Tülin Aydemir

Partial purification and characterization of polyphenoloxidase from peppermint (Mentha piperita)

Abstract Polyphenoloxidase (PPO) of peppermint leaves ( Mentha piperita) was isolated by (NH4)2SO4 precipitation and dialysis. Its pH and temperature optima were 7.0 and 30°C, respectively. On heat-inactivation, half of the activity was lost after 6.5 and 1.5 min of treatment at 70 and 80°C, respectively. Sucrose, (NH4)2SO4, NaCl and KCl appeared to be protective agents of peppermint PPO against thermal denaturation. Km of this enzyme ranged from 6.25×10−3 M with catechol to 9.00×10−3 M with L-dopa. The I50 values of inhibitors studied on PPO were determined by means of activity percentage (I) diagrams. Values were 1.4×10−4 M, 1.7×10−4 M, 9.7×10−5 M, 2.45×10−4 M, 2.16×10−1 M, 1.83×10−5 M, 6.5×10−5 M, 1.4×10−2 M, 7.5×10−5 M, for potassium cyanide, glutathione, ascorbic acid, thiourea, sodium azide, sodium metabisulfite, dithioerythritol, β-mercaptoethanol and sodium diethyl dithiocarbamate respectively. Therefore, sodium metabisulfite was the most effective inhibitor.

Immobilization of tyrosinase on chitosan–clay composite beads

Tyrosinase was immobilized on glutaraldehyde crosslinked chitosan-clay composite beads and used for phenol removal. Immobilization yield, loading efficiency and activity of tyrosinase immobilized beads were found as 67%, 25% and 1400 U/g beads respectively. Optimum pH of the free and immobilized enzyme was found as pH 7.0. Optimum temperature of the free and immobilized enzyme was determined as 25-30 °C and 25 °C respectively. The kinetic parameters of free and immobilized tyrosinase were calculated using l-catechol as a substrate and K(m) value for free and immobilized tyrosinase were found as 0.93 mM and 1.7 mM respectively. After seven times of repeated tests, each over 150 min, the efficiency of phenol removal using same immobilized tyrosinase beads were decreased to 43%.

Removal of lead (II) ions from aqueous solution by using crosslinked chitosan-clay beads

Abstract A simple and effective biodegradable material known as chitosan-clay composite beads were prepared to remove Pb(II) ions from aqueous solution. For this purpose, various important parameters such as contact time, pH and temperature were examined on the adsorption of Pb(II) ions onto crosslinked chitosan-clay composite beads. Maximum adsorption capacity of Pb(II) was observed at pH 4.5 and 25°C and calculated as 7.93 mg/g according to Langmuir isotherm model. Thermodynamic parameters namely ΔG°, ΔH° and ΔS° of the Pb(II) adsorption process have been calculated as 7.889 kJ/mol, −15.131 kJ/mol and −0.0785 kJ/molK respectively. EDTA was the best eluent for the desorption of Pb(II) ions from the crosslinked chitosan-clay beads. Scanning electron microscope (SEM) was used to characterize the surface morphology of the crosslinked chitosan-clay beads.

An efficient removal of RB5 from aqueous solution by adsorption onto nano-ZnO/Chitosan composite beads

In this study, the removal of Reactive Black 5 (RB-5) by nano-ZnO/Chitosan composite beads (nano-ZnO/CT-CB) from aqueous solution was investigated. ZnO nanoparticles were prepared by the via the microwave-assisted combustion technique. And then nano-ZnO/Chitosan composite beads were prepared by polymerization in the presence of nano-ZnO and chitosan. Characterization of composite beads were conducted using SEM, TEM, FTIR, TGA and XRD. Several important parameters influencing the removal of RB 5 such as contact time, pH and temperature were investigated systematically by batch experiments. At optimum conditions of pH 4 and adsorbent concentration of 0.2g, dye removal efficiency was found 76%. Langmuir, Freundlich and Temkin adsorption models were used to describe adsorption isotherms and constants. The maximum adsorption capacity (qm) by Langmuir isotherm has been found to be 189.44mg/g. Isotherms have also been used to obtain the thermodynamic parameters such as free energy, enthalpy and entropy of adsorption. The positive value of the enthalpy change (32.7kJ/mol) indicated that the adsorption is an endothermic process. The obtained results showed that the tested adsorbents are efficient and alternate low-cost adsorbent for removal of dyes from aqueous media.

Selected Kinetic Properties of Polyphenol Oxidase Extracted from Rosmarinus Officinalis L.

Polyphenol oxidase from Rosmarinus officinalis L. (PPO, EC 1:14:18.1) was extracted and partially purified by using (NH4)2SO4 precipitation and dialysis. Stable and highly active PPO extracts were obtained using 0.5% (w/v) PEG and 0.5% (w/v) Triton X-100 in 0.1 M phosphate buffer 7.0. KM values were found to be as 14.3 mM for catechol. Four isoenzymes of Rosemary PPO were detected by PAGE with DL-dopa substrate. The enzyme was strongly inhibited by dithiotreitol, sodium metasulfite, sodium thiosulfate, ascorbic acid, and L-cysteine. Metal ions Ca++, Mg++, and Mn++ were poor inhibitors of rosemary PPO at 10 Mm.

Antioxidant responses of lentil and barley plants to boron toxicity under different nitrogen sources

In this study, the effects of different nitrogen sources on lentil (Lens clunaris) and barley (Hordeum vulgare) plants, exposed to 5 and 10 mM boron stress previously, were studied. After ten-day germination, the lentil (native) and barley (Tokak157/37) were incubated 16 h light and 8 h dark per day for 7-day growth cycle under the conditions of boron stress via different nitrogen sources (10 mM nitrogen in NH4 Cl, KNO3 and urea). As a result of the changes in the nitrogen sources of the plants, there were determined decreases in the relative growth rate (%) and total chlorophyll content related to boron stress, (p < 0.05) and (p < 0.01), respectively. The changes in the lentil were obtained much more than those in the barley. In addition, the changes in the groups in which NH4 + was used as nitrogen source were obtained at lowest levels. The concentrations of MDA, H2O2 and proline showed increases under boron stress (p < 0.05). The effect of boron toxicity on the activities of SOD, GPX and LOX was similar in the two species but the levels of CAT and APX activities were different in both species under 5 and 10 mM boron stress (p < 0.01). SOD, GPX and LOX activities increased in the roots and shoots of boron treated plants as compared with the controls (p < 0.01). Although, lentil CAT and APX, activities decreased; in barley, CAT and APX, activities increased under boron toxicity. In conclusion, the fertilizers which contain NH4 + should be used in the boron stressed farmlands.

Purification and Characterization of Catalase from Chard (Beta vulgaris var. cicla)

Catalase is a major primary antioxidant defence component that primarily catalyses the decomposition of H2O2 to H2O. Here we report the purification and characterization of catalase from chard (Beta vulgaris var. cicla). Following a procedure that involved chloroform treatment, ammonium sulfate precipitation and three chromatographic steps (CM-cellulose, Sephadex G-25, and Sephadex G-200), catalase was purified about 250-fold to a final specific activity of 56947 U/mg of protein. The molecular weight of the purified catalase and its subunit were determined to be 235000 and 58500 daltons, indicating that the chard catalase is a tetramer. The absorption spectra showed a soret peak at 406 nm, and there was slightly reduction by dithionite. The ratio of absorption at 406 and 275 nanometers was 1.5, the value being similar to that obtained for catalase from other plant sources. In the catalytic reaction, the apparent Km value for chard catalase was 50 mM. The purified protein has a broad pH optimum for catalase activity between 6.0 and 8.0. The enzyme had an optimum reaction temperature at 30 °C. Heme catalase inhibitors, such as azide and cyanide, inhibited the enzyme activity markedly and the enzyme was also inactivated by β-mercaptoethanol, dithiothreitol and iodoacetamide.

Characterization of β-glucosidase immobilized on chitosan-multiwalled carbon nanotubes (MWCNTS) and their application on tea extracts for aroma enhancement

β-Glucosidase was covalently immobilized on chitosan-MWCNTs carrier and its aroma enhancement effect in different tea samples was investigated. Chitosan-MWCNTs carrier was prepared by mixing chitosan with MWCNTs (5:1w/w) and characterization of prepared composite carrier was done by FTIR, TGA and SEM analysis. β-Glucosidase was covalently immobilized on the composite carrier after glutaraldeyde activation. After optimization of the immobilization conditions, immobilization yield was achieved as 95.22%. Optimum pH was found as pH 6.0 and pH 5.0 for free and immobilized enzyme, respectively. Optimum temperature of the enzyme was shifted from 35°C to 45°C after immobilization. The Km and Vmax values for immobilized β-glucosidase calculated as 5.55mM and 7.14U/mg protein respectively. Immobilized β-glucosidase showed better pH and storage stability than free enzyme. After storage at +4°C for 50days, the immobilized enzyme retained its 68.4% of the initial activity. The calculated half-life (t1/2) of immobilized enzyme was 115.8min. After 10 cycles of reuse, immobilized β-glucosidase showed 72.83% of its initial activity.

Purification and Characterization of Glutathione-S-Transferase from Chicken Erythrocyte

The glutathione-s-transferases are a family of multifunctional enzymes involved in the detoxification of electrophilic xenobiotics primarily through conjugation to reduce glutathione. A form of the enzyme, designated GSH-S transferase ρ, was purified chicken erythrocyte by acetone precipitation, ethanol-chloroform treatment, DEAE-Cellulose, Q-Sepharose, Sephadex G-100 chromatography. The molecular weight of GST purified from chicken erythrocyte was estimated as 47,500 Da by gel filtration. The subunit molecular weight of chicken erythrocyte GST as determined by electrophoresis in the presence of sodium dodecyl sulfate was predicted as 24,000 Da. The specific activity was found to be 20.39 U/mg. The km for CDNB calculated from Lineweaver-Burk plot was 0.71 mM. Optimum temperature of maximum GST activity was 28°C for CDNB. The maximal activity of the enzyme was observed at pH 7.5. The activity of purified GST is inhibited by DDT, urea, CDNB, Triton X-100, DTNB.

Characterization and immobilization of Trametes versicolor laccase on magnetic chitosan–clay composite beads for phenol removal

Abstract Laccase from Trametes versicolor was immobilized on magnetic chitosan–clay composite beads by glutaraldehyde crosslinking. The physical, chemical, and biochemical properties of the immobilized laccase and its application in phenol removal were comprehensively investigated. The structure and morphology of the composite beads were characterized by SEM, TGA, and FTIR analyses. The immobilized laccase showed better storage stability and higher tolerance to the changes in pH and temperature compared with free laccase. Moreover, the immobilized laccase retained more than 75% of its original activity after 10 cycles. The efficiency of phenol removal by immobilized laccase was about 80% under the optimum conditions after 4 h.