Preparation And Separation Of Degradable Peptide From Spirulina Protein And Research Of Antibacterial Property

The present study was performed to isolate, purify and determine the activity of a novel antibacterial peptide from Arthrospira (Spirulina) innermongoliansis by ultra-filtration and G-25 gel-filtration. Spirulina phycobiliprotein was prepared by the combination of repeatedly freezing-thaw and ultrasonic wave cracking method. Spirulina phycobiliprotein was hydrolyzed by the combination of papain and alkaline protease. The main results were as follows:(1) Suspending liquid of Spirulina was frozen by liquid nitrogen and melted in 37℃water-bath for 4 times. The dissolution rate of Spirulina phycobiliprotein would reach at 81.5%.(2) By using of the combination of papain and alkaline protease, hydrolysis degree could gain higher. On the base of the single factor experiments, the method of response surface analysis (RSA) with three factors and three levels was adopted. By means of regression analysis, the optimal process conditions were determined. Spirulina phycobiliprotein hydrolysis by alkaline proteinase, enzyme dosage 4300U/g, time 160min, pH value 7.0, temperature 55℃; after destroying enzyme, Spirulina phycobiliprotein hydrolysis by papain protease, enzyme dosage (E/S) 4.5%, time 210min, pH value 6.5, temperature 60℃. The actual hydrolysis degree could reach at 32%.(3) The Spirulina peptide solution were ultrafiltrated with 5000D hollow fiber membrane. After that, components less than 5000D was isolated and purified by G-25 gel-filtration column.In peptide which is less than 5000D, there were four kinds of components. The component having antibacterial activity is the second component. According to the standard curve of Standard peptide molecular weight distribution, we could know the Molecular weight of this component is 360.OOD. Through the speculation, this peptide should be composed by 2 to 5 amino acids.(4) The effects of Spirulina peptide having antibacterial activity on inhibition of S.aureus,B.subtilis,E.coil and B.cereus were studied, taking saline solution as the negative comparison. The result indicated that S.aureus, B.subtilis and E.coil were determined as the tested bacteria which could be used in minimum inhibitory concentration (MIC) tests. The MIC of S.aureus, B. subtilis and E.coil were detected by filter paper method. The MIC were 3.125mg/mL,3.125mg/mL,1.5625mg/mL respectively.The MIC of peptides on E.coli was smaller than sorbic acid, and the MIC of peptides on Staphylococcus aureus and Bacillus subtilis was closely with sorbic acid. Sorbic acid were widely used in food, beverages, tobacco, pesticides, cosmetics and other industries.The peptides of spirulina were nontoxic to humans, and had rich nutrition. So, in future, the spirulina peptides would have a broad application prospects as a natural preservative.