论文总字数：34203字

摘 要

柔红霉素（DNR）主要用于各种类型的急性白血病(包括粒细胞性、淋巴细胞性和单核细胞性以及粒-单核细胞性)、红白血病、慢性粒细胞性白血病、恶性淋巴瘤，也可用于神经母细胞病、尤因肉瘤和肾母细胞瘤等。

本课题选用可降解高分子材料聚乳酸羟基乙酸（PLGA）、聚赖氨酸（PLL）、聚乙二醇（PEG）作为合成新型纳米载药系统的基体，通过改善反应条件，制得PLGA-PLL-PEG聚合物。采用该聚合物包载抗白血病药柔红霉素（DNR）及汉防己甲素（Tet）。具体研究内容与结果如下：

⑴通过PLGA的羧基与PLL-CBZ主链氨基反应形成PLGA-PLL-CBZ，使用33%的溴化氢乙酸溶液脱保护得到PLGA-PLL，再使用过量活化的CDI-PEG-CDI与PLGA-PLL侧链的伯氨基反应，最终形成PLGA-PLL-PEG-CDI。通过优化反应条件，并对制备载体进行结构确证，最终高产率（61%）；

⑵建立紫外分光光度法测定DNR含量的方法，空白辅料及Tet对其测定无干扰，回收率为98.70～100.67%，精密度好（小于2%），并建立了高效液相色谱法测定Tet的方法，专一性高，方法的精密度考察RSD均小于2%，重复性试验RSD为0.71%；

⑶选用细胞粉碎仪进行复乳化溶剂挥发法制备共载DNR和Tet的PLGA-PLL-PEG纳米粒，最终产品为一种PLGA-PLL-PEG共聚物纳米粒。其特征在于PLGA与PLL连接为嵌段结构，PLL与PEG连接为接枝结构，PLGA-PLL-PEG聚合物的分子量40000~500000，所述纳米粒子粒径范围为180-230，其平均粒径为213.0±12nm，PI值为0.075，zeta电位为-19.16mv，外观规则圆形。纳米粒中DNR载药量为3.63±0.15%，包封率为70.23±1.91%，Tet载药量为4.27±0.12%，包封率为86.5±0.7%

关键词 PLGA-PLL-PEG 柔红霉素 汉防己甲素 纳米粒

ABSTRACT

Daunorubicin (DNR) is mainly used for various types of acute leukemia(including granulocyte, lymphocyte and monocyte and granulocyte monocyte) , leukemia, chronic myelogenous leukemia, malignant lymphoma, can also be used for neuroblastoma cell disease, Ewing's sarcoma and nephroblastoma.

In our research, we use the biodegradable materials including poly(lactic-co-glycolic acid) (PLGA), poly-L-lysine (PLL), polyethylene glycol (PEG) to synthesis novel nanodrug carrier system. We optimize the reaction conditions to synthesis PLGA-PLL-PEG, then use it to delivery the classical antileukemic drug daunorubicin (DNR) and the MDR reversal agents of tetrandrine (Tet). The contents and results are as follows

⑴We synthesis PLGA-PLL-CBZ through the reaction between carboxyl of PLGA and amino of PLL-CBZ, then use 33% Hydrogen bromide acetic acid to expose the protected primary amines. At last, PLGA-PLL-PEG-CDI is combined by the reaction of excessive CDI-PEG-CDI and primary amines on the side chain of PLGA-PLL. We optimize the reaction conditions and confirm the polymer’s structure, the yield of the reaction is high（61%）.

⑵We establish UV spectrophotometry method to determination the daunorubicin, the recovery rate is between 98.70% to 100.67%, the precision is high(RSDlt; 2%). We also establish HPLC method for the determination of tetrandrine,it has high specificity ,the precision of the method of RSD was less than 2%, repetitive test of RSD was 0.71%

⑶We chose double-emulsion method using the cell crusher to prepare DNR/Tet-loaded PLGA-PLL-PEG nanoparticle. The final product is a kind of PLGA-PLL-PEG copolymer nanoparticles. The featheris that PLGA and PLL connection with block structure, PLL connected with PEG as grafting structure, the molecular weight of PLGA-PLL-PEG polymer 40000~500000, the particle size range of 180-230.its average size was 213.0 ± 12 nm with polydisperse index (PI) of 0.075, and the zeta potential was -19.16 mv. The drug loading of nanoparticles DNR was 3.63 ± 0.15%, encapsulation efficiency was 70.23 ± 1.91%. The drug loading of Tet was 4.27 ± 0.12%, the encapsulation efficiency was 86.5 ± 0.7%.

KEYWORDS: PLGA-PLL-PEG；Daunorubicin；Tetrandrine；Nanoparticles；

目 录

摘要………………………………………………………………………………………I

ABSTRACT……………………………………………………………………………II第一章 研究背景………………………………………………………………………3

1.1肿瘤治疗的现状…………………………………………………………………3

1.2固体脂质纳米粒的现状…………………………………………………………3

1.3 PLGA -PLL-PEG纳米粒载体的现状……………………………………………4

1.4药物介绍…………………………………………………………………………6

1.4.1盐酸柔红霉素……………………………………………………………6

1.4.2汉防己甲素………………………………………………………………6

1.5共聚物纳米粒制备方法研究进展………………………………………………7

1.5.1乳化溶剂挥发法……………………………………………………………7

1.5.2溶剂扩散法…………………………………………………………………8

1.5.3.喷雾干燥法…………………………………………………………………8

第二章 制备及分析PLGA-PLL-PEG-CDI共聚物 ………………………………9

2.1背景资料…………………………………………………………………………9

2.2实验内容…………………………………………………………………………9

2.2.1实验材料与仪器 …………………………………………………………9

2.2.2 PLGA-PLL-PEG-CDI合成路线图 ………………………………………10

2.2.3 Lys(Z)-NCA聚合成PLL-CBZ……………………………………………11

2.2.4 PLGA-PLL-CBI的合成 …………………………………………………11

2.2.5 PLGA-PLL-CBI脱保护 …………………………………………………11

2.2.6 PLGA-PLL-PEG-CDI的合成 ……………………………………………12

2.3实验结论………………………………………………………………………12

第三章 建立柔红霉素及汉防己甲素体外分析方法…………………………13

3.1背景资料………………………………………………………………………13

3.2实验内容………………………………………………………………………13

3.2.1实验材料与仪器 ………………………………………………………13

3.2.2 盐酸柔红霉素及汉防己甲素最大吸收波长的确定…………………14

3.2.3 柔红霉素体外分析方法的建立………………………………………14

3.2.4 汉防己甲素体外分析方法的建立……………………………………14

3.3 实验分析 ……………………………………………………………………16

3.3.1汉防己甲素、柔红霉素最大吸收波长的确定…………………………16

3.3.2柔红霉素体外分析方法的建立 ………………………………………16

3.3.3汉防己甲素体外分析方法的建立 ……………………………………18

3.4实验结论………………………………………………………………………23

第四章 PLGA-PLL-PEG纳米粒的制备及质量研究…………………………24

4.1背景资料………………………………………………………………………24

4.2实验内容………………………………………………………………………24

4.2.1实验材料与仪器 ………………………………………………………24

4.2.2载药纳米粒的制备（复乳化溶剂挥发法）……………………………25

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