水溶性壳聚糖修饰甘草酸脂质体的制备及其释药性能的研究
|
摘要
脂质体是具有类似生物膜结构的双分子层膜,作为药物载体具有靶向性能和缓释性能,能降低给药剂量和药物毒性。但未经修饰的传统脂质体,其靶向性主要集中于网状内皮细胞丰富的器官,对其他组织器官进行治疗其靶向性则不明显;同时传统脂质体存在体内外稳定性差、存储期间磷脂易氧化水解、双分子膜易变相、脂质体粒子之间易相互聚集融合、包裹在其中的药物易渗漏等缺点。
近年来为了改善脂质体的靶向性和体内外稳定性,很多学者对脂质体膜进行表面修饰,即在脂质体的膜材组成中加入一定的辅助剂,如胆固醇、大豆甾醇、聚乙烯醇及聚乙二醇等,增加脂质双层膜的稳定性,提高脂质体体外、体内稳定性。本课题主要研究利用可生物降解、无毒、生物相容性良好的水溶性壳聚糖衍生物对脂质体进行表面修饰,以期构建具有较高稳定性和靶向性的脂质体载药传递系统。主要研究内容和结论如下:
1、以壳聚糖为原料,在碱性条件下与氯乙酸进行醚化反应,合成得到水溶性的羧甲基壳聚糖;在硼氢化钠存在下,羧甲基壳聚糖与乳糖进行还原胺化反应,合成得到半乳糖基羧甲基壳聚糖。采用红外光谱对合成产物进行结构表征,同时通过双突跃电位滴定法对羧甲基取代度和氨基上半乳糖基取代度进行测定。结果表明:羧甲基化反应主要发生在C_6位羟基上,取代度为0.65左右;半乳糖化反应发生在C_2位氨基上,取代度为0.20左右。
2、采用薄膜分散-超声法制备修饰脂质体。以包封率为指标,通过正交实验对脂质体进行制备处方筛选和工艺优化,采用扫描电镜和超细粒度及Zeta电位检测仪对脂质体进行结构表征。结果表明:修饰脂质体的最佳处方为磷脂/甘草酸质量比10:1,羧甲基壳聚糖(半乳糖基羧甲基壳聚糖)浓度为0.3%,在羧甲基取代度为0.65和半乳糖基取代度为0.22时,羧甲基壳聚糖修饰的甘草酸脂质体平均粒径为122nm,Zeta电位为-53.2mV;半乳糖基羧甲基壳聚糖修饰的甘草酸脂质体平均粒径为112nm,Zeta电位为-50.7mV;脂质体形貌规整,粒径较小符合正态分布,同时具有较高的药物包封率,达到80%左右。
3、研究修饰脂质体的体外释药性能,考察修饰材料的不同浓度和取代度,以及不同pH释药介质对修饰脂质体体外释药性能的影响。结果表明:经羧甲基壳聚糖和半乳糖基羧甲基壳聚糖修饰的甘草酸脂质体具有药物缓释控释性能,在释药初期1h内平均累积释药15%左右,随后缓慢释药,12h内平均累积释药45%左右,能够构建稳定性好、具有潜在主动肝靶向性能的脂质体载药控释系统;并且经羧甲基壳聚糖修饰后的甘草酸脂质体具有良好的体外pH敏智能控释性能。
Liposomes have a biological structure similar to bilayer membrane.As drug carriers,they have ability of targeting,delayed release,reducing the dose and drug toxicity.But the targeting of unmodified liposomes focus on organs that are rich in reticuloendothelial cell,while in other parts of organs their targeting are not obvious during the therapy.Meanwhile,traditional liposomes are not stable in vivo and in vitro, easily oxidized and hydrolyzed,and their bilayer membrane will also easily convert the phase,aggregation and fusion will happen between liposomes,then the drug encapsulated in liposomes will easily seepage from it,and so on.
In recent years,in order to improve the targeting and stabiliy in vivo and in vitro of liposomes,many scholars have prepared the liposome membrane of surface modification,which is adding certain auxiliary agents in the composition of membrane material,such as cholesterol,soy sterol,polyvinyl alcohol and polyethylene glycol to increase the stability of lipid bilayer membrane and enhance the stability of liposomes in vitro and in vivo.The study of this dissertation is mainly to use biodegradable,non-toxic,water-soluble,low-molecular-weight and good biocompatibie chitosan derivatives on liposomes surface-modifying,for building a high stability and special targeting liposomal drug delivery systems.Main contents and conclusions are as follows:
Firstly,chitosan as raw materials,hydrosoluble carboxymethyl chitosan was synthesized through the etherealization reaction under basic condition between chitosan and chloroacetic acid.Then,with existence of sodium borohydride,reductive amination reaction happened between carboxymethy chitosan and lactose,and galactose carboxymethyl chitosan was obtained.We took the method of infrared spectrum to characterized the structure of synthetic products,at the same time we measured the degree of substitution of carboxymethyl and galactose group on amino-group by the method of two abrupt change conductometric titration.The result indicated that the reaction of carboxymethyl mainly happened on the group of hydroxyl of C6,and the degree of substitution was about 0.65.While the reaction of galactose happened on amino group of C2,the degree of substitution was 0.20 or so.
Secondly,we prepared the liposomes by the film-ultrasonic wave dissolving techniques.Taking entrapment efficiency as an index,the preparing prescription of liposomes were screened and the process of liposomes were optimized through the orthogonal experiment,using scanning electron microscopy and Zeta potential of ultra-fine-grained and detector for characterization of liposomes.The experimental results showed as followed.The best prescription was that mass ratio of phospholipids /glycyrrhizic acid was 10:1,and the concentration of carboxymethyl chitosan (galactosyl-carboxymethyl chitosan) was 0.3%.When the substitution degree of carboxymethyl was 0.65 and the substitution degree of galactosyl was 0.22,the mean diameter of liposomes modified by carboxymethyl was 122nm,and the zeta electric potential was -53.2mV.While the mean diameter of liposoems that modified by galactosyl was 112nm,and Zeta electric potential was -50.7mV,the morphology of liposomes was more structured and uniform,with smaller particle size distribution,at the same time had a high drug entrapment efficiency,about 80%.
Finally,the dissertation studied liposomal release properties in vitro,investigated different concentrations of material,substitutional degrees and releasing mediuns at different pH values,which had effect on the performance of liposomal releasing properties.Through the experimental study of release in vitro,it was found that glycyrrhizin liposomes modified by both carboxymethyl chitosan and galactosyl-carboxymethyl chitosan showed slow-release and controlled release performance.At preliminary stage of an hour,drug releasing accumulated to 15%on average,then it slowly released drug,in twelve hours drug releasing accumulated to 45%or so.It is showed that the liposomes can build a potential initiative liver-targeted liposome drug release delivery system.And glycyrrhizin liposomes modified by carboxymethyl chitosan have good pH sensitivity of the intellective controlled release characteristics in vitro.
近年来为了改善脂质体的靶向性和体内外稳定性,很多学者对脂质体膜进行表面修饰,即在脂质体的膜材组成中加入一定的辅助剂,如胆固醇、大豆甾醇、聚乙烯醇及聚乙二醇等,增加脂质双层膜的稳定性,提高脂质体体外、体内稳定性。本课题主要研究利用可生物降解、无毒、生物相容性良好的水溶性壳聚糖衍生物对脂质体进行表面修饰,以期构建具有较高稳定性和靶向性的脂质体载药传递系统。主要研究内容和结论如下:
1、以壳聚糖为原料,在碱性条件下与氯乙酸进行醚化反应,合成得到水溶性的羧甲基壳聚糖;在硼氢化钠存在下,羧甲基壳聚糖与乳糖进行还原胺化反应,合成得到半乳糖基羧甲基壳聚糖。采用红外光谱对合成产物进行结构表征,同时通过双突跃电位滴定法对羧甲基取代度和氨基上半乳糖基取代度进行测定。结果表明:羧甲基化反应主要发生在C_6位羟基上,取代度为0.65左右;半乳糖化反应发生在C_2位氨基上,取代度为0.20左右。
2、采用薄膜分散-超声法制备修饰脂质体。以包封率为指标,通过正交实验对脂质体进行制备处方筛选和工艺优化,采用扫描电镜和超细粒度及Zeta电位检测仪对脂质体进行结构表征。结果表明:修饰脂质体的最佳处方为磷脂/甘草酸质量比10:1,羧甲基壳聚糖(半乳糖基羧甲基壳聚糖)浓度为0.3%,在羧甲基取代度为0.65和半乳糖基取代度为0.22时,羧甲基壳聚糖修饰的甘草酸脂质体平均粒径为122nm,Zeta电位为-53.2mV;半乳糖基羧甲基壳聚糖修饰的甘草酸脂质体平均粒径为112nm,Zeta电位为-50.7mV;脂质体形貌规整,粒径较小符合正态分布,同时具有较高的药物包封率,达到80%左右。
3、研究修饰脂质体的体外释药性能,考察修饰材料的不同浓度和取代度,以及不同pH释药介质对修饰脂质体体外释药性能的影响。结果表明:经羧甲基壳聚糖和半乳糖基羧甲基壳聚糖修饰的甘草酸脂质体具有药物缓释控释性能,在释药初期1h内平均累积释药15%左右,随后缓慢释药,12h内平均累积释药45%左右,能够构建稳定性好、具有潜在主动肝靶向性能的脂质体载药控释系统;并且经羧甲基壳聚糖修饰后的甘草酸脂质体具有良好的体外pH敏智能控释性能。
Liposomes have a biological structure similar to bilayer membrane.As drug carriers,they have ability of targeting,delayed release,reducing the dose and drug toxicity.But the targeting of unmodified liposomes focus on organs that are rich in reticuloendothelial cell,while in other parts of organs their targeting are not obvious during the therapy.Meanwhile,traditional liposomes are not stable in vivo and in vitro, easily oxidized and hydrolyzed,and their bilayer membrane will also easily convert the phase,aggregation and fusion will happen between liposomes,then the drug encapsulated in liposomes will easily seepage from it,and so on.
In recent years,in order to improve the targeting and stabiliy in vivo and in vitro of liposomes,many scholars have prepared the liposome membrane of surface modification,which is adding certain auxiliary agents in the composition of membrane material,such as cholesterol,soy sterol,polyvinyl alcohol and polyethylene glycol to increase the stability of lipid bilayer membrane and enhance the stability of liposomes in vitro and in vivo.The study of this dissertation is mainly to use biodegradable,non-toxic,water-soluble,low-molecular-weight and good biocompatibie chitosan derivatives on liposomes surface-modifying,for building a high stability and special targeting liposomal drug delivery systems.Main contents and conclusions are as follows:
Firstly,chitosan as raw materials,hydrosoluble carboxymethyl chitosan was synthesized through the etherealization reaction under basic condition between chitosan and chloroacetic acid.Then,with existence of sodium borohydride,reductive amination reaction happened between carboxymethy chitosan and lactose,and galactose carboxymethyl chitosan was obtained.We took the method of infrared spectrum to characterized the structure of synthetic products,at the same time we measured the degree of substitution of carboxymethyl and galactose group on amino-group by the method of two abrupt change conductometric titration.The result indicated that the reaction of carboxymethyl mainly happened on the group of hydroxyl of C6,and the degree of substitution was about 0.65.While the reaction of galactose happened on amino group of C2,the degree of substitution was 0.20 or so.
Secondly,we prepared the liposomes by the film-ultrasonic wave dissolving techniques.Taking entrapment efficiency as an index,the preparing prescription of liposomes were screened and the process of liposomes were optimized through the orthogonal experiment,using scanning electron microscopy and Zeta potential of ultra-fine-grained and detector for characterization of liposomes.The experimental results showed as followed.The best prescription was that mass ratio of phospholipids /glycyrrhizic acid was 10:1,and the concentration of carboxymethyl chitosan (galactosyl-carboxymethyl chitosan) was 0.3%.When the substitution degree of carboxymethyl was 0.65 and the substitution degree of galactosyl was 0.22,the mean diameter of liposomes modified by carboxymethyl was 122nm,and the zeta electric potential was -53.2mV.While the mean diameter of liposoems that modified by galactosyl was 112nm,and Zeta electric potential was -50.7mV,the morphology of liposomes was more structured and uniform,with smaller particle size distribution,at the same time had a high drug entrapment efficiency,about 80%.
Finally,the dissertation studied liposomal release properties in vitro,investigated different concentrations of material,substitutional degrees and releasing mediuns at different pH values,which had effect on the performance of liposomal releasing properties.Through the experimental study of release in vitro,it was found that glycyrrhizin liposomes modified by both carboxymethyl chitosan and galactosyl-carboxymethyl chitosan showed slow-release and controlled release performance.At preliminary stage of an hour,drug releasing accumulated to 15%on average,then it slowly released drug,in twelve hours drug releasing accumulated to 45%or so.It is showed that the liposomes can build a potential initiative liver-targeted liposome drug release delivery system.And glycyrrhizin liposomes modified by carboxymethyl chitosan have good pH sensitivity of the intellective controlled release characteristics in vitro.
引文
[1]周力,吴肖群.脂质体及其应用.现代化工,1997,9(3):18-20
[2]张树彪,许英梅,胡剑钧等.脂质体研究综述.大连民族学院学报,2004,6(3):6-9
[3]陆彬.药物新剂型和新技术.北京:人民卫生出版社,1998,107-164
[4]Dan D,Lasic.Novel applications of liposomes.Trends Biotechnol,1998,3(16):307-321
[5]张志荣,靶向治疗分子基础与靶向药物设计.北京:科学出版社,2005,14-15
[6]Torchilin VP.Affinity liposomes in vivo:factors influencing target accumulation.J Mol Recognit,1996,9(5):335-346
[7]Lee R J,Huang L.Folate-targeted anionic liposome-entrapped polylysine-condensed DNA for tumor cell-specific gene transfer.J Biol Chem,1996,271(14):8481-8487
[8]张灵芝,凌世长,赵晶等.脂质体导向给药治疗心脑血管疾病的实验研究.北京医科大学学报,1994,26(S1):144-151
[9]Briscoc P,Caniggia I,Graves A,et al.Delivery of supeoxide dismutase to pulmonary eptihlium via pH-sensitive liposomes.Am J PHysiol,1995,268(3):374-380
[10]Chelvi TP,Jain SK,Ralhan R.Hyperthermia-mediated targeted delivery of thermosensitive liposome-encapsulated melpHalan in murine tumors.Oncol Res,1995,7(7):393-398
[11]Chelvi TP,Ralhan R.Hyperthermia potentiates antitumor effect of thermosensitive liposomeencapsulated melpHalan and radiation in murine melanoma.Tumour Biol,1997,18(4):250-260
[12]程建峰,扈本荃,刘梅等.甘草酸脂质体的处方及制备工艺.第四军医大学学报,2006,27(7):655-657
[13]全东琴,苏德森,顾学裘.药物载体空白脂质体前体的制备及性质的研究.沈阳药科大学学报,1999,16(3):160-164
[14]黄黎,李成文.脂质体在医学中的研究进展.国际生物制品学杂志,2006,29(3):130-132
[15]Lian T,Ho RJ.Trends and developments in liposome drug delivery systems.J PHarm Sci,2001,90(6):667-680
[16]李彦超,蔡宝昌.脂质体在抗肿瘤药物研究中的应用.南京中医药大学学报,2003,19(4):250-253
[17]Wu NZ,Da D,Rudull TL,et al.Increased microvascular permeability contributes to preferential accumulation of stealth liposomes in tumor tissue.Cancer Res,1993,53(16):3765-3770
[18]周筱青.无环鸟苷脂质体混悬液的含量分析.中国医院药学杂志,1997,17(3):115-116
[19]Payne NI,Timmins P,Ambrose CV,et al.Proliposomes:a novel solution to an old problem.J PHann Sci,1986,75(4):325-329
[20]Remaut K,Lucas B,Braeckmans K,et al.Protection of oligonucleotides against nucleases by pegylated and non-pegylated liposomes as studied by fluorescence correlation spectroscopy.J Controlled Release,2005,110(1):212-226
[21]Catitel L,Cerutim M,Dosio F.From conventional to stealth liposomes a new forongtier in cancer chemotherapy.Tumor,2003,89(3):237-249
[22]蒋挺大.壳聚糖.北京:化学工业出版社,2007,14-15
[23]Vandevord P,Matthew H,Desilva S,et al.Evaluation of the biocompatibility of a chitosan scaffold in mice.J Biomed Mater Res,2002,59(3):585-590
[24]Senkoylu A,Simsek A,Sahin F,et al.Interaction of cultured chonddrocytes with chitosan scaffold.J Bioact Compat Polym,2001,16(7):136-144
[25]陆广汇.脱乙酰壳多糖作为药用辅料的研究进展.福建医药杂志,2000,22(6):116-117
[26]卢凤琪,曹宗顺.壳聚糖对尼莫地平的缓释作用.中国生化药物杂志,1995,16(5):205-207
[27]Tsai G,Su W.Antibacterial activity of shrimp chitosam against escherichia coli.J Food Prot,1999,62(3):239-243
[28]Struszczyk M.Chitin and chitosan-PartⅢ:Some aspect of biodegradation and bioactivity.PolymJ,2002,47(5):619-629
[29]Cuero R,Osuji G,Washington A.N-carboxymethyl chitosan inhibition of aflatoxin production:role of zinc.Biotechnol Lett,1991,13(6):441-444
[30]高怀生,黄是,张世达等.壳聚糖-诺氟沙星烧伤生物敷料的研究.军事医学科学院院刊,1993,17(3):16-18
[31]王志国,张敏卿等.壳聚糖在靶向制剂中的研究进展.化工进展,2004,23(4):375-379
[32]姚金成,李万忠,胡领.壳聚糖在被动靶向制剂中应用.中医药学刊,2004,22(3):562-564
[33]张庆云,张利平,胡迎庆等.壳聚糖在靶向制剂中的应用进展.天津药学,2004,16(2):56-58
[34]Zheng J,Liu C,Bao D.Preparation and evaluation of floating-bioadhesive microparticles containing clarithromycin for the eradication of helicobacter pylori.J Appl Polym Sci,2006,15(102):2226-2232
[35]Lee M,Heo J,Song C.Radioevaluation of PAMs,CMs,and PS-Lip as an oral carrier for vaccine delivery into intestinal Peyer's patches,Drug Develop Res,2006,18(67):884-889
[36]Kockisch S,Rees G,Young S,et al.Polymeric microspHeres for drug delivery to the oral cavity:An in vitro evaluation of mucoadhesive potential.J PHarm.Sci,2003,23(92):1614-1623
[37]Ishikawa K,Takag S,Chow L.Reaction of calcium pHospHate cements with different amounts of tetracalcium pHospHate and dicalcium pHospHate anhydrous.J Biomed Mater Res,1999,18(46):504-510
[38]BernKop-Schnurch A,Schwarz V,Steininger S..Polymers with thiol groups:a nev generation of mucoadhesive polymers.Pharm.Res,1999,12(16):876-881
[39]Yamamoto H,Kuno Y.Surface-modified PLGA nanospHere with chitosan improved pulmonary delivery of calcitonin by mucoadhesion and opening of the intercellular tight junctions.J Controlled Release,2005,18(102):373-381
[40]张立彦,曾庆孝,李作为等.溶菌酶降解壳聚糖条件的研究.功能高分子学报,2004,17(3):391-395
[41]马如,黄明智.壳聚糖的溶菌酶降解.北京化工大学学报,2002,29(6):40-43
[42]王征,刘万顺,韩宝芹等.低分子量壳聚糖及其衍生物的制备与溶菌酶对其降解的实验研究.海洋科学,2007,31(10):36-40
[43]孙慎侠,付昌斌,范钦信.壳聚糖的生物活性及其在中医药领域中的应用.贵阳中医学院学报,2003,1(25):39-41
[44]易英.壳聚糖衍生物的合成及其性能的研究:[硕士学位论文].武汉:武汉大学化学学院,2005
[45]Abe M,Akbar F,Hasebe A,et al.Glycyrrhizin enhance interleukin-10 production by liver dendritic cells in micewith hepatitis.J Gastroenterol,2003,38(10):962-967
[46]张翠玲.甘草酸药理作用及临床应用.社区医学杂志,2008,6(2):13-14
[47]Yi H,Nakashima I,Isobe K.Enhancement of nitric oxide production from activated macropHages by glycyrrhizin.Am J Chin Med,1996,24(3):271-278
[48]白木公康.甘草酸抗乙肝病毒的作用机理.和汉医药学杂志,1995,12(1):24-25
[49]颜宁.复方甘草酸治疗慢性乙型肝炎的临床疗效观察.中国药房,2004,15(6):355-356
[50]宋惠东,舒建昌,宋方闻等.甘草酸治疗肝硬化疗效观察.中华实用医药杂志,2004,6(4):112-113
[51]涨亚西.复方甘草酸治疗慢性乙型肝炎的临床疗效及抗病毒作用观察.传染病信息,2005,18(2):89-90
[52]陆海英,霍娜,王广发等.复方甘草酸治疗传染性非典型肺炎(SARS)的临床研究.中国药房,2003,14(14):610-612
[53]Ashwell G,Harford J.Carbohydrate-specific receptors of the livery.Annu Rev Biochem,1982,51(1):531-554
[54]Hashida M,Nishikawa M,Yamashita F,et al.Cell-specific delivery of genes with glycosylated carries.Adv Drug Deliv Rev,2001,52(3):187-196
[55]徐颖,周世文.半乳糖介导的肝靶向性研究进展.中国医院药学杂志,2001,21(2):108-110
[56 范锋,孙晓飞.半乳糖介导的肝靶向药物研究进展.中南药学,2007,5(1):62-65
[57]Park Y,Shin B,Choi E,et al.Galactosylated chitosan-graft-dextran as hepatocyte-targeting DNA carrier.J Controlled Release,2000,52(69):97-108
[58]连佳芳,秦葵,徐彪.靶向给药的研究进展.白求恩军医学院学报,2006,4(2):106-108
[59]孔维军,郭伟英.新型强化材料修饰脂质体的研究进展.中国新药杂志,2007,16(19):1565-1568
[60]马宁,汪琴,孙胜玲等.甲壳素和壳聚糖化学改性研究进展.化学进展,2004,16(4):643-653
[61]Pang H T,Chen X G,Park H,et al.Preparation and rheological properties of deoxycholate-chitosan and carboxymethyi-chitosan in aqueous systems.Carbohydr Polym,2007,53(69):419-425
[62]Zhu A P,Jin W J,Yuan L H,et al.O-carboxymethyi chitosan-based novel gatifloxacin delivery system.Carbohydr Polym,2007,52(68):693-700
[63]Sun S L,Wang A Q.Adsorption properties of carboxymethyi-chitosan and cross-linked carboxymethyi-chitosan resin with Cu(Ⅱ)as template.Sep Purif Technol,2006,49(3):197-204
[64]王爱勤.甲壳素化学.北京:科学出版社,2008,186-187
[65]Simoes S,Moreira JN,Fonseca C,et al.On the formulation of pH-sensitive liposomes with long circulation times.Adv Drug Deliv Rev,2004,56(7):947-965
[66]Venugopalan P,Jain S,Sankar S,et al.pH-sensitive liposomes mechanism of triggered release to drug and gene delivery prospects.PHarmazie,2002,57(10):659-671
[67]Hafez M,Ansell S,Cullis PR.Tunable pH-sensitive liposomes composed of mixturea of cationic and anionic lipids.BiopHys J,2000,79(3):1438-1446
[68]张宏娟,丁娅,张灿等.O-羧甲基-N-半乳糖基壳聚糖衍生物的设计、合成和表征.中国天然药物,2004,2(6):354-358
[69]时念秋,马毅,张大同等.长循环脂质体在抗肿瘤药物中的应用进展.中国天然药物,2008,17(3):190-194
[70]Kunihito W,Ikuo S,Yoshihiro M,et al.Antimetastatic activity of neocarzinostain incorporated into controlled release gels of CM-chitin.Carbohydr Polym,1992,17(1):29-37
[71]Komazawa H,Saiki I,Igarashi Y,et al.The conjugation of RGDS peptide with CM-chitin augments the peptide-mediated inhibition of tumor metastasis.Carbohydr Polym,1993,21(4):299-307
[72]Tokura S,Miura Y,Johmen M,et al.Induction of drug specific antibody and the controlled release of drug by 6-O-carboxymethyi-chitin.J Controlled Release,1994,28(13):235-241
[73]Maghami G G,Roberts G A.The molecular weight of chitosans studied by laser lightscattering.CarbohydrRes,1988,28(189):195-200
[74]Vasudevan T K,Ran V S R.Preferred conformations and flexibility of aminoacyl side chain of penicillins.Int J Bio Macromol,1982,4(6):347-351
[75]韩笑,谭伟天.羧甲基壳聚糖制备新工艺研究.北京化工大学学报,2000,27(3):1-4
[76]赵中华,孙海洲,孙明昆.一点法测定羧甲基壳聚糖的特性粘度.海洋水产研究,2002,23(3):41-43
[77]Muzzarelli R A A,Tanfani F.N-(carboxymethylidene)-chitosan and N-(carboxymethyl)-chitosan:novel chelating polyampHolytesobtained from chitosan glyoxylate.Carbohydr Res,1982,4(107):199-214
[78]钟超.N,O-羧甲基壳聚糖的制备及表征:[硕士研究生学位论文].北京:北京化工大学,2004
[79]张灿,丁娅,平其能.半乳糖化季铵壳聚糖衍生物的设计、合成和表征.中国现代应用药学杂志,2005,22(5):386-388
[80]邹晓青.半乳糖基羧甲基壳聚糖纳米粒的制备及其释药性能的研究:[硕士学位论文].武汉:武汉理工大学化学工程学院,2008
[81]王爱勤.甲壳素化学.北京:科学出版社,2008,172-173
[82]Olga B,Anabela C S,Stefan G R,et al.Uptake studies in rat Peyer's patches,cytotoxicity and release studies of alginate coated chitosan nanoparticles for mucosal vaccination.J Controlled Release,2006,32(114):348-358
[83]宴亦林,叶勇,周莉玲等.磷酸川芎嗪微透析体外回收率的研究.中药新药与临床药理,2008,19(2):117-119
[84]于波涛,张志荣,刘文胜.提高脂质体包封率的方法及其研究进展.中国医药工业杂志,2002,33(11):564-568
[85]周丽娟,刘清飞,陈曦等.尼莫地平微乳的包封率和体外释药特性的研究.中国药事,2008.22(7):564-568
[86]Liu H,Tang R,He X X,et al.Effects of liposomes formulation and preparation method on the stablity of acyclovir palmitate liposomes.Acta PHarm Sin,2002,37(7):563-566
[87]Li S H.Study on the liver targeted aclacinomycin A solid lipid nanoparticles[D].Chengdu:Si Chuan Univ,2003
[88]苗彩云,邓树海,陈江飞.氧化苦参碱负电荷脂质体的制备及理化性质研究.中国药学杂志,2006,41(18):1400-1404
[89]杨莉,成丽,田聆等.甘露聚糖修饰的靶向纳米脂质体的抗肿瘤作用实验研究.四川大学学报(医学版),2006,37(3):357-360
[90]毛声俊,候世祥,金辉等.肝细胞靶向甘草次酸表面修饰脂质体的制备.中国中药杂志,2003,28(4):328-331
[91]孙萍,邓树海,于维萍.PEG修饰大蒜素长循环脂质体的制备及药物动力学研究.实用心脑肺血管病杂志,2006,14(6):454-456
[92]邓英杰,陈妍,陈勇等.膜修饰脂质体对缺氧心肌细胞的保护作用.沈阳药科大学学报,2006,23(5):299-302
[93]谭燕,周建平,仝新勇等.F-68修饰后紫杉醇脂质体的大鼠体内药动学.中国药科大学学报,2006,37(3):230-232
[94]林友文,苏燕评,蒋智清.羧甲基壳聚糖水凝胶的pH敏感性及体外释药性能.福建医科大学学报,2005,39(3):331-334
[2]张树彪,许英梅,胡剑钧等.脂质体研究综述.大连民族学院学报,2004,6(3):6-9
[3]陆彬.药物新剂型和新技术.北京:人民卫生出版社,1998,107-164
[4]Dan D,Lasic.Novel applications of liposomes.Trends Biotechnol,1998,3(16):307-321
[5]张志荣,靶向治疗分子基础与靶向药物设计.北京:科学出版社,2005,14-15
[6]Torchilin VP.Affinity liposomes in vivo:factors influencing target accumulation.J Mol Recognit,1996,9(5):335-346
[7]Lee R J,Huang L.Folate-targeted anionic liposome-entrapped polylysine-condensed DNA for tumor cell-specific gene transfer.J Biol Chem,1996,271(14):8481-8487
[8]张灵芝,凌世长,赵晶等.脂质体导向给药治疗心脑血管疾病的实验研究.北京医科大学学报,1994,26(S1):144-151
[9]Briscoc P,Caniggia I,Graves A,et al.Delivery of supeoxide dismutase to pulmonary eptihlium via pH-sensitive liposomes.Am J PHysiol,1995,268(3):374-380
[10]Chelvi TP,Jain SK,Ralhan R.Hyperthermia-mediated targeted delivery of thermosensitive liposome-encapsulated melpHalan in murine tumors.Oncol Res,1995,7(7):393-398
[11]Chelvi TP,Ralhan R.Hyperthermia potentiates antitumor effect of thermosensitive liposomeencapsulated melpHalan and radiation in murine melanoma.Tumour Biol,1997,18(4):250-260
[12]程建峰,扈本荃,刘梅等.甘草酸脂质体的处方及制备工艺.第四军医大学学报,2006,27(7):655-657
[13]全东琴,苏德森,顾学裘.药物载体空白脂质体前体的制备及性质的研究.沈阳药科大学学报,1999,16(3):160-164
[14]黄黎,李成文.脂质体在医学中的研究进展.国际生物制品学杂志,2006,29(3):130-132
[15]Lian T,Ho RJ.Trends and developments in liposome drug delivery systems.J PHarm Sci,2001,90(6):667-680
[16]李彦超,蔡宝昌.脂质体在抗肿瘤药物研究中的应用.南京中医药大学学报,2003,19(4):250-253
[17]Wu NZ,Da D,Rudull TL,et al.Increased microvascular permeability contributes to preferential accumulation of stealth liposomes in tumor tissue.Cancer Res,1993,53(16):3765-3770
[18]周筱青.无环鸟苷脂质体混悬液的含量分析.中国医院药学杂志,1997,17(3):115-116
[19]Payne NI,Timmins P,Ambrose CV,et al.Proliposomes:a novel solution to an old problem.J PHann Sci,1986,75(4):325-329
[20]Remaut K,Lucas B,Braeckmans K,et al.Protection of oligonucleotides against nucleases by pegylated and non-pegylated liposomes as studied by fluorescence correlation spectroscopy.J Controlled Release,2005,110(1):212-226
[21]Catitel L,Cerutim M,Dosio F.From conventional to stealth liposomes a new forongtier in cancer chemotherapy.Tumor,2003,89(3):237-249
[22]蒋挺大.壳聚糖.北京:化学工业出版社,2007,14-15
[23]Vandevord P,Matthew H,Desilva S,et al.Evaluation of the biocompatibility of a chitosan scaffold in mice.J Biomed Mater Res,2002,59(3):585-590
[24]Senkoylu A,Simsek A,Sahin F,et al.Interaction of cultured chonddrocytes with chitosan scaffold.J Bioact Compat Polym,2001,16(7):136-144
[25]陆广汇.脱乙酰壳多糖作为药用辅料的研究进展.福建医药杂志,2000,22(6):116-117
[26]卢凤琪,曹宗顺.壳聚糖对尼莫地平的缓释作用.中国生化药物杂志,1995,16(5):205-207
[27]Tsai G,Su W.Antibacterial activity of shrimp chitosam against escherichia coli.J Food Prot,1999,62(3):239-243
[28]Struszczyk M.Chitin and chitosan-PartⅢ:Some aspect of biodegradation and bioactivity.PolymJ,2002,47(5):619-629
[29]Cuero R,Osuji G,Washington A.N-carboxymethyl chitosan inhibition of aflatoxin production:role of zinc.Biotechnol Lett,1991,13(6):441-444
[30]高怀生,黄是,张世达等.壳聚糖-诺氟沙星烧伤生物敷料的研究.军事医学科学院院刊,1993,17(3):16-18
[31]王志国,张敏卿等.壳聚糖在靶向制剂中的研究进展.化工进展,2004,23(4):375-379
[32]姚金成,李万忠,胡领.壳聚糖在被动靶向制剂中应用.中医药学刊,2004,22(3):562-564
[33]张庆云,张利平,胡迎庆等.壳聚糖在靶向制剂中的应用进展.天津药学,2004,16(2):56-58
[34]Zheng J,Liu C,Bao D.Preparation and evaluation of floating-bioadhesive microparticles containing clarithromycin for the eradication of helicobacter pylori.J Appl Polym Sci,2006,15(102):2226-2232
[35]Lee M,Heo J,Song C.Radioevaluation of PAMs,CMs,and PS-Lip as an oral carrier for vaccine delivery into intestinal Peyer's patches,Drug Develop Res,2006,18(67):884-889
[36]Kockisch S,Rees G,Young S,et al.Polymeric microspHeres for drug delivery to the oral cavity:An in vitro evaluation of mucoadhesive potential.J PHarm.Sci,2003,23(92):1614-1623
[37]Ishikawa K,Takag S,Chow L.Reaction of calcium pHospHate cements with different amounts of tetracalcium pHospHate and dicalcium pHospHate anhydrous.J Biomed Mater Res,1999,18(46):504-510
[38]BernKop-Schnurch A,Schwarz V,Steininger S..Polymers with thiol groups:a nev generation of mucoadhesive polymers.Pharm.Res,1999,12(16):876-881
[39]Yamamoto H,Kuno Y.Surface-modified PLGA nanospHere with chitosan improved pulmonary delivery of calcitonin by mucoadhesion and opening of the intercellular tight junctions.J Controlled Release,2005,18(102):373-381
[40]张立彦,曾庆孝,李作为等.溶菌酶降解壳聚糖条件的研究.功能高分子学报,2004,17(3):391-395
[41]马如,黄明智.壳聚糖的溶菌酶降解.北京化工大学学报,2002,29(6):40-43
[42]王征,刘万顺,韩宝芹等.低分子量壳聚糖及其衍生物的制备与溶菌酶对其降解的实验研究.海洋科学,2007,31(10):36-40
[43]孙慎侠,付昌斌,范钦信.壳聚糖的生物活性及其在中医药领域中的应用.贵阳中医学院学报,2003,1(25):39-41
[44]易英.壳聚糖衍生物的合成及其性能的研究:[硕士学位论文].武汉:武汉大学化学学院,2005
[45]Abe M,Akbar F,Hasebe A,et al.Glycyrrhizin enhance interleukin-10 production by liver dendritic cells in micewith hepatitis.J Gastroenterol,2003,38(10):962-967
[46]张翠玲.甘草酸药理作用及临床应用.社区医学杂志,2008,6(2):13-14
[47]Yi H,Nakashima I,Isobe K.Enhancement of nitric oxide production from activated macropHages by glycyrrhizin.Am J Chin Med,1996,24(3):271-278
[48]白木公康.甘草酸抗乙肝病毒的作用机理.和汉医药学杂志,1995,12(1):24-25
[49]颜宁.复方甘草酸治疗慢性乙型肝炎的临床疗效观察.中国药房,2004,15(6):355-356
[50]宋惠东,舒建昌,宋方闻等.甘草酸治疗肝硬化疗效观察.中华实用医药杂志,2004,6(4):112-113
[51]涨亚西.复方甘草酸治疗慢性乙型肝炎的临床疗效及抗病毒作用观察.传染病信息,2005,18(2):89-90
[52]陆海英,霍娜,王广发等.复方甘草酸治疗传染性非典型肺炎(SARS)的临床研究.中国药房,2003,14(14):610-612
[53]Ashwell G,Harford J.Carbohydrate-specific receptors of the livery.Annu Rev Biochem,1982,51(1):531-554
[54]Hashida M,Nishikawa M,Yamashita F,et al.Cell-specific delivery of genes with glycosylated carries.Adv Drug Deliv Rev,2001,52(3):187-196
[55]徐颖,周世文.半乳糖介导的肝靶向性研究进展.中国医院药学杂志,2001,21(2):108-110
[56 范锋,孙晓飞.半乳糖介导的肝靶向药物研究进展.中南药学,2007,5(1):62-65
[57]Park Y,Shin B,Choi E,et al.Galactosylated chitosan-graft-dextran as hepatocyte-targeting DNA carrier.J Controlled Release,2000,52(69):97-108
[58]连佳芳,秦葵,徐彪.靶向给药的研究进展.白求恩军医学院学报,2006,4(2):106-108
[59]孔维军,郭伟英.新型强化材料修饰脂质体的研究进展.中国新药杂志,2007,16(19):1565-1568
[60]马宁,汪琴,孙胜玲等.甲壳素和壳聚糖化学改性研究进展.化学进展,2004,16(4):643-653
[61]Pang H T,Chen X G,Park H,et al.Preparation and rheological properties of deoxycholate-chitosan and carboxymethyi-chitosan in aqueous systems.Carbohydr Polym,2007,53(69):419-425
[62]Zhu A P,Jin W J,Yuan L H,et al.O-carboxymethyi chitosan-based novel gatifloxacin delivery system.Carbohydr Polym,2007,52(68):693-700
[63]Sun S L,Wang A Q.Adsorption properties of carboxymethyi-chitosan and cross-linked carboxymethyi-chitosan resin with Cu(Ⅱ)as template.Sep Purif Technol,2006,49(3):197-204
[64]王爱勤.甲壳素化学.北京:科学出版社,2008,186-187
[65]Simoes S,Moreira JN,Fonseca C,et al.On the formulation of pH-sensitive liposomes with long circulation times.Adv Drug Deliv Rev,2004,56(7):947-965
[66]Venugopalan P,Jain S,Sankar S,et al.pH-sensitive liposomes mechanism of triggered release to drug and gene delivery prospects.PHarmazie,2002,57(10):659-671
[67]Hafez M,Ansell S,Cullis PR.Tunable pH-sensitive liposomes composed of mixturea of cationic and anionic lipids.BiopHys J,2000,79(3):1438-1446
[68]张宏娟,丁娅,张灿等.O-羧甲基-N-半乳糖基壳聚糖衍生物的设计、合成和表征.中国天然药物,2004,2(6):354-358
[69]时念秋,马毅,张大同等.长循环脂质体在抗肿瘤药物中的应用进展.中国天然药物,2008,17(3):190-194
[70]Kunihito W,Ikuo S,Yoshihiro M,et al.Antimetastatic activity of neocarzinostain incorporated into controlled release gels of CM-chitin.Carbohydr Polym,1992,17(1):29-37
[71]Komazawa H,Saiki I,Igarashi Y,et al.The conjugation of RGDS peptide with CM-chitin augments the peptide-mediated inhibition of tumor metastasis.Carbohydr Polym,1993,21(4):299-307
[72]Tokura S,Miura Y,Johmen M,et al.Induction of drug specific antibody and the controlled release of drug by 6-O-carboxymethyi-chitin.J Controlled Release,1994,28(13):235-241
[73]Maghami G G,Roberts G A.The molecular weight of chitosans studied by laser lightscattering.CarbohydrRes,1988,28(189):195-200
[74]Vasudevan T K,Ran V S R.Preferred conformations and flexibility of aminoacyl side chain of penicillins.Int J Bio Macromol,1982,4(6):347-351
[75]韩笑,谭伟天.羧甲基壳聚糖制备新工艺研究.北京化工大学学报,2000,27(3):1-4
[76]赵中华,孙海洲,孙明昆.一点法测定羧甲基壳聚糖的特性粘度.海洋水产研究,2002,23(3):41-43
[77]Muzzarelli R A A,Tanfani F.N-(carboxymethylidene)-chitosan and N-(carboxymethyl)-chitosan:novel chelating polyampHolytesobtained from chitosan glyoxylate.Carbohydr Res,1982,4(107):199-214
[78]钟超.N,O-羧甲基壳聚糖的制备及表征:[硕士研究生学位论文].北京:北京化工大学,2004
[79]张灿,丁娅,平其能.半乳糖化季铵壳聚糖衍生物的设计、合成和表征.中国现代应用药学杂志,2005,22(5):386-388
[80]邹晓青.半乳糖基羧甲基壳聚糖纳米粒的制备及其释药性能的研究:[硕士学位论文].武汉:武汉理工大学化学工程学院,2008
[81]王爱勤.甲壳素化学.北京:科学出版社,2008,172-173
[82]Olga B,Anabela C S,Stefan G R,et al.Uptake studies in rat Peyer's patches,cytotoxicity and release studies of alginate coated chitosan nanoparticles for mucosal vaccination.J Controlled Release,2006,32(114):348-358
[83]宴亦林,叶勇,周莉玲等.磷酸川芎嗪微透析体外回收率的研究.中药新药与临床药理,2008,19(2):117-119
[84]于波涛,张志荣,刘文胜.提高脂质体包封率的方法及其研究进展.中国医药工业杂志,2002,33(11):564-568
[85]周丽娟,刘清飞,陈曦等.尼莫地平微乳的包封率和体外释药特性的研究.中国药事,2008.22(7):564-568
[86]Liu H,Tang R,He X X,et al.Effects of liposomes formulation and preparation method on the stablity of acyclovir palmitate liposomes.Acta PHarm Sin,2002,37(7):563-566
[87]Li S H.Study on the liver targeted aclacinomycin A solid lipid nanoparticles[D].Chengdu:Si Chuan Univ,2003
[88]苗彩云,邓树海,陈江飞.氧化苦参碱负电荷脂质体的制备及理化性质研究.中国药学杂志,2006,41(18):1400-1404
[89]杨莉,成丽,田聆等.甘露聚糖修饰的靶向纳米脂质体的抗肿瘤作用实验研究.四川大学学报(医学版),2006,37(3):357-360
[90]毛声俊,候世祥,金辉等.肝细胞靶向甘草次酸表面修饰脂质体的制备.中国中药杂志,2003,28(4):328-331
[91]孙萍,邓树海,于维萍.PEG修饰大蒜素长循环脂质体的制备及药物动力学研究.实用心脑肺血管病杂志,2006,14(6):454-456
[92]邓英杰,陈妍,陈勇等.膜修饰脂质体对缺氧心肌细胞的保护作用.沈阳药科大学学报,2006,23(5):299-302
[93]谭燕,周建平,仝新勇等.F-68修饰后紫杉醇脂质体的大鼠体内药动学.中国药科大学学报,2006,37(3):230-232
[94]林友文,苏燕评,蒋智清.羧甲基壳聚糖水凝胶的pH敏感性及体外释药性能.福建医科大学学报,2005,39(3):331-334