兩相性幾丁聚醣包覆磁性奈米粒子之製備與生醫上的應用

Abstract

長久以來，由於其特殊性質，磁性奈米粒子 (MNPs) 的合成受到科學和技術上的重視。在生物醫學領域MNPs已經廣泛地應用於組織影像、強化核磁共振的造影及有選擇性地和具功能性的脫氧核醣核酸混合。在本研究中，我們用過碘酸鈉 (NaIO4) 將幾丁聚醣環狀結構開環而得到醛基化幾丁聚醣。接著在醛基化幾丁聚醣上鍵結正十六胺，進而合成具有疏水和親水端的幾丁聚醣。改質後的兩性烷基化幾丁聚醣可以藉由形成微胞去包覆由有機相合成的MNPs使其從有機相轉至水相中穩定懸浮。根據X光繞射分析儀 (XRD) 和震動樣品磁強計 (VSM) 的測試結果，MNPs顯現出超順磁的特性。由穿透式電子顯微鏡 (TEM) 的影像，MNPs和被兩性烷基化幾丁聚醣包覆的磁性奈米粒子 (CMNPs) 大小分別為7奈米和25奈米。於基因轉殖實驗中，CMNPs經由帶正電之幾丁聚醣與綠色螢光蛋白表達載體 (pEGFP-N1) 形成複合體 (complex) 在磁場的有無條件下與大腸癌HT-29細胞株共同培養。藉由共軛焦顯微鏡的偵測和細胞存活率的測試，HT-29細胞有著超過90%的存活率且在磁場下展現比在無磁場下較多的綠色基因螢光表達。於光動力療法實驗鍵結上孟加拉玫瑰素 (rose bengal) 之CMNPs與HT-29細胞共同培養後，經由540奈米波長的發光二極體 (LED) 照射後會導致自由基 (singlet oxygen) 的產生進而殺死細胞。

Synthesis of magnetic nanoparticles (MNPs) has long been of scientific and technical interest due to their potential applications in tissue images, contrast enhancement to magnetic resonance imaging (MRI), and selective hybridization to nucleic acids. In this study, we employed NaIO4 to open the cyclic structure of chitosan and obtain periodate-oxidized chitosan that is further grafted with 1-hexadecylamine to form modified chitosan with amphiphilic feature. The alkylated chitosan could convert organically synthesized MNPs into aqueous phase by virtue of hydrophobic interaction. As shown in XRD and VSM analysis, MNPs exhibited superparamagnetic characteristics. TEM data showed that the average size of MNPs and the amphiphilic chitosan-capped MNPs (CMNPs) were 7 and 25 nm, respectively. In gene transfection study, CMNPs utilized positively charged chitosan to form complexes with plasmid pEGFP-N1 were co-cultured with colon HT-29 cancer cells with and without magnetic field. According to confocal microscopic images and cell viability assay, HT-29 cells could remain over 90% cell viability and express more green fluorescence proteins in the presence of magnetic field. In photodynamic therapy study, CMNPs conjugated with rose bengal (RB) were used to challenge HT-29 cells and then irradiated with 540 nm light emitting diode (LED) light. The singlet oxygen released from the illuminated cells were detected and accounted for the destruction of HT-29 cells.