Preparation,characterization Of Zein-based Nano/microparticles And Their Biocompatibility And Immunogenicity Evaluation

Due to low cost,good biodegradability and strong ductility,zein and zein-based delivery systems have received more and more attention.In this thesis,the effects of various factors of phase separation method on the properties of zein nanoparticles were investigated in detail.A simple and efficient method for the controlled preparation of zein nanoparticles was finally obtained.The related mechanism was also discussed.Subsequently,we further investigated the histocompatibility and immunogenicity of zein nanoparticles in mice.This thesis provided valuable and important information on in vivo application of zein nano/microparticles.In the part of fabrication of zein nano/microparticles,we found that zein nanoparticles with good reproducibility,controlled particle size and zeta potential,and low polydispersity index could be obtained by tuning different parameters.The faster the mixing rate,the lower the temperature,and the lower the initial concentration of zein,the smaller the particle size of the resultant nanoparticles were.Combining the factors of mixing rate and zein concentration enabled a controlled preparation of zein particles from nanometer to micrometer scale.Introducing sodium caseinate into the system eliminated the influence of the above parameters on particle size.However,with the two-step method of adsorption onto preformed zein nanoparticles,core-shell zein/sodium caseinate nanoparticles were obtained in a size-controllable manner.The resultant carriers had obvious advantages in drug encapsulation efficiency and controlled drug release profiles compared with traditional hybrid zein/sodium caseinate carriers.The results of histocompatibility experiment showed that injection of 800 μg zein nanoparticles into mice via intramuscular and subcutaneous routes resulted in varying degrees of inflammation in the injection site on the third day and the first day,respectively.However,the corresponding tissues could recover within the third day and the seventh day,respectively.The above results suggested that zein nanoparticles led to a transient inflammation and is acceptable in terms of biocompatibility.The results of immunogenicity investigation showed that intramuscular injection of zein nanoparticles with different particle sizes at the same dose induced similar antibody levels in vivo in Balb/c mice,and the type of immune response was Th2 biased humoral immune response.The level of Ig G1/Ig G2 a in the 250 nm group was higher than those of others,suggested a higher tendency to induce Th2-type immune response.Further studies demonstrated that the immunogenicity of zein was administration route-dependent.The highest antibody titer produced in the intramuscular injection group was slightly higher than that of the subcutaneous injection group,and it was dose-dependent in the range of 200μg,600μg and 800μg.However,there was no significant difference between the three subcutaneous injection groups with different doses.When the antibody titer reached to plateau at the low level,mice were given forth injection.The Ig G antibody titer rapidly increased to a comparable maximum level as before and maintained for a long time.Further antibody subtype analysis showed that Ig G1/Ig G2 a was higher in the subcutaneous injection group than that of the intramuscular injection group at the highest antibody titer.However,the Ig G1/Ig G2 a values were not dose-dependent in both groups inoculated intramuscularly and subcutaneously.The above results indicated that the zein nanoparticles could induce a memory Th2 type immune response in mice.