Preparation And Releasing Characteristics Of Recombinant Penaeidin3-2 Microcapsules

Recombinant antimicrobial peptide is a kind of minor polypeptides produced through genetic engineering method. Due to their small molecular weight, heat stability, broad-spectrum of antimicrobial activities, the AMP were considered to be excellent candidates for potential novel antibiotic agents. This protein drugs should be generally delivered by oral approach for the aquaculture characteristics. However the clinic applications through oral delivery systems are still not successful for most of them because of their short half-life resulting from acid-catalytic or proteolytic degradation in the gastrointestinal tracts. A potential solution is microencapsulation through which the protein drugs are encapsulated, preventing from the degradation, and releasing in sustained and controlled manners.In this paper, the recombinant antimicrobial peptides (penaeidin 3-2) were purified with affinity chromatography, and recombinant antimicrobial peptides sodium alginate microcapsules were prepared by using coacervation technology. The morphology and encapsulation rate were introduced as indexes of microcapsules formulation. Afterward, in vitro releasing characteristics of the optimal microcapsules was investigated. The results showed that the microcapsules with a diameter of 1.1mm and an encapsulation rate of 83.87% were prepared when the concentrations of calcium chloride and sodium alginate were 1.5% and 2.0%, respectively. The releasing rate of recombinant AMP after standing for 2 h in simulated gastric fluid (pH2.0) was less than 14%. However, after standing in simulated intestinal fluid (PH7.8) for 5 h, the releasing rate of recombinant AMP reached 98%.Which indicated that the intestinal-lysis microcapsules have the potential to be used as sustained-release and controlled-release preparations, providing the experimental basis for oral drug delivery system in aquaculture.Recombinant antimicrobial peptides-sodium alginate microcapsules were orally given to Tilapia and fishes in the control groups were given with special and ordinary forage.20d after that liver, stomach, intestine, sera and muscle were analyzed by Western Blot. The recombinant AMP was not discovered in simples of all groups. In this case, the oral administration of recombinant AMP still need to be further researched.Using purified recombinant AMP as model protein and PLGA as controlled-release carrier, recombinant AMP-PLGA microspheres were prepared by W/O/W multiple emulsion volatilizing method. Using entrapment efficiency, particle size, and 24h-release amount as the evaluating indicators, we optimized the 7 factors that influenced the preparation technique for microspheres by orthogonal factorization method. The morphology was investigated using scanning electron microscope(SEM) .The results showed that the microspheres seemed to be smooth and uniform with mean particle size of 3μm under conditions of recombinant AMP of 10 mg, PLGA of 200 mg, 1%PVA, ultrasonic power of 100 W, 5%PEG, 10%NaCl. However, the encapsulation efficiency of microspheres was only 10%.