| The polyol pathway called as sorbitol way is a small part of glycose metabolism. The pathway is regarded as an important element in the pathogenesis of various diabetic complications in individuals with diabetes mellitus. The pathway plays a crucial role in hyperglycemia. The polyol pathway contains two enzymes as aldose reductase (AR) and sorbitol dehydrogenase (SDH). In the present study, AR and SDH were purified from sheep liver by using simple chromatographic methods. AR was purified with a yield of 2.11% and approximately 162 fold and SDH was purified with a yield of 0.53% and approximately 9.07 purification fold. Sodium dodecyl sulfate polyacrylamide gel electrophoresis (SDS-PAGE) was performed to check the purity and determine the subunit molecular weights of the enzymes. Subunit molecular weights of AR and SDH were 38.82 kDa and 37.74 kDa, respectively. Optimal pH, optimal ionic strength, optimal temperature, activation energy, activation enthalpy, Q value and stable pH were determined as 5.5, 10 mM, 50 °C, 2.16 kcal, 1.52 kcal, 1.33 and 8.0 for AR enzyme, respectively. The kinetic parameters K and V for AR were determined as 0.45 mM and 0.55 EU/mL, respectively. These parameters were studied for only AR in the present study, because it was previously studied for SDH. IC values of 3,4-dihydroxybenzoic acid, 3,5-dihydroxybenzoic acid, 3-hydroxybenzoic acid, 4-hydroxybenzoic acid, salicylic acid, p-coumaric acid, ellagic acid, gallic acid, ferrulic acid and caffeic acid on AR and SDH activities were found as 120, 92.0, 49.0, 39.0, 40.0, 690, 7.00, 103, 84.0, 3.00 μM for AR enzyme and 5060, 8350, 6730, 7070, 5780, 24.0, 13.0, 26.0, 17.0, 21.0 μM for SDH enzyme, respectively. According to these results, caffeic acid was the strongest inhibitor for AR activity compared to the other phenolic acids and ellagic acid was also the strongest inhibitor for SDH activity. |
Doç. Dr. Hatice Esra DURAN
Şükrü Beydemir
