Showing papers by "Nazan Demir published in 2005"

Identification of protease from Euphorbia amygdaloides latex and its use in cheese production.

Abstract: In this research, protease enzyme was purified and characterized from milk of Euphorbia amygdaloides. (NH4)2SO4 fractionation and CM-cellulose ion exchange chromatography methods were used for purification of the enzyme. The optimum pH value was determined to be 5, and the optimum temperature was determined to be 60 degrees C. The V(max) and K(M) values at optimum pH and 25 degrees C were calculated by means of Linewearver-Burk graphs as 0.27 mg/L min(-1) and 16 mM, respectively. The purification degree was controlled by using SDS-PAGE and molecular weight was found to be 26 kD. The molecular weight of the enzyme was determined as 54 kD by gel filtration chromatography. These results show that the enzyme has two subunits. In the study, it was also researched whether purified and characterized protease can be collapsed to milk. It was determined that protease enzyme can collapse milk and it can be used to produce cheese.

Purification and characterization of carbonic anhydrase from bovine stomach and effects of some known inhibitors on enzyme activity.

Abstract: Carbonic anhydrase (CA) was purified from four different cell localisation (outer peripheral, cytosolic, inner peripheral and integral) in bovine stomach using affinity chromatography with Sepharose-4B-L-tyrosine sulphanilamide. During the purification steps, the activity of the enzyme was measured using p-nitrophenyl acetate at pH 7.4. Optimum pH and optimum temperature values for all CA samples were determined, and their K(m) and V(max) values for the same substrate by Lineweaver-Burk graphics. The extent of purification for all CA localizations was controlled by SDS-PAGE. The K(m) values at optimum pH and 20 degrees C were 0.625 mM, 0.541 mM, 0.785 mM and 0.862 mM with p-nitro phenyl acetate, for all CA localizations. The respective V(max) values at optimum pH and 20 degrees C were 0.875 micromol/L min, 0.186 micromol/L min, 0.214 micromol/L min and 0.253 micromol/L min with the same substrate. The K(i) and I50 values for the inhibitors sulphanilamide, KSCN, NaN3 and acetazolamide were determined for all the CA localizations.

Enzymatic Zinc Determination in Human Pancreatic Juice

Abstract: Objective: Evaluation of an enzymatic method for determining trace amount of zinc was in human pancreatic juices. Materials and Methods: Bovine carbonic anhydrase was purified from erythrocyte hemolisate by affinity chromatography. Zinc present in active site of enzyme was removed by dialysis against dipicolinic acid and resulting in apo carbonic anhydrase obtained at a ratio of 100%. The activity of the enzyme was determined by the esterase activity on pnitrophenyl acetate. Standard curve obtained with constant apoCA (3.6x10 -5 mol/L) and changing Zn 2+ concentrations. Human pancreatic juices were taken from 10 patients and zinc concentrations with enzymatic method were determined using standard curve. For comparison, the same samples were analyzed in atomic absorption. Results: When the results of the AA and apoCA methods were compared, it was found from the r and p values that there is a good correlation between the two methods. Conclusion: The enzymatic method can easily be used in human pancreatic juice zinc determination.