本文選題：自熒光 + 介孔二氧化硅�。� 參考：《蘭州大學(xué)》2016年博士論文

【摘要】：近年來(lái),藥物載體作為降低藥物毒副作用、改善藥物體內(nèi)分布與代謝、提高藥物治療效率的一種手段,已經(jīng)成為生物醫(yī)學(xué)領(lǐng)域的研究熱點(diǎn)之一。其中,介孔二氧化硅納米粒子由于其具有有序的介孔結(jié)構(gòu)、比表面積大、生物相容性好、藥物負(fù)載率高等優(yōu)點(diǎn),引起了人們的廣泛關(guān)注,且被認(rèn)為是最有前景的藥物載體之一。然而,該納米粒子在實(shí)際應(yīng)用中尚存在一些亟待解決的問(wèn)題,如在水中的分散性差、易聚集;功能相對(duì)單一,現(xiàn)有的修飾方法存在潛在毒性;且各種功能分子之間的分配存在競(jìng)爭(zhēng)性矛盾;實(shí)現(xiàn)不同藥物之間的協(xié)同作用的方法可控性低等。本論文針對(duì)上述問(wèn)題展開(kāi)了自熒光介孔二氧化硅納米粒子的制備、表征及其作為藥物載體的應(yīng)用等研究。本論文制備了一系列自熒光介孔二氧化硅納米粒子,采用傅立葉變換紅外光譜(FTIR)和熱重分析(TGA)對(duì)介孔二氧化硅納米粒子的修飾過(guò)程進(jìn)行了監(jiān)測(cè),采用透射電子顯微鏡(TEM)對(duì)介孔二氧化硅納米粒子修飾前后的形貌變化及粒徑變化進(jìn)行了觀察,通過(guò)N2吸附-脫附對(duì)介孔二氧化硅納米粒子修飾前后的孔徑、孔容及比表面積進(jìn)行了測(cè)定,采用熒光分光光度計(jì)、熒光顯微鏡及共聚焦熒光顯微鏡對(duì)介孔二氧化硅納米粒子的熒光性質(zhì)及熒光成像能力進(jìn)行了評(píng)價(jià),采用四甲基偶氮唑鹽微量酶反應(yīng)比色法(MTT法)初步考察了介孔二氧化硅納米粒子的體外細(xì)胞毒性,通過(guò)體外釋放實(shí)驗(yàn)考察了介孔二氧化硅納米粒子作為藥物載體在刺激響應(yīng)性藥物控制釋放行為方面的可行性,具體研究?jī)?nèi)容及結(jié)果如下:(1)采用PAMAM樹(shù)枝體硅烷偶聯(lián)劑修飾介孔二氧化硅納米粒子,制備了具有pH敏感性和自熒光的藥物載體,研究結(jié)果發(fā)現(xiàn)經(jīng)PAMAM樹(shù)枝體硅烷偶聯(lián)劑修飾后,介孔二氧化硅納米粒子表面含有大量的伯胺基團(tuán),有利于進(jìn)一步進(jìn)行修飾,引入其他官能團(tuán)分子,如靶向分子;同時(shí),經(jīng)PAMAM樹(shù)枝體硅烷偶聯(lián)劑修飾后,介孔二氧硅納米粒子的載藥量得到了進(jìn)一步提高。此外,研究還發(fā)現(xiàn),通過(guò)改變?nèi)芤旱膒H值,可以改變PAMAM樹(shù)枝狀分子的構(gòu)象,進(jìn)而改變其在粒子外層的密度,實(shí)現(xiàn)藥物的控制釋放,并可以防止藥物在中性條件下的突釋;熒光發(fā)射強(qiáng)度和熒光顏色可隨著藥物負(fù)載率和藥物分子的釋放而發(fā)生變化,從而具有在跟蹤或檢測(cè)方面應(yīng)用的潛力。研究結(jié)果表明,PAMAM樹(shù)枝體可同時(shí)作為ph敏感的封堵劑和熒光標(biāo)記物。(2)基于硫酸軟骨素(cs)和pamam樹(shù)枝體采用層層自組裝的方法制備了多功能介孔二氧化硅納米粒子藥物載體。研究發(fā)現(xiàn),在中性條件下,pamam/cs層層自組裝膜可以有效地將藥物進(jìn)行封堵,抑制了介孔中藥物的提前釋放;該藥物載體具有刺激響應(yīng)性藥物控制釋放性質(zhì),即模型藥物阿霉素和姜黃素在中性ph值條件下釋放緩慢,在酸性(ph5.5)條件下釋放較快;考察了pamam/cs層層自組裝膜對(duì)粒子分散性質(zhì)的影響,結(jié)果表明,層層自組裝膜能使介孔二氧化硅納米載體具有更好的分散性;通過(guò)溶血實(shí)驗(yàn)發(fā)現(xiàn)層層自組裝膜可以有效屏蔽硅羥基與血紅細(xì)胞膜的相互作用,從而提高該納米粒子的血液相容性;熒光光譜和熒光顯微鏡結(jié)果初步表明該納米粒子具有熒光成像的能力;cs的靶向功能可以促進(jìn)該納米粒子藥物載體進(jìn)入腫瘤細(xì)胞,增強(qiáng)其細(xì)胞毒性。(3)采用負(fù)載了熒光抗癌藥物姜黃素的f127膠束(cf127)作為介孔二氧化硅納米粒子的封堵劑和熒光標(biāo)記物,制備了多功能介孔二氧化硅納米粒子藥物載體。研究結(jié)果表明,cf127可以作為介孔二氧化硅納米粒子的封堵劑;兩種藥物載體(介孔二氧化硅納米粒子和膠束)的組合提供了一種普適性的設(shè)計(jì)制備多功能藥物載體的方法,可同時(shí)實(shí)現(xiàn)刺激性控制藥物釋放和熒光成像功能;體外藥物釋放實(shí)驗(yàn)結(jié)果表明,cf127膠束通過(guò)席夫堿鍵鍵接在介孔二氧化硅納米粒子表面后,可以實(shí)現(xiàn)藥物的ph響應(yīng)性控制釋放,這可以歸因于席夫堿對(duì)ph的敏感性,即在中性條件下,席夫堿鍵可以較穩(wěn)定存在,因此可以阻止藥物釋放,而在ph值為5.5時(shí),席夫堿鍵斷裂,釋放藥物;熒光實(shí)驗(yàn)結(jié)果發(fā)現(xiàn),cf127膠束聚集在介孔二氧化硅納米顆粒表面后,cf127膠束的熒光強(qiáng)度會(huì)明顯增強(qiáng),該現(xiàn)象有助于熒光成像;此外,通過(guò)考察該載體在連續(xù)紫外激發(fā)光照射下的熒光強(qiáng)度變化,發(fā)現(xiàn)cf127膠束具有良好的抗光漂白性,這些結(jié)果顯示cf127膠束可作為介孔二氧化硅納米粒子藥物載體的熒光標(biāo)記物。(4)采用抗癌藥物姜黃素分子作為介孔二氧化硅納米粒子的封堵劑和熒光成像劑,制備了具有自熒光功能和刺激響應(yīng)性的藥物載體(lpcc-c-f127)。研究結(jié)果表明,姜黃素封堵劑可以有效地把藥物分子保留在介孔里,避免藥物分子進(jìn)入細(xì)胞前的提前泄露;體外藥物釋放實(shí)驗(yàn)結(jié)果表明,在ph5.5和谷胱甘肽(gsh)存在下,姜黃素上的β-硫酯可以被有效水解,從而使姜黃素分子從介孔上脫落,實(shí)現(xiàn)藥物的刺激響應(yīng)性控制釋放;采用熒光分光光度計(jì)分析后發(fā)現(xiàn),通過(guò)多步功能化修飾后,納米藥物載體的熒光強(qiáng)度隨著姜黃素封堵劑上姜黃素分子的增加而增強(qiáng);分散性實(shí)驗(yàn)結(jié)果表明,f127膜層可以通過(guò)聚乙二醇(peg)殼層提高該藥物載體的分散性,同時(shí)還可創(chuàng)建一個(gè)疏水的微環(huán)境提高該藥物載體的熒光強(qiáng)度;mtt法實(shí)驗(yàn)結(jié)果表明,所制備的藥物載體具有良好的細(xì)胞生物相容性。(5)基于姜黃素分子采用一鍋法制備了姜黃素聚合物殼封堵的多功能介孔二氧化硅納米粒子藥物載體。體外藥物釋放實(shí)驗(yàn)結(jié)果表明,姜黃素聚合物殼可以有效地將藥物分子封堵在介孔里,并能實(shí)現(xiàn)介孔的開(kāi)關(guān),實(shí)現(xiàn)介孔內(nèi)藥物分子的控制釋放;降解實(shí)驗(yàn)結(jié)果表明,姜黃素聚合物殼可以被谷胱甘肽(GSH)有效降解,并且其降解速度與GSH的濃度成正比;通過(guò)熒光分光光度計(jì)和熒光顯微鏡測(cè)定發(fā)現(xiàn)姜黃素聚合物殼可以使該納米粒子藥物載體具有熒光功能;分散性實(shí)驗(yàn)結(jié)果發(fā)現(xiàn),所得姜黃素聚合物殼可以改善介孔二氧化硅藥物載體在磷酸緩沖液中的分散性;通過(guò)體外細(xì)胞實(shí)驗(yàn)結(jié)果證實(shí)該藥物載體能夠有效殺死A549癌細(xì)胞。
[Abstract]:In recent years, as a means to reduce the side effects of drug toxicity, improve the distribution and metabolism of drugs and improve the efficiency of drug treatment, it has become one of the hot spots in the field of biomedicine. It has attracted wide attention and is considered to be one of the most promising drug carriers. However, there are still some problems to be solved in its practical application, such as poor dispersivity and easy aggregation in water, relatively single function, potential toxicity of existing modification methods, and various functional molecules. In this thesis, a series of self fluorescent mesoporous silica nanoparticles have been prepared. The modification process of mesoporous silica nanoparticles was monitored by Fu Liye transform infrared spectroscopy (FTIR) and thermogravimetric analysis (TGA). The changes of morphology and particle size of mesoporous silica nanoparticles were observed by transmission electron microscopy (TEM), and the mesoporous silica nanoparticles were adsorbed by N2 adsorption and desorption. The pore size, Kong Rong and specific surface area were measured before and after the subsurface modification. Fluorescence spectrophotometer, fluorescence microscope and confocal fluorescence microscope were used to evaluate the fluorescence properties and fluorescence imaging ability of mesoporous silica nanoparticles. The mesopore was preliminarily investigated by four methyl azazolase Microenzyme reaction Colorimetry (MTT method). The cytotoxicity of silica nanoparticles in vitro was investigated in vitro. The feasibility of mesoporous silica nanoparticles as drug carriers in stimulating response to drug control release behavior was investigated. The specific content and results were as follows: (1) modified mesoporous silica nanoparticles with PAMAM dendrane coupling agent, The drug carriers with pH sensitivity and self fluorescence were prepared. The results showed that the surface of mesoporous silica nanoparticles had a large number of primary amine groups modified by PAMAM dendritic silane coupling agent, which was beneficial to the further modification and the introduction of other functional groups, such as target molecules, and modified by the PAMAM dendranes silane coupling agent. After that, the drug loading of mesoporous two oxygen silicon nanoparticles has been further improved. In addition, it is found that by changing the pH value of the solution, the conformation of PAMAM dendrites can be changed and the density of the particles in the outer layer of the particles can be changed to control the release of drugs, and the release of the drugs under neutral conditions can be prevented; the intensity of fluorescence emission and the intensity of fluorescence emission can be prevented. The fluorescence color changes with the drug loading rate and the release of drug molecules, and thus has the potential to be used for tracking or detection. The results show that PAMAM dendrites can be used as pH sensitive plugging agent and fluorescent marker. (2) a layer self-assembly method based on chondroitin sulfate (CS) and PAMAM branches is prepared. A multifunction mesoporous silica nanoparticle drug carrier. It is found that, under neutral conditions, pamam/cs layer self assembled monolayers can effectively block drugs and inhibit the early release of drugs in the mesoporous. The drug carrier has the properties of stimulating response drug control release, that is, the model drug adriamycin and curcumin in neutral pH value. The release was slow under the condition of acid (pH5.5) condition. The effect of pamam/cs layer self assembled film on the dispersion properties of particles was investigated. The results showed that the layered self assembled monolayer could make the mesoporous silica nanoscale have better dispersion, and the hydroxyl group and blood red fine could be effectively shielded by the hemolysis experiment. The interaction of the cell membrane to improve the blood compatibility of the nanoparticles; the fluorescence and fluorescence microscopy results showed that the nanoparticle had the ability of fluorescence imaging; the targeting function of CS could promote the nanoparticle drug carrier to enter the tumor cells and enhance its cytotoxicity. (3) a fluorescent anti-cancer drug, Jiang Huang, was used. F127 micelle (cf127) is used as the plugging agent and fluorescent marker of mesoporous silica nanoparticles, and the drug carrier of multifunction mesoporous silica nanoparticles is prepared. The results show that cf127 can be used as a plugging agent for mesoporous silica nanoparticles, and the combination of two kinds of drug carriers (mesoporous silica nanoparticles and micelles) A universal method for the preparation of multifunctional drug carriers is provided, which can simultaneously achieve stimulant control of drug release and fluorescence imaging. In vitro drug release experiment results show that cf127 micelles can be controlled by the pH response of the mesoporous silica nanoparticles after the Schiff base bond is connected to the mesoporous silica nanoparticles. It can be attributed to the sensitivity of Schiff base to pH, that is, under the neutral condition, the Schiff base bond can be more stable and thus can prevent the release of the drug. When the pH value is 5.5, the Schiff base bond breaks and releases the drug. The fluorescence test results show that the fluorescence intensity of the cf127 micelles will be evident after the cf127 micelles are aggregated on the mesoporous two oxygen silicon nanoparticles. In addition, the cf127 micelles have good anti photobleaching properties by examining the changes in the fluorescence intensity of the carrier in continuous UV irradiation. These results show that the cf127 micelle can be used as a fluorescent marker for the carrier of the mesoporous silica nanoparticles. (4) the anti-cancer drug Jiang Huang is used. As the plugging agent and fluorescent imaging agent of mesoporous silica nanoparticles, a drug carrier (lpcc-c-f127) with self fluorescence function and stimulus responsiveness is prepared. The results show that the curcumin plugging agent can effectively keep the drug molecules in the mesoporous and avoid the advance leakage of drug molecules before the cells; in vitro drugs. The results of the release experiment showed that the beta thioester on Curcumin could be hydrolyzed effectively in the presence of pH5.5 and glutathione (GSH), so that the curcumin molecules fell off the mesoporous and controlled the response of the drug to be controlled and released. The fluorescence spectrophotometer was used to detect the fluorescence of the nano drug carrier after multi-step functionalization. The strength was enhanced with the increase of curcumin molecules on the curcumin plugging agent. The results of dispersion experiment showed that the F127 film could improve the dispersibility of the drug carrier by the polyethylene glycol (PEG) shell, and also create a hydrophobic microenvironment to improve the fluorescence intensity of the drug carrier. The results of the MTT method showed that the prepared drug was loaded. The body has good biocompatibility. (5) the drug carrier of multifunction mesoporous silica nanoparticles is prepared by one pot method based on the curcumin molecule. The results of drug release in vitro show that the curcumin polymer shell can effectively block the molecule in the mesoporous and realize the mesoporous. The results showed that the curcumin polymer shell could be effectively degraded by glutathione (GSH), and the degradation rate was proportional to the concentration of GSH. The fluorescence spectrophotometer and fluorescence microscope showed that the gingiber polymer shell could make the nanoparticle drug carrier. The results showed that the curcumin polymer shell could improve the dispersibility of the mesoporous silica drug carrier in the phosphoric acid buffer solution, and the results of cell experiment in vitro proved that the drug carrier could effectively kill the A549 cancer cells.
【學(xué)位授予單位】：蘭州大學(xué)
【學(xué)位級(jí)別】：博士
【學(xué)位授予年份】：2016
【分類號(hào)】：TQ127.2;TB383.1
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本文編號(hào)：1961494