Study On Characterization Of Three Digestive Enzymes And Preliminary Purification Of Salivary Gland Protease From Rapana Venosa

Combining the remaining studies and the strong feeding ability of Rapana venosa,this paper selected three digestive glands as the basic research objects.The digestive glands were salivary gland,leiblein's gland(esophageal gland)and liver.To explore the main properties of the three digestive enzymes in Rapana venosa for productive practice,hoping to get new enzymes,the enzyme characterization pf protease,amylase and lipase of the three digestive glands were tested.Meanwhile,the protease purification of the salivary gland has been done.The results of this study are summarized as follows:1.The protease properties of the three digestive glands were tested.It's been found that the protease properties was affected by pH?temperature?metal ion concentration?concentration of organic solvent?the surfactant concentration and protease inhibitors concentration.The protease ativity of the salivary gland was much higher than the leiblein's gland and liver.The protease ativities of the three digestive glands of the same concentration when pH7.0,25? was:salivary gland>liver>leiblein's gland.The optium temperature of salivary gland,leiblein's gland and liver protease were 40?,40?,3O?,and optium pH were 8.0,2.0,8.5.The appropriate storage temperature were-20?,-20 ?,4?.It was concluded from the experiment that most of the metal ions restricted the protease ativity,but Ca2+?Mg2+?Mn2+?Ba2+ activated the salivary gland protease ativity,Mg2+?low concentration of Ba2+?Cu2+?Ca2+ and high oncentration of Mn2+?Fe3+ promoted the leiblein's gland protease activity.Different concentrations of methanol,isopropanol and xylene had different effects on the three digestive glands.The tested concentrations of Tween-80 promoted the digestive glands protease ativity,while 4%Tween-80 restricted the leiblein's gland protease activity.Different concentrations of PMSF and EDTA inhibited the salivary gland and the liver protease ativity in different degrees.0.1mmol/L EDTA restricted the leiblein's gland protease activity,while 10 mmol/L EDTA promoted it to a large extent.2.The amylase activity and lipase charaterization of the three digestive glands were also measured.It's been found that the three digestive glands contained amylase and lipase.When pH7.0,25?,the order of amylase ativities of the three digestive glands was:leiblein's gland>salivary gland>liver.When pH7.0,37??the order of lipase ativities of the three digestive glands was:salivary gland>liver>leiblein's gland.The optium temperature of salivary gland,leiblein's gland and liver lipase were 37?,17?,37?,and optium pH were 8,9,7.1 mmol/L Ca2+?Ba2+?Mg2+?Mn2+exhibited different effects on the three digestive glands,while 1 mmol/L Cu2+ inhibited them.40%methanol,dimethyl sulfoxide and ethanol had different effects on the three digestive glands,while xylene and isopropanol inhibited them.3.There was more than one kind of protease in Rapana venosa salivary gland by the native page electrophoresis.High activity and purification fold enzyme extracts were got through(NH4)2SO4 grading salting-out,DEAE Sepharose Fast Flow ion exchange and Sephacryl S-200 HR molecular sieving.