The Preparation Of Chitosan-based Nanosystems And In Vitro Study Of Their Applications As Carriers For The Antitumor Drugs

Chitosan, the only kind of naturally occurring basic polysaccharide discovered so far, is widely present in the shell of crab, shrimp and silkworm pupa, and shows wide applications in the medicine field. Recently, many studies have found that some chitosan derivatives by chemical modification can be used as the carrier materials to exploit good delivery nanosystem for antitumor drugs. This study mainly investigated the extraction process of chitosan from silkworm pupa shell, the preparation of chtiosan-based nanosystems and the feasibility of their application as the carrier for antitumor drugs.1. The extraction conditions of chitosan from silkworm pupa shellIn this section, serial chitosan samples were extracted from silkworm pupa shell under different conditions and their properties were subsequently determined, thus optimized the extraction conditions. First, the multifactor-multilevel research method could be used to optimize the extraction process of the pupa chitosan. Next, the properties of extracted chitosan, e.g., appearance, yield, water content, viscosity, and deacetylation degree, were assessed by the naked eye-observation, weighing, desiccation, Ubbelohde viscometer and fourier transform infrared spectroscopy (FTIR) methods, respectively. There are four steps in the extraction process. During each step, the concentration of reagents, reaction time and reaction temperature will affect the properties of the pupa chitosan. The optimized extraction conditions of chitosan from silkworm pupa shell can be obtained by comparing the conditions of these four steps.2. Nanoparticle carrier based lactosylated chitosan for antitumor drugThis section is to prepare lactosylated chitosan-based nanoparticles used as the carrier for the anticancer drug:methotrexate (MTX) and to provide the reference data for the study of liver-targeting drug carrier systems. First, N-lactosyl-chitosan conjugate (CLCS) was synthesized and characterized by FTIR and proton nuclear magnetic resonance (’H-NMR), respectively. Second, CLCS nanoparticles were prepared by the self-assembly method; their morphologies and sizes were assessed by the transmission electron microscopy (TEM) and dynamic laser light scattering (DLS). Third, MTX was phaysically loaded into CLCS self-assembled nanoparticles using microdialysis method. MTX loading efficiency and content were determined by Ultraviolet (UV) method. MTX in vitro releases from CLCS nanoparticles were valuated by the dynamic dialysis method. The preliminary experiment results showed that CLCS nanoparticles can effectively load MTX, and realize the sustained release of MTX in vitro.3. Chitosan-based nanoparticles for the cancer combinational therapyThis section studied the preparation of a chitosan-based multifunctional nanoparticle carrier and its drug loading and release properties. First, chemotherapeutic agent, MTX, was conjugated to chitosan to systhesize polymeric prodrug of MTX-CS. Chemical structure of MTX-CS was characterized by IR and1H NMR, respectively. MTX-CS nanoparticles were prepared by the self-assembly method in aqueous media. Anti-angiogenic agent, combretastatin A-4(CA-4), was physically loaded into MTX-CS nanoparticles. The morphologies and sizes of MTX-CS nanoparticles and CA-4loaded MTX-CS (CA-4/MTX-CS) nanoparticles were assessed by the transmission electron microscopy (TEM) and dynamic laser light scattering (DLS); drug loading content and efficiency were determined by UV method; drug in vitro releases from CA-4/MTX-CS nanoparticles were valuated by the dynamic dialysis method. The preliminary experiment results showed that two antitumor drugs with different mechanisms can be efficiently loaded into chitosan-based nanoparticles via chemical bonding and physical encapsulation, and can be orderly released from nanoparticles in vitro, which was favorable for the cancer combinational therapy.