فرزانه رسولزاده

Design of experiments is frequently used to study chemical systems. If enzymatic reactions, as a chemical system, can be modeled using these methods, it will be possible to research with lower prices. Xanthine oxidase and xanthine were selected as enzyme and substrate, consequetively. Initial rate (V) of oxidation of xanthine was studied. Several experiments were designed using central composite design (CCD) method. Three parameters, temperature (T), pH and concentration of substrate, were selected and studied in this research. The reaction was started by adding the enzyme. Absorbance was recorded at 286 nm in different time then initial rate was calculated. Relation between response (initial rate) and selected parameters was studied and suitable mathematical equation was extracted and related response surfaces were plotted. The extracted equation was validated using some tests and other different experiments. CCD is one of the experimental methods that was used to study the oxidation reaction rate of xanthine as a function of T, pH and substrate concentration.These results show that the enzyme parameters can be predicted using extracted mathematical equation.

Quercetin and hesperetin are natural flavonoids with many important therapeutic properties. The interaction of these polyphenolic compounds with bovine milk xanthine oxidase as one of their major target proteins was studied using fluorescence quenching method for the first time. ThisEnzyme (3.0710-8M) incubated with varying concentrations of hesperetin and quercetin (0-100 μM) for 5 min and the fluorescence spectra were recorded at 296, 303 and 310 K. It was found that these compounds quenched the fluorescence of XO. The binding constants and the number of binding sites of quercetin and hesperetin with XO were obtained.It was shown that the fluorescence quenching of quercetin and hesperetin with xanthine oxidase occur through a static, static and dynamic mechanism, respectively. The thermodynamic parameters (enthalpy and entropy changes) were also calculated at different temperatures. Both hydrogen and hydrophobic binding are involved in the interaction of quercetin with xanthine oxidase and hydrophobic binding exists in the interaction of hesperetin with xanthine oxidase.