Study On Purification And Enzymatic Properties Of The Barley Root Phospholipase

In this paper， with response surface methodology, we investigated that thermaldenaturation temperature、thermal denaturation time and pH had an effect on phospholipasespecific activity in the barley root. Further studied the conditions of separation andpurification of barley root phospholipase were studied with the SephadexG-100gelchromatography and CM-Sephadex C-50ion exchange chromatography, we got a more purephospholipase, and studied the enzymatic properties of the enzyme.1. The experiment found that the best extraction phospholipase conditions were asfollows: thermal denaturation temperature was40.97℃,thermal denaturation time was23.76min and pH was4.96,the maximal phospholipase specific activity was up to30.03U/mg,the extracting purity was the highest in such a case.2. In the further course of the study，results showed that two elution peaks wereobtained,and only S1peak had enzyme activity by SephadexG-100gel chromatographyseparation,and S1peak had5electrophoretic bands by SDS-PAGE, the concentrated enzymeliquid was purified4.86times, and coefficient of recovery of enzyme activity was40.51%.With CM-Sephadex C-50ion exchange chromatography,5elution peaks wereobtained, and only the C3peak had enzyme activity, the vitality peak had only oneelectrophoretic band, concentrated enzymes liquid was purified8.8times coefficient ofrecovery of enzyme activity was16.3%.3. On the enzymatic properties of the study，the results showed that the enzymemolecular mass of about5.28×10⁴Da，in the experiment within the framework of metal ions，Ca²⁺on the phospholipase activity of the role of promoting, K⁺、Na⁺、Mg²⁺、Fe²⁺、Fe³⁺、Li⁺、Co²⁺on the enzyme activity has a slight inhibitory effect, Cu²⁺、Cr³⁺、As³⁺、Se⁴⁺on theenzyme activity significantly inhibited, Se⁴⁺on the enzyme activity of which inhibition of themost significant.