Preparation and Application of Novel Functional Microspheres

Author:Ge Chun Ling

Supervisor:tan tian wei


Degree Year:2017





Functional microsphere is the polymer material or polymer composite material in the near-perfect sphere shape with the size in micrometer or nanometer.The functional microsphere not only has versatility,but also has better biocompatibility and mechanical property.It can be widely used in biomedicine,immunology,bioseparation and biological catalysis field.This work synthesized different type polymer microsphere and used them after modification in enzyme immobilization,separation and drug delivery.The main contents are as follows:1.pGMA microspheres were prepared use glycidyl mathacrylate(GMA)as monomer,1,3,5-Tri-2-propenyl-1,3,5-triazine-2,4,6(1H,3H,5H)-trione(TAIC)and ethylene glycol dimethacrylate(hydroxyethyl methacrylate(EGDMA)as crosslinking agent and toluene as porogen by suspension polymerization method.The preparation condition was optimized including the crosslinking agent amount,stirring speed and washing conditions.The different crosslink degree of 10%,20%,30%,50%,75%and 100%were prepared with 100%porogen ratio.The mean size of macropore and uniform particle can be comtrolled in the range of 400-500 nm and 70~450 μm.Finally,the enzyme has higher activity when the crosslink degree in the range of 10%~30%and can be used in lipase immobilization for further optimization.2.The agar microsphere with different concentration(4%-4B,6%-6B,and 10%-10B)was prepared by emulsion precipitation technique.The obtained agar microspheres with mean diameter 90 μm have regular spherical shapes.Bi-crosslinking of the obtained microsphere with epichlorohydrin and 1,4-butanediol diglycidyl ether effectively enhanced their mechanical and thermal stability.Compared with the control(<1.5 ml/min)and previous crosslink method(<4 ml/min),the new crosslinked microsphere packed in column can be used in higher flow rate(>6 ml/min).What is more,the alkaline condition in the cross-linking process also decreased the content of sulfate groups of agar microspheres thus reducing the non-specific adsorption of positively charged molecules.Finally,the cross-linked agar microspheres were functionalized with branched poly(ethyleneimine)(BPEI)and 2-diethylaminoethyl chloride hydrochloride(DEAE)to separate proteins in an anion-exchange mode.The mixture proteins can be separated in 40 mins after the elution condition optimizated.3.The gold nanoparticles(GNPs,)and amphiphilic poly(ethylene glycol methyl ether acrylate)(POEGA)were prepared by Frens method and RAFT method,respectively.The molecular weight of reduced POEGA was 10840 g/mol with exposed mercapto group which have specific binding interaction with GNPs surface.The obtained GNPs with a mean diameter 46 nm were coated with POEGA polymer to increase the colloidal stability.The PCL photothermal triggered microspherical depot composed of poy(ε-caprolactone),BSA or horseradish peroxidase(HRP)as model protein,polymer-coated GNPs(PGNP)as photothermal component was prepared by W/O/W emulsion method.The mean size and drug encapsulation efficiency of prepared microspheres for PCL2k、PCL10k and PCL43kwere 40 μm,45 μm,48 μm,and 44%,45%,80%,respectively.4.The prepared photothermal triggered microspheres were used to delivery protein drug.Firstly,the photorheometer property was studied using blue light(400-500 nm):after given blue light at 180 s,the visicosity will be significantly reduced by 81%and 95.5%,respectively.The process is reversible after the light turned off.Secondly,the effect of light intensity,concentration of PGNP and different molecular weight of PCL on the release behavior was studied.Then the folded structure integrity and the bioactivity of released protein were assessed.The results confirmed the released protein was still intact and retained high bioactivity by in vitro H2O2 toxicity mitigation study.Besides,the photothermal system is also biocompatible and easy to implant within the host tissue as protein depot proved by injection experiment with a narrow 26 gauge needle in porcine skin.Furthermore,this system showed no cytotoxicity confirmed by the cell viability results following the ISO-10993 standard protocols.So this photothermal-triggered system has a potential for remotely controlled delivery of therapeutic proteins from an implanted depot.