A.pernyi Silk Microspheres Prepared By Ion-induced Self-assembly And Its Application In Drug Delivery

With its excellent biocompatibility, biodegradability and breathable oxygenpermeability, silk fibroin of domestic silkworm Bombyx mori (B. mori SF) is widely usedin various field, such as surgical sutures, wound dressings, enzyme immobilizationmaterials, drug delivery and tissue engineering etc. However, the Antheraea pernyi(A.pernyi) silk fibroin not only has the advantage of B. mori SF in biological medicinefield, but also contains the tripeptide sequence Arg-Gly-Asp(RGD) which is known toexhibit specific interactions with mammal cells. Therefore, it is accepted to have betterbiocompatibility and effect than B.mori as a wound dressing, a scaffold for tissueengineering, a drug carrier and a cell culture substrate. On the base of researching in theproperties of regenerated A.pernyi silk fibroin solution (ASF), in this paper, we adopt a newmethod to prepare the A.pernyi silk microspheres by ion-induced self-assembly. At thesame time, to prepare the small molecule drug ibuprofen sodium/ASF microspheres withnegative charge, ibuprofen/ASF microspheres with neutral charge and adriamycin/ASFmicrospheres, a cancer drug with positive charge. Furthermore, the reasons with differentrelease effects were also discussed on the system.As a kind of high purity protein that secreted by the endothelial cells, ASF hasexcellent biocompatibility, which amino acid composition is given priority to with glycineand alanine and serine. Based on the observation and analysis of the preliminaryexperiment results, It is found that the regenerated ASF solution was sensitive and highlyunstable. So the microspheres were formed easily in ASF solution and the average sizes ofthe silk particles greatly increased form3nm to1.5μm. It offers experimental basis for theprepartion of ASF nanoparticles. Mixing the different valences and concentration of ionswith A.pernyi silk fibroin to prepare the A.pernyi silk microspheres by ion-inducedself-assembly. Its characteristic is that the shape of prepared microspheres were regularspherical structure, uniform distribution and the production rate of microspheres can reach more than96%in the condition of the test. Meanwhile, it is found that ASF microspheres,with small size ranging form0.1to0.4μm, can be prepared within a short time and isbiodegradable. The merits of this novel method included a rather simple productionapparatus and no potentially toxic agents, such as cross-linking agent, initiators, surfactant,or organic regent. The mechanism of ASF microspheres formation by ion-inducedself-assembly was also discussed in this paper: the rapid transition of ASF from sol tomicros is based on the hydrophobic effect and electrostatic interactions. With increased theionic concentration, strong hydrophobic forces and electrostatic effect can trigger fasterfibroin unfolding. Simultaneously, ASF molecules self-assembled into stable β-sheetstructures, ultimately leading to associating composite particles.This paper also studies on the preparation of small molecule drug ibuprofensodium/ASF microspheres with negative charge, ibuprofen/ASF microspheres with neutralcharge and adriamycin/ASF microspheres, a cancer drug with positive charge. Through thestudy of different charge drug microspheres, the results showed that the drug loading rateand drug encapsulation efficiency are associated with the inputs of drugs, but due to ASFmicrospheres themselves show negative charge, so the the drug loading rate and drugencapsulation efficiency are different under the drugs of different charge. Positiveadriamycin/ASF microspheres compared with the other two drugs with high encapsulationefficiency and drug loading. When adriamycin increased to16%, the rate of thedrug-loading and encapsulation efficiency can reach11.4%and82.5%, respectively. Onthe contrary, ibuprofen sodium/ASF microspheres with negative charge have lowerdrug-loading rate and encapsulation efficiency. When ibuprofen sodium up to60%, the rateof drug-loading was2.5%and the encapsulation efficiency as low as4.08%. However,ibuprofen/ASF microspheres were between the two. At the same time, the release ofmicrospheres with different charge of drug has a pH sensitive features, In vitro testsmanifested that the release of the adriamycin/ASF microspheres were faster under theenvironment of low pH (5.2), the release was slowly after one day. Finally, there are stillabout65.4%drug are not released which showed that the slow-release effect of the drugmicrospheres was significant. But There is an obvious sudden release of ibuprofensodium/ASF microspheres and the phenomenon more obvious up to88.2%under theenvironment of high pH (8.0). Ibuprofen/ASF microspheres with neutral charge has moreobvious sustained release properties and under the environment of high pH (8.0) release faster. Finally, there are still about65.4%drug are not released after10hours.At last, on the basis of those, the cellular compatibility of CaCl2/ASF microspheresand the suppression tumor performance of ADM/ASF microspheres were discussed.Experiments show that CaCl2/ASF microspheres were not only able to support cell growthand proliferation but also cell dispersed evenly and morphology were better, there are nosignificant difference with pure ASF microspheres. ADM/ASF microspheres have obviousinhibitory effect to the cancer cells which showed that the slow-release effect of the drugmicrospheres was significant after3days. Meanwhile, when the concentration is lowerthan50ug/ml, ADM/ASF microspheres were not obvious effect on normal cells.