甘草次酸衍生物受体靶向复方脂质体的制备及其体外释放研究

目的：合成两亲性导向分子甘草次酸衍生物3-琥珀酸-30-硬脂醇甘草次酸酯（18-GA-Suc）,研究其掺入甘草次酸-丹参酮ⅡA-丹酚酸B复方脂质体（GTS-Lip）的制备工艺,并考察其体外释放规律。方法：采用1HNMR和13CNMR表征18-GA-Suc的结构,通过单因素考察确定18-GA-Suc的投料量,低速离心法测定其掺入比率,比较修饰前后复方脂质体的理化性质,以平衡透析法考察甘草次酸衍生物受体靶向的甘草次酸-丹参酮ⅡA-丹酚酸B复方脂质体（Suc-GTS-Lip）中3种成分的体外释放规律。结果：优化的处方工艺为：18-GA-Suc投料量为膜材的10%（mol·mol-1）,掺入比率为96.58%。Suc-GTS-Lip形态圆整,分布均匀,其中甘草次酸（GA）、丹参酮ⅡA（TSN）和丹酚酸B（SalB）的平均包封率分别为86.15%,81.70%,91.05%,平均粒径为128.7 nm,平均Zeta电位为-15.5mV。GA和TSN体外释放规律符合Higuchi方程,SalB体外释放规律符合Hixon-crowell方程。结论：甘草次酸衍生物（18-GA-Suc）能在脂质体膜上成功表达,本脂质体制备工艺稳定,GA、TSN和SalB 3种成分在体外均具有一定的缓释作用,为进一步研究其肝靶向性奠定了基础。     

甘草次酸衍生物配体18-GA-Ala介导脂质体的制备及肝靶向性研究

目的:合成制备甘草次酸衍生物配体丙氨酸丁二酸甘草次酸十八烷酯(18-GA-Ala),并修饰包埋药物丹酚酸B(Sal B)和丹参酮ⅡA(TSN)的脂质体,探讨该脂质体对小鼠的肝靶向作用。方法:采用薄膜分散-高压乳匀法制备18-GA-Ala配体修饰的TSN-Sal B脂质体及未修饰配体的脂质体,考察粒径、电位、包封率、多分散系数和配体结合率;尾静脉给药后不同时间点获取血浆样本及小鼠心、肝、脾、肺、肾组织样本,超高效液相色谱法测定各样本中Sal B和TSN的含量,评价18-GA-Ala配体的肝靶向效果。结果:配体18-GA-Ala修饰后的脂质体中,Sal B在肝脏的药时曲线下面积(AUC)分别是脾、肺和肾的1.19、1.39、63.67倍,TSN在肝脏的AUC分别为脾、肺、肾的4.64、0.36、14.12倍。结论:甘草次酸衍生物配体18-GA-Ala修饰后的脂质体可增加Sal B和TSN在肝脏中的峰浓度,表现出一定的肝靶向作用。     

甘草次酸衍生物18-GA-Gly介导肝主动靶向脂质体的制备及体内相关研究

基于国内外对甘草次酸配体介导主动肝靶向脂质体的研究情况,该文主要对以甘草次酸为母核的肝靶向配体18-GA-Gly修饰的脂质体的肝靶向效果进行研究。采用薄膜分散-高压乳匀法制备18-GA-Gly配体介导的丹参酚酸B-丹参酮ⅡA脂质体及未修饰配体的脂质体,考察2种脂质体的制剂学性质,考察粒径、电位、包封率和配体结合率;尾静脉给药后不同时间点获取血浆样本以及小鼠心、肝、脾、肺、肾组织样本,UPLC测定各样本中丹参酚酸B(Sal B)和丹参酮IIA(TSN)含量,评价18-GA-Gly配体的肝靶向效果。结果显示脂质体Gly-TS-Lip和TS-Lip的粒径、电位、包封率、配体结合率基本符合要求;体内靶向性考察结果显示Gly-TS-Lip组脂质体与TS-Lip组脂质体血浆数据无显著性差别;甘草次酸衍生物配体18-GA-Gly介导脂质体可以增加Sal B和TSN在肝组织的峰浓度,但并未显示明显的肝靶向效果。     

甘草次酸修饰的长循环阳离子脂质体DNA复合物的制备

 目的 合成甘草次酸-聚乙二醇-胆固醇偶合物,并运用该材料制备具有肝细胞靶向作用的阳离子脂质体DNA复合物。方法 丁二酰化甘草次酸和丁二酰化胆固醇与聚乙二醇通过酯键偶联,合成甘草次酸-聚乙二醇-胆固醇偶合物,并通过芘荧光探针法测定其临界胶束浓度;采用前、后聚乙二醇化法制备阳离子脂质体DNA复合物,以粒径为指标优选制备工艺。结果 甘草次酸-聚乙二醇-胆固醇偶合物是一种具有低临界胶束浓度的载体材料,临界胶束质量浓度为5.9×10-4～3×10-3g·L-1;前、后聚乙二醇化法制备的阳离子脂质体DNA复合物粒径分别为（297.9±8.16,（178.7±5.4nm,Zeta电位分别为（28.34±1.23,（26.72±0.52mV。结论 具有低临界胶束浓度的甘草次酸-聚乙二醇-胆固醇偶合物合成工艺简单、成本低廉,宜用于聚乙二醇化法制备具有潜在肝细胞靶向的阳离子脂质体DNA复合物。     

活体成像系统检测甘草次酸修饰脂质体在小鼠体内的分布

目的:研究胆固醇-聚乙二醇-甘草次酸(Chol-PEG-GA)修饰脂质体(CPGL)在小鼠体内的长循环型和分布情况,为肝肿瘤的靶向治疗提供理论依据.方法:将普通脂质体(LP)和CPGL分别包裹荧光显像剂NIRD-15,通过NIRD-15跟踪检测LP和CPGL在小鼠体内分布情况.将小鼠随机分为LP组和CPGL组,分别尾静脉注射,与药物注射后5,10,15,30,60,120,180,360,600 min采用活体成像系统观察小鼠活体及离体器官中药物荧光强度.结果:尾静脉注射LP后随循环系统迅速分布到各器官,肝靶向性较差.尾静脉注射CPGL 15 min后,药物开始聚集在肝脏,600 min后在肝脏中的药物浓度仍显著高于LP组,说明用Chol-PEG-GA修饰的脂质体能主动靶向于肝细胞,并延长其在肝组织的滞留时间.结论:CPGL在体内有明显的长循环性和肝靶向性,延长其在体内的半衰期,减少药物代谢率,靶向到达肝脏,延长在肝脏的滞留时间.     

甘草次酸固体脂质纳米凝胶的制备及体外透皮效应

目的制备甘草次酸固体脂质纳米凝胶并考察其体外透皮效应。方法采用微乳液法制备甘草次酸固体脂质纳米粒并考察其包封率、粒径与表面电位,以研和法制备固体脂质纳米粒凝胶;采用改良Franz立式扩散池法进行体外透皮实验,HPLC法测定甘草次酸,评价甘草次酸固体脂质纳米粒凝胶的经皮渗透结果。结果甘草次酸固体脂质纳米粒外观为圆球形或椭球形;甘草次酸固体脂质纳米粒的包封率为(64.75±1.36)%,粒径范围(46.13±20.10)nm,电位分布范围为(-53.4±7.11)mV。24 h甘草次酸固体脂质纳米粒凝胶较甘草次酸固体脂质纳米粒的累积透过量提高66%。结论甘草次酸固体脂质纳米粒凝胶能提高甘草次酸的透皮速率,有望成为甘草次酸透皮给药的新型制剂。     

甘草次酸及其衍生物在肝靶向载药系统中的应用

目前临床上肝炎、肝纤维化、肝硬化和肝癌等肝脏疾病已成为多发病和常见病。开发肝靶向载药系统是肝病治疗的研究热点。甘草次酸(GA)是甘草的主要有效成分之一,研究表明肝细胞膜上存在能够与甘草次酸特异性结合的位点。对以甘草次酸及其衍生物为载体的靶向前药、脂质体、纳米粒等肝靶向载药系统的最新研究进展进行综述。     

甘草次酸修饰的人参皂苷Rh2脂质体的制备及其体外评价

目的:制备由甘草次酸修饰的人参皂苷Rh2脂质体,评价其体外理化性质和对人肝癌细胞SMMC-7721的生长抑制作用。方法:采用薄膜分散法制备普通的人参皂苷Rh2脂质体(Rh2-L)和甘草次酸修饰的人参皂苷Rh2脂质体(GA-Rh2-L),利用UPLC测定人参皂苷Rh2的含量,流动相乙腈-水(28∶72),流速0.3 m L·min-1,检测波长203 nm。考察脂质体的包封率、粒径、体外释放率及Zeta电位,运用噻唑蓝(MTT)法评价人参皂苷Rh2溶液,Rh2-L和GA-Rh2-L对SMMC-7721细胞体外增殖的抑制作用。结果:与Rh2-L相比,GA-Rh2-L的包封率、粒径、体外释放率及Zeta电位等理化性质均无显著性差异。Rh2-L和GARh2-L的包封率分别为(91.67±1.05)%,(95.54±2.23)%,粒径(138.6±45.8),(146.5±48.9)nm,Zeta电位-(12.75±0.34),-(14.79±0.67)m V,72 h的体外释放率(84.67±7.23)%,(89.03±8.61)%。人参皂苷Rh2溶液在12 h内释药率达(91.23±5.17)%。细胞用药2 d后,GA-Rh2-L对SMMC-7721的半抑制率(IC50)分别为人参皂苷Rh2和Rh2-L的0.524,0.596倍,并呈浓度和时间以依赖关系。结论:甘草次酸修饰不会影响人参皂苷Rh2脂质体得理化性质,还可增加其对细胞的靶向性,增加了细胞毒性,为肝肿瘤的靶向治疗提供了新思路。     

羟基喜树碱磁性脂质体的制备及其靶向性特征试验

目的:优选羟基喜树碱(HCPT)磁性脂质体的制备工艺并验证其体内靶向聚集性.方法:以经油酸表面改性的Fe3O4磁性纳米粒作为材料,大豆卵磷脂、胆固醇为原料,利用薄膜分散-超声法制备羟基喜树碱磁性脂质体,以包封率为评价指标,采用正交试验法对羟基喜树碱脂质体的制备工艺进行优选.引入微透析技术,动态、连续地考察磁性脂质体的肿瘤局部聚集性.结果:羟基喜树碱磁性脂质体的最佳制备工艺为药脂比25:1,磷脂-胆固醇80:30,制备温度45 ℃,水化介质pH6.8的磷酸盐缓冲液(PBS).在此条件下,最高包封率66.8％,载药量2.57％.在荷瘤小鼠体内磁靶向定位试验中,HCPT磁性脂质体外加磁场组的肿瘤部位药物浓度较不加磁场组和注射液组高,表明HCPT磁性脂质体在外加磁场引导下具有明显的磁靶向性效果.结论:使用油酸改性的Fe3O4联合应用薄膜分散-超声法制备的HCPT磁性脂质体粒径均一,稳定分散,包封率较高,磁靶向性好.     

甘草次酸脂质体的制备及其药剂学性质的研究

目的研究甘草次酸阳离子脂质体的制备方法并考察其药剂学性质。方法采用正交设计筛选处方,乙醇注入法制备甘草次酸脂质体;用葡聚糖凝胶G-50柱分离脂质体和游离药物,用HPLC法测定包封率;用透射电镜观察脂质体的外观形态,并用粒径分析仪测定脂质体的粒径和zeta电位;进一步考察脂质体的释放规律。结果所得脂质体包封率为(91.61±1.16)%;形态为粒径均匀的球形和类球形,粒径为(141±10)nm,Zeta电位为(35.9±5)mV;脂质体的体外释放符合Higuchi方程;具有较好的稳定性。结论优选得到的甘草次酸脂质体处方和制备工艺合理、稳定,其体外释放具有缓释特点。     

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??OBJECTIVE To evaluate the physical stability of glycyrrhetinic acid derivatives-mediated coumarin 6(Cou6) liposomes and confirm the applicability of different stability tests on liposomes. METHODS Film dispersion-ultrasonic method was used to prepare Cou6 liposomes, PEG-modified liposomes and glycyrrhetinic acid-mediated liposomes. The stability constants, membrane stability, serum stability and leakages of the six kinds of liposomes were studied. RESULTS The physical stability of the liposomes without modification was poor. As for the glycyrrhetinic acid-mediated liposomes, the stability constants at 15 min were 5.37-7.32 and the concentrations of Triton X-100 were 0.207??-0.380?? when half liposome membranes were destroyed. The serum stability in 24 h and leakages in 7 or 14 d showed good stability with little change. CONCLUSION The physical stability is one of the key pharmaceutical properties of liposomes. The stability constant, serum stability and leakage tests and the method of membrane stability we have established can be used to study the stability of liposomes.     

酶促合成3种半乳糖-胆固醇配体及其与肝靶向性的构效关系研究

目的酶促法构建3种不同类型的半乳糖-胆固醇配体修饰的阿霉素(doxorubicin,DOX)脂质体,研究其在小鼠体内的药动学及组织分布特征。方法利用脂肪酶在非水相中合成了3种不同结构的半乳糖-胆固醇配体:CHS-C8-Gal NAc、CHS-C8-GAL、CHS-C8-LA;利用MS、NMR鉴定产物结构;采用薄膜分散-梯度载药法制备DOX脂质体并对其理化性质进行表征;以小鼠为实验对象,采用尾iv给药,通过比较3种配体修饰的DOX脂质体在小鼠体内的药动学与组织分布特征来阐明配体中半乳糖基的结构与去唾液酸糖蛋白受体之间的构效关系。结果产物经MS、NMR鉴定均为目标产物,产率均90%;采用薄膜分散-梯度载药法制备的DOX脂质体粒径90 nm,PDI0.1,包封率99%,24 h渗漏率5%,Zeta电位接近于0;3种配体分子对肝脏亲和力大小依次为CHS-C8-Gal NAcCHS-C8-LACHS-C8-Gal;肝脏对CHS-C8-Gal NAc修饰的脂质体的摄取几乎完全被预注射的asialofetuin所抑制(P0.01),而对CHS-C8-Gal及CHS-C8-LA修饰的脂质体无显著的抑制效果(P0.05)。结论含有N-乙酰半乳糖胺基(N-acetylgalactosamine,Gal NAc)为末端的配体分子能被去唾液酸糖蛋白受体高效识别,而含有D-半乳糖基(D-galactose,Gal)或乳糖醇基(lactitol,LA)为末端的配体分子易被肝枯否细胞上的半乳糖粒子受体所竞争性结合。     

甘草次酸脂质体中乙醇残留量的顶空气相色谱法测定

目的建立甘草次酸脂质体中乙醇残留量的气相色谱测定方法。方法采用毛细管气相色谱法,色谱柱为HP-5,载气为氮气,检测器为FID,柱温采用程序升温,初始温度为70℃,保持3 min,以5℃.min-1升温至90℃,再以10℃.min-1升温至200℃,保留5 min,顶空进样,进样体积为1 ml。结果乙醇在20.1～1 005 mg.L-1内线性关系良好,r=0.999 3(n=3),最低检测限和定量限分别为0.63 mg.L-1和2.51 mg.L-1。高、中、低浓度的回收率分别为97.94%,99.46%和96.56%。结论该毛细管气相色谱法灵敏、准确、可靠,适用于甘草次酸脂质体中乙醇残留量的测定。     

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??OBJECTIVE To develop a kind of multifunctional targeting epirubicin liposomes for the treatment of brain tumor, to characterize their physicochemical properties, and to observe their targeting effects on the brain microvascular endothelial cells(BMVECs)and on the brain glioma cells.METHODS The 2-amino-2-deoxy-??-D-glucopyranose(NH₂-Glu)was used as a targeting molecule and conjugated with a cholesterol-polyethylene glycol derivative (Chol-PEG₂₀₀₀-NHS) for obtaining the targeting functional material aimed at targeting to glucose transporter-1(Glut-1)on the BMVECs of blood-brain barrier and further targeting to the glioma cells. To prepare the multifunctional targeting epirubicin liposomes, the targeting functional material was modified onto the surface of liposomes, and epirubicin was loaded into the core of liposomes as the anticancer drug. The encapsulation efficiency, particle size, polydispersity indexes and Zeta potential of the liposomes were measured, their cellular uptakes were performed on the BMVECs and the glioma cells. The inhibitory effect was performed on the glioma cells. RESULTS The analysis by MALDI-TOF-MS demonstrated that the targeting functional material, Chol-PEG₂₀₀₀-Glu, was successfully synthesized. The multifunctional targeting epirubicin liposomes were prepared, and had an average particle size of approximately 125 nm, and were negatively charged. The encapsulation efficiency of epirubicin in the liposomes was about 93%. Results from flow cytometry indicated that the multifunctional targeting epirubicin liposomes had the highest cellular uptakes by BMVECs and by two kinds of brain glioma cells as compared with no-targeting epirubicin liposomes. The cytotoxic study showed that the multifunctional targeting epirubicin liposomes had the strongest inhibitory effect to brain glioma cells as compared with free epirubicin or no-targeting epirubicin liposomes. CONCLUSION A new targeting material (Chol-PEG₂₀₀₀-Glu) and the multifunctional targeting epirubicin liposomes are developed, and the multifunctional targeting epirubicin liposomes exhibit the potential fortransporting across the blood-brain barrier (BBB), and selectively inhibiting the brain glioma cells.