微卡計於液膜交互作用力量測之應用----微脂粒系統穩定度之探討

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劉得任(De-Rin Liu) 查詢紙本館藏

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化學工程與材料工程學系

論文名稱

微卡計於液膜交互作用力量測之應用----微脂粒系統穩定度之探討

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摘要(中)

關於微脂粒聚集融合行為之研究，微脂粒脂雙層結構以及微脂粒粒子間交互作用力兩者都扮演著一相當重要之影響，本研究利用流場剪應力以及脂雙層磷脂質游離釋放實驗討論微脂粒脂雙層結構之穩定性，同時利用恆溫滴定微卡熱量計測量微脂粒懸浮液之稀釋熱並將稀釋熱配合簡單數學模式之演譯計算找出微脂粒懸浮液系統之第二維里係數b2，利用第二維里係數觀點說明微脂粒粒子間交互作用行為，此外更配合Square-Well 能井方程式說明微脂粒粒子間聚集融合行為之能量障壁參數(b2/B0)。
就微脂粒粒子間交互作用力，實驗結果顯示PC微脂粒粒子間淨交互作用力場為斥力場，而且其有效斥力場範圍可遠及其自身半徑長之80倍(約8

摘要(英)

The behavior aggregation and fusion of loposome, both the structure of lipid bilayer and the interactions between liposomes play a very important role. In this research, the stability of lipid bilayer structure is discussed by using the shear stress coupled with the lipid released from the liposomal bilayer. At the same time, the dilution heat of liposomal suspension can be measured by isothermal titration calorimetry (ITC). In addition, the second virial coefficient b2 in the system of liposomeal suspension are also determined by using a mathematical model plus the measurement data from dilution heat, moreover, the behavior of interactions between liposomes can be explained from the standpoint of the second virial coefficient. Furthermore, the energy barrier parameter (b2/B0) relating to the aggregation and fusion behavior of liposomes can be interpreted in conjunction with Square-Well energy equation.
Our experimental data indicates that the net interaction force between PC liposomes are repulsive. Also, the effective range of repulsive force between PC liposomes can reach as faraway as 80 times of their diameter (around 8 x 10-4cm). So far as the incorporation of cholesterol into the PC bilayer, our experimental results also indicates that the absolute zeta potential becomes more negative, this data indicate that the repulsive electrostatic force between liposomes have become larger. In addition, the values of b2 and b2/B0 have become larger as well. Another way of explaining this data that incorporating cholesteril into the PC bilayer promotes the net repulsive force (2.5 times of its effective repulsive force) between PC liposomes, in addition to the increment of energy barrier in respect of the behavior of aggregation and fusion. Moreover, incorporating cholesterol into the PC bilayer make the liposome more rigid and sustains more severs shear stress in our shear forces studies; in other words, incorporating cholesterol into the PC bilayer can promote the stability of PC bilayer structure. More importantly, incorporating cholesterol into the bilayer can reduce the amount of free lipid, pointing out that this incorporation can increase the interactive force of lipid molecules and enhancing the stability of PC bilayer. Finally, our experimental results also indicated that the incorporation of cholesterol into the bilayer would reduce the liposomal size changes. In other words, incorporation of cholesterol can inhibite the behavior of aggregation and fusion and enhance the physical stability of liposome.
This work also reveals that temperature and PC liposomal stability are correlated with each other. The stability of PC liposomes are likely to decline with a rise in the temperature, and the elevated temperature effect is evidence of the rigidity or curvature of the PC liposome. Furthermore, the repulsive interaction potential (or energy barrier) between the PC liposome declines with a rise in temperature
The second part of this research is to examines how PEG2000-grafted lipid affect the surface properties of the PC liposomal bilayer membrane by the zeta potential and interaction potential measurement by microcalorimetry. In this studies Liposome were prepared from mixtures of egg- phosphatidylcholine (PC), cholesterol and distearoyl-phosphatidylethanolamine covalently attached poly (ethylene glycol) of molecular weight 2000 (DSPE-PEG2000).
Experimental results demonstrated that the absolute value of the zeta potential of PEG2000-grafted liposome decreased from

關鍵字(中)

★ 物理穩定度
★ 第二維里係數
★ 膽固醇
★ 界面位能
★ 交互作用力
★ 微脂粒

關鍵字(英)

論文目次

中文摘要………………………………………………………………I
英文摘要……………………………….……………………………IV
目錄………………………………………………………….….………VIII
圖表索引……………………………………………………….….…IX
符號說明……………………………………………….……………XII
第壹章: 前言………………………………………………………1
第貳章: 研究原理……………………………………………...4
2-1 微脂粒之簡介…………………………………….……………4.
2-1-1 微脂粒之組成…………………………………….……….4
2-1-2 微脂粒之形成與機制………………………..…………12
2-1-3 微脂粒之製備方法………………………………………17
2-1-4 微脂粒的物理結構分類………………………………..21
2-1-5 微脂粒之代謝機制─微脂粒與細胞作用機制….23
2-2 微脂粒膜之特性…………………………………………….26
2-2-1 相轉移………………………………………………………26
2-2-2 相分離………………………………………………………29
2-3 微脂粒之穩定度……………………………………………30
2-3-1微脂粒之生理穩定度…………………………………..30
2-3-2微脂粒之化學穩定度…………………………………30
2-3-2-1氧化………………………………………………….31
2-3-2-2水解…………………………………………………..32
2-3-3 微脂粒之物理穩定度…………………………………36
2-3-3-1 微脂粒的聚集……………………………………..38
2-3-3-2 微脂粒的融合………………………………………40
2-3-3-3 陽離子添加對微脂粒物理穩定性之影響…..43
2-4 微脂粒懸浮液系統微脂粒粒子間交互作用……….52
2-4-1 微脂粒脂雙層間之交互作用力………………………..52
2-4-1-1 凡得瓦爾引力…………………………………………52
2-4-1-2 庫侖靜電斥力…………………………………………54
2-4-1-3 水合作用力…………………………………………….58
2-4-1-4 立體空間力…………………………………………….61
2-4-2 添加物對微脂粒脂雙層(或微脂粒)間之交互作用
影響…………………………………………………………..63
2-4-2-1 膽固醇之影響……………………………………….63
2-4-2-2 親水性高分子(Lipid-PEG)之影響…………..69
2-4-3 微脂粒粒子間之交互作用力………………………… 71
2-4-4 膠體懸浮液中膠質粒子之交互作用……………75
2-4-4-1 維里方程式及維里係數………………………76
2-4-4-2 膠體懸浮液系統膠質粒子第二維里係數數學
模式之討論………………………………………77
2-4-4-3 膠體懸浮液系統膠質粒子第二維里係數與膠
質粒子間交互作用之討論……………………86
第參章: 研究動機暨實驗目的………………………90
第肆章：實驗設備與實驗設計………………………96
4-1 實驗設備…………………………………………………………96
4-1-1 實驗藥品……………………………………………………96
4-1-1-1緩衝溶液之配製…………………………………….96
4-1-1-2 微脂粒之製備……………………………………….96
4-1-1-3 其它化學藥品……………………………………….97
4-1-2 儀器設備…………………………………………………….98
4-1-2-1黏度儀………………………………………………….98
4-1-2-2 恆溫滴定微卡熱量計…………………………….101
4-1-2-3 其它儀器設備……………………………………..104
4-2 實驗設計…………………………………………………106
4-2-1 游離磷脂質之釋放……………………………..106
4-2-2 剪應力實驗……………………………………….108
4-2-3 微脂粒界面電位及粒徑之量測…………….108
4-2-4 微脂粒懸浮液之稀釋熱-恆溫滴定微卡熱量計
…………………………………………………..………….109
4-2-5 第二維里系數之計算…………………………110
第伍章：實驗結果與討論………………………..112
5-1 膽固醇之嵌入微脂粒脂雙層對微脂粒之影響
……………………………………………………………….……..114
5-1-1 磷脂質(PC）游離釋放之影響…………………114
5-1-2 剪應力流場對微脂粒脂雙層之磷脂質游離釋放之
影響…………………………………………………..117
5-1-3 剪應力(shear stress)對微脂粒結構之影響.122
5-1-4 微脂粒界面電位……………………………………123
5-1-5 微脂粒粒子交互作用-ITC量測微脂粒懸浮液之稀
釋熱實驗部份………………………………………126
5-1-6 微脂粒之粒徑變化………………………………….132
5-2 PEG-lipid之嵌入微脂粒脂雙層對微脂粒之
影響……………………………………………………….140
5-2-1 剪應力(shear stress)對微脂粒結構之影響140
5-2-2 界面電位…………………………………………………142
5-2-3 微脂粒粒子交互作用-ITC量測微脂粒懸浮液之稀
釋熱實驗部份………………………………………….143
5-2-4 微脂粒之粒徑變化……………………………………..146
第陸章結論……………………………………………………….150
參考資料………………………………………………………………153
附錄…………………………………………………………………… 176
圖表索引
圖2.1 微脂粒之結構示意圖…………………………………….4
圖2.2 磷脂質的結構示意圖…………………………………….5
圖2.3 磷脂基膽鹼示意圖………………………………………..6
圖2.4 磷脂基乙醇氨示意圖…………………………………….7
圖2.5 磷酸基甘油示意圖………………………………………..9
圖2.6 常見磷脂質之結構示意圖……………………………..11
圖2.7 微脂粒形成機制示意模型圖…………………………..16
圖2.8 超音波震盪法製備微脂粒示意圖…………………….19
圖2.9 微脂粒的物理結構分類示意圖………………………..23
圖2.10 磷脂質脂肪酸鏈的反式、鈍式結構圖……………….26
圖2.11 磷脂質的DSC圖形………………………………………..29
圖2.12 脂質的氧化反應機制示意圖…………………………….32
圖2.13 磷脂醯膽鹼(PC)的水解反應機制………………………33
圖2.14 pH值對磷脂質水解速率的影響……………………….35
圖2.15 膠體粒子之距離─位能示意圖…………………………..37
圖2.16 微脂粒聚集融合程序示意圖……………………………..42
圖2.17 膠體粒子layer 分佈示意圖………………………………55
圖2.18 膽固醇嵌入於脂雙層中位置結構示意圖……………..64
圖2.19 膽固醇嵌入於脂雙層中對相轉移溫度之影響………65
圖2.20 脂雙層間作用力量測的儀器示意圖……………………66
圖2.21 微脂粒粒子間之交互作用距離-位能示意圖…………74
圖4.1 Cone/Plate 黏度儀及水浴法控溫示意圖……………100
圖4.2 恆溫滴定微卡計-恆溫部分儀器外觀圖……………..103
圖4.3 恆溫滴定微卡計-注射與反應器之外觀圖…………..104
圖5.1 脂雙層中不同嵌入膽固醇量於不同溫度系統下，隨
時間變化磷脂質游離釋放圖…………………………….116
圖5.2 脂雙層中不同嵌入膽固醇量於277K儲存溫度環境
下受其到剪應率流場300sec-1，五分鐘環境下，隨
貯存時間變化磷脂質游離釋放圖……………………119
圖5.3 雙層中不同嵌入膽固醇量於277K儲存溫度環境下
受其到剪應率流場800sec-1，五分鐘環境下，隨貯
存時間變化磷脂質游離釋放圖……..……………….120
圖5.4 脂雙層中不同嵌入膽固醇量於310K儲存溫度環境
下受其到剪應率流場300sec-1，五分鐘環境下，隨
貯存時間變化磷脂質游離釋放圖……………………121
圖5.5 不同剪應力對不同嵌入膽固醇莫爾量PC微脂粒其
於不同溫度下粒徑之變化……………………………..123
圖5.6 微脂粒懸浮液於ITC稀釋熱之圖譜……………….130
圖5.7 儲存溫度277K下，不同莫爾含量膽固醇之PC微
脂粒其隨儲存時間之粒徑變化圖………………….136
圖5.8 儲存溫度298K下，不同莫爾含量膽固醇之PC
微脂粒其隨儲存時間之粒徑變化圖……………….137
圖5.9 儲存溫度310K下，不同莫爾含量膽固醇之PC微
脂粒其隨儲存時間之粒徑變化圖………………….138
圖5.10 不同剪應力對不同DSPE-PEG2000量微脂粒其於不
同溫度下，粒徑之變化……………………………….141
圖5.11 儲存溫度298K下，不同含量DSPE-PEG2000之微
脂粒其隨時間變化之粒徑變化圖………………..148
圖5.12 儲存溫度310K下，不同含量DSPE-PEG2000之微
粒其隨時間變化之粒徑變化圖……………………149
表2.1 不同碳鏈與頭基之磷脂質的相轉移溫度(Tc) ………28
表5.1 不同嵌入膽固醇量之微脂粒於不同溫度下之界面電
位值……………………………………………………………125
表5.2 不同嵌入膽固醇量之微脂粒於不同溫度下之b2, B0
及b2 / B0值…………………………………………………..131
表5.3 膽固醇之嵌入與無膽固醇嵌入之微脂粒系統其b2值
比較表………………………………………………………..131
表5.4 相同組成之微脂粒於不同溫度下其b2值比較表…132
表5.5 298K系統下，不同膽固醇莫爾含量之PC微脂粒其
隨時間變化之b2值……………………………………….139
表5.6 310K系統下，不同膽固醇莫爾含量之PC微脂粒其
隨時間變化之b2值……………………………………….139
表5.7 不同嵌入DSPE-PEG2000量之微脂粒於不同溫度下
之界面電位值………………………………………………143
表5.8 不同嵌入DSPE-PEG2000量之微脂粒於不同溫度下
之b2, B0及b2 / B0 值…………………………………….145

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指導教授

陳文逸(Wen-yih Chen)

審核日期

2000-8-18

推文