Research On Purification And Separation Of Eucommia Leaf Protein And The Pharmacological Activity

Eucommia ulmoides for the domestic medicinal species is a kind of frequently used as a clinical medicine. Because all is used skin medicine which cost larger and scarcer, People gradually use of eucommia ulmoides leaves instead of medicine According to the study, the leaves mainly include liver and kidney loss, lumbar debility, pain, fetus, and so on. But all the studies are concentrated on the small molecular compounds of eucommia ulmoides leaves, less on the study of macromolecular protein.The experiment that were separate and purify a variety of biological active protein from the eucommia ulmoides leaves, laying a foundation for the development and utilization of the health care industry and healthy. This experiment in eucommia ulmoides leaves as the research material, concluded that the optimal extraction method of water-soluble protein through crushing, flash type water extraction,salting out, alcohol precipitation, acetone precipitation etc. And through the dialysis, dextran gel column and gel electrophoresis method from eucommia ulmoides leaves extract bioactive protein components.According to the determination of protein content and the analysis of the antibacterial, antioxidant etc.results are as follows:1. With bovine serum albumin solution as the standard solution, using the Bradford method, draw standard curve. The linear regression equation is y=0.0959 x +0.0005 and the correlation coefficient is0.9994.2. Albumin extract of eucommia ulmoides leaves protein content as indexes, single factor experiment, orthogonal experiment was carried out respectively, and combined with industrial benefit,which concluded that the optimal process condition is the material liquid than 1:20, mention 10 times, the leaching time of 60 min, and 4? extraction, speeding centrifugal 15 min at 5000 rpm, will clear liquid on the merger.3. Using ammonium sulfate, ethanol, acetone respectively extract water-soluble protein from protein solution for extraction, which weighing 0.0451 g, 0.1076 g and 0.1940 g. IT is concluded that the ammonium sulfate laboratory conditions of the method is simple and efficient extraction method.4. Using the final concentration of 30%, 45%, 60%, 75%, 90% ammonium sulfate on the eucommia ulmoides leaves respectively extract water-soluble protein, which weighing 0.1928 g,0.159 g,0.1301 g, 0.2238 g, 0.2025 g and concentrating 0.1495 ?g/?l, 0.12581 ?g/?l, 0.11481 ?g/?l, 0.10975?g/?l, 0.12099 ?g/?l and yielding 372.5 ?g/g, 314.519 ?g/g,287.013 ?g/g, 274.375 ?g/g and 302.463 ?g/g.IT is concluded that optimizing 30% final concentration of ammonium sulfate can be effective for rough separation of water-soluble protein barks of eucommia ulmoides leaves.5. Using the methods of gel electrophoresis, explores the different concentrations of ammonium sulfate on the effects of eucommia ulmoides leaves protein extraction components, which is obtained using the final concentration of 30% ammonium sulfate to extract protein components.6. Using dialysis, filtering, glucan gel chromatography and other methods, which concluded two kinds of different molecular weight of proteins.7. Thronging the experiment of DPPH free radical scavenging, proved that the five protein concentration in the 0.0-1.0 mg/mL, can produce different degree of scavenging effect, and decreases as the dose, removing capacity is larger. The final concentration of 30%, 90% ammonium sulfate precipitation of crude protein in the concentration of 1.0 mg/L, is larger clearance than others, which are 39.4%, 41.7%respectively. And the final concentration of 30% ammonium sulfate and after purification of two differences proteins showed strong antioxidant capacity.8. By escherichia coli, staphylococcus aureus, bacteriostatic activity of the experiment, proved that the final concentration of 30%, 45%, 60%, 75%, 90% ammonium sulfate of crude protein do not produce of escherichia coli and staphylococcus aureus antibacterial activity at the concentration of 1.5×104 mg/L.9. The experiment of the SH-SY5 Y tumor cell proliferation activity shows that eucommia ulmoides leaves can inhibit protein components, respectively, and promote the role of the tumor cells. The final concentration of 30%, 75%, 90%crude protein of ammonium sulfate is inhibition, and the final concentration of 45%, 60% crude protein of ammonium sulfate is promoting role.