Photosensitive Nano-drug Delivery Of O-nitrobenzyl Ester For Controlled Release

At present,most of the small molecule antitumor drugs have such issues as poor water solubility,a large dose,short half-life in vivo,lack of selectivity and severe side effects.These issues can be improved and resolved by changing the existing drug release methods.After the drug is attached to a special delivery system,it can be controllably released under the specific conditions.In the field of biological therapy,light has these major characteristics,for example cleanliness,controllability,high efficiency and non-toxicity.Therefore,the synthesis and performance research of photosensitive drug delivery systems are one of the research focuses in polymer drug science.In this dissertation,the structures of O-nitrobenzyl ester group with photosensitivity were introduced into the structural design of polymeric drug carriers.Two types of nanocarriers that can be used for encapsulating lipophilic drugs were prepared and their structures and performance were analyzed.The specific contents included:（1）O-nitrophenyl glycol was synthesized,using O-nitroacetophenone as material and a series of substitutional addition reactions.Two novel types of photosensitivecrosslinkerof1-（2-nitrophenyl）ethane-1,2-diylbis（2-methylacrylate）（CL1）and1-（2-nitrophenyl）ethane-1,2-diylbis（4-chlorobutanoate）（CL2）were designed and synthesized by the reaction of o-nitrophenyl glycol with methacryloyl chloride and 4-chlorobutyryl chloride,respectively.The structure of the crosslinker was verified by a ultraviolet spectrum,a hydrogen nuclear magnetic resonance spectroscopy（¹H-NMR）and a carbon nuclear magnetic resonance spectroscopy(¹³C-NMR).（2）Preparation and properties of light-sensitive nanospheres PMAA-CL1（PMCL1）.A series of photo/pH dual-sensitive polymethacrylic acid nanospheres（PMCL1）were prepared by distillation precipitation polymerization process,using methylα-methacrylic acid（MAA）as monomers,and CL1 containing O-nitrobenzyl as photosensitive cross-linker,and the effect of cross-linker ratio on photosensitivity was studied.Fourier transform infrared spectrum（FT-IR）showed that CL1 has incorporated into the polymer structure successfully.Thermogravimetric analysis（TGA）analysis confirmed PMCL1 nanospheres have excellent thermal stability,and the maximum decomposition at temperature of about 450℃.Ultraviolet spectrum confirmed O-nitrophenyl carbonyl and carboxyl groups appeared in CL1 and PMCL1 upon 15 min UV,verifing CL1and PMCL1 had photosensitivity.The particle size of nanospheres detected by DLS were 553.1 nm,and the potentials were-27.8 mV,as the cross-linker ratio is 10%,demonstrating the surface of the nanospheres was negatively charged.The particle size and distribution of the nanospheres were relatively uniform,the shape of the particles was intact,using SEM as a characterization.The light/p H controlled release ability of this nanospheres for liposoluble drugs were studied using Nile red（NR）as the model molecules in vitro.According to the best NR release efficiency（about 77%and 64%）in a pH-acidic reaction environment（pH=4.4）,the nanospheres demonstrated obvious photosensitivity and some pH sensitivity.Package load capacity of this nanospheres for liposoluble drugs in vitro at different time was analyzed were studied using PMCL1-10%nanospheres as drug carrier and doxorubicin（DOX）as the model molecules,showing DOX loading was 4.13%,DOX release with no UV and 15 min UV were 34.66%and 56.16%,respectively.The uptake and intracellular release of DOX-PMCL1-10%nanospheres were investigated using fluorescence microscopy as a characterization,and fluorescein diacetate（FDA）as the model molecules in vivo,verifing DOX-PMCL1-10%nanospheres can be taken into the cell,and release the drug upon UV.Finally,cytotoxicity were analyzed by3-（4,5）-dimethylthiahiazo（-z-y1）-3,5-di-phenytetrazoliumromide colorimetry（MMT）assay,indicating the nanospheres and their photodegradation have pretty good biocompatibility（cell viability 95.0%¹05.3%）;and the cytotoxicity of nanospheres was obvious increased upon UV（cell viability as low as 34.75%）,indicating DOX-PMCL1-10%nanospheres have obvious photosensitivity and anti-tumor effect.（3）Preparation and properties of photosensitive quaternary ammonium cross-linked micelles mPEG-b-p（DEAM-co-MMA-CL2）（PDMCL2）.A linear amphiphilic block copolymer mPEG-b-p（DEAM-co-MMA）（PDM）and a quaternary ammonium cross-linked photosensitive micelle PDMCL2 were synthesized using atom transfer radical polymerization（ATRP）,with methyl methacrylate（MMA）and beta-（Diethylamino）ethyl methacrylate（DEAM）as monomer,N,N,N,N,N,N-penta methyl divinyltriamine（PMDETA）as ligand,CL2 as cross-linker,and methoxy polyethylene glycol methyl bromide（mPEG-Br）as initiator.The structures and average molecular weights of PDM and PDMCL2 were characterized by gel permeation chromatography（GPC）and¹H-NMR.¹H-NMR analysis showed CL2 was successfully added to the structure of the polymer,and the molecular weights of PDM and PDMCL2 were 11854g/mol and 23705 g/mol;the average molecular weights of PDM and PDMCL2were 11198 g/mol and 35665 g/mol,using GPC as a characterization.Ultraviolet spectrum confirmed O-nitrophenyl carbonyl and carboxyl groups appeared in CL2 and PDMCL2 upon 15 min UV,verifing CL1 and PMCL1 had photosensitivity.The particle size of micelles detected by DLS was about 1000nm and the potential was 37.6 mV,demonstrating the surface of the micelles was positive charged.The particle size and distribution of the micelles were relatively uniform,the shape of the particles was intact,and some degradation happened upon UV,using SEM as a characterization.The light/pH controlled release ability of this micelles for liposoluble drugs were studied using Nile red（NR）as the model molecules.According to the best NR release efficiency（about64%）in a pH-neutral reaction environment（p H=7.4）,the micelles demonstrated obvious photosensitivity and some pH sensitivity.Package load capacity of this micelles for liposoluble drugs in vitro at different time was analyzed were studied using PDMCL2 micelles as drug carrier and CPT as the model molecules,showing CPT loading was 12.41%,CPT release with no UV and 15 min UV were 35.53%and 98.48%,respectively.Finally,cytotoxicity were analyzed by MMT assay,indicating that the micelles and their photodegradation have pretty good biocompatibility（cell viability 80.9%¹11.7%）;and the cytotoxicity of micelles was obvious increased upon UV（cell viability as low as 19.16%）,indicating CPT-PDMCL2 micelles has obvious photosensitivity and anti-tumor effect.