of conformational changes caused only in the pepsin molecule

Kinetics of 1,8-dihydroxy-2-(pyrazol-5-ylazo)-naphthalene-3,6-disulphonic acid immobilization on anion exchangers

dThe sorption of 1,8-dihydroxy-2-(pyrazol-5-ylazo)-naphthalene-3,6-disulphonic acid (PACA) onto the anion exchange resins, Dowex 1-X8 and Dowex 2-X8, from aqueous solutions with pH between 1 and 10 was studied spectrophotometrically. Conditions for obtaining modified sorbents with a given capacity for the azo compound were determined. The absorption and reflectance spectra of the sorbed dye were recorded. The immobilized reagent retained similar chromic characteristics as in the solution. The sorption mechanism was investigated under static conditions in the temperature range from 0 to 25 degreesC, by determining the amount of sorbed dye as a function of contact time. The sorption was a first order process in all cases. The pore diffusion was found to be a rate-limiting step in the sorption process. The diffusion coefficient values from 6.0 to 7.4 x 10(-7) cm(2) s(-1) were calculated from the experimental results. The activation energy of PACA sorption is - 8.56 and - 8.05 kJ mol(-1) for Dowex 1-X8 and Dowex 2-X8, respectively. (C) 2002 Elsevier Science B.V. All rights reserved

Unusually sluggish microemulsion system with water, toluene and a technical branched alkyl polyethoxylate

Microemulsion systems with water, toluene and nonionic surfactants are interesting for application due to the excellent solvent properties of aromatic hydrocarbons. In this study, the pseudo-binary phase diagram (“fish-diagram”) of such a ternary system was investigated using a branched technical alkyl polyethoxylate. Lutensol ON 50 (i-C10E5) was considered a suitable surfactant. The system with technical branched i-C10E5 exhibits very long and for compositions near optimum solubilization, i.e. the minimum content of surfactant needed for a bicontinuous microemulsion, extremely long times for equilibration. In addition to visual observation, qualitative measurements of turbidity were performed with a UV-Vis spectrometer for characterizing the behavior of this unusually sluggish system. Isothermal phase diagrams at 20, 25 and 30°C yielded inconsistent results after the change of temperature and the application of mechanical stress for different treatment of the samples. Ternary mixtures of water, toluene, and Lutensol ON 50 seem to form long-living metastable states. The sluggishness and the ambiguous phase behavior of the system are discussed. Inconsistent results are attributed to the slow kinetics of the formation and destruction of liquid crystals and the ability of the system to form miniemulsions within an unusual concentration range

Thermal denaturation of pepsin at acidic media: Using DSC, MALDI-TOF MS and PAGE techniques

The thermal stability of pepsin in a strong acid medium as a function of pH has been investigated using differential scanning calorimetry (DSC), UV absorbance, polyacrylamide gel electrophoresis (PAGE) and MALDI-TOF MS methods. The two independent two-state transitions model with view of physiological function of pepsin was used for analyzing thermal denaturation curves. The transition temperature (T-m) values ranging from 305.15 to 319.15K for the first transition and from 322.15 to 349.15K for the second transition in the examined pH range implicating the higher stability at pH 4 are in good agreement with MALDI-TOF MS results. The corresponding maximum, Delta G degrees(25), was stability obtained at pH 4 with values of 65.3 kJ mol(-1). (C) 2013 Elsevier B.V. All rights reserved

Iron complexes of dietary flavonoids: Combined spectroscopic and mechanistic study of their free radical scavenging activity

Combined spectroscopic (UV/visible, Raman, MS) and theoretical approaches were used to assess interaction of iron(III) with quercetin and baicalein in aqueous buffered solutions. Obtained results implicated formation of two iron quercetin complexes, with pH-dependent stoichiometries of 1:2 and 1:1, and one iron baicalein complex with stoichiometry of 1:1. Results of vibrational analysis and theoretical calculations implicated 3-hydroxy-4-carbonyl and the 3'-hydroxy-4'-hydroxy group of catechol as chelating sites for quercetin. For baicalein 5-hydroxy-6-hydroxy group is energetically the most favourable, although 5-hydroxy-4-carbonyl and 6-hydroxy-7-hydroxy chelating sites are energetically similar. Antiradical activity. reaction stoichiometry and number of inactivated DPPH molecules per mole of antioxidant indicated quercetin as a better antioxidant than its iron complex, baicalein and iron baicalein complex. The same structural features appeared to be important both in complexation and antioxidant activity. The equilibrium geometries, optimised using the B3LYP/6-31G (d, p) level of theory, predicted structural modifications between the ligand molecules in free state and in the complex structures. Correlation between experimental and theoretical results was very good

The influence of alkyl polyglucosides (and highly ethoxylated alcohol boosters) on the phase behavior of a water/toluene/technical alkyl polyethoxylate microemulsion system

The influence of additives (alkyl polyglucoside, Glucopon 600 CS UP and alcohol ethoxylate C18E100) on the behavior of the water/toluene/Lutensol ON 50 (technical oxoalcohol, i-C10E5) microemulsion system as a function of temperature and composition has been investigated. The phase behavior of the microemulsions was determined by vertical sections through the Gibbs phase prism (fish-like phase diagrams). Alkyl polyglucoside shifts the one phase region to lower temperatures compared with water/toluene/Lutensol ON 50 mixtures. This is contrary to the expectation, considering the extreme hydrophilic nature of the sugar headgroup. The addition of hydrophilic alcohol ethoxylate (C18E100) to the water/toluene/Lutensol ON 50 system increases the solubilization capacity of the surfactant, even if the co-surfactant is used in small quantities, and shifts the one-phase region to higher temperature by a few degrees Celsius. [Projekat Ministarstva nauke Republike Srbije, br. 172051