發(fā)布時間：2018-01-27 12:22

  本文關(guān)鍵詞： 大豆淀粉 淀粉微球 微載體 細(xì)胞培養(yǎng)　出處：《西北民族大學(xué)》2017年碩士論文　論文類型：學(xué)位論文

【摘要】：淀粉微球在藥物緩釋、包埋劑和吸附劑方面的應(yīng)用已有很多報道,但在貼壁動物細(xì)胞培養(yǎng)方面的應(yīng)用卻鮮有研究報道。用淀粉制備細(xì)胞培養(yǎng)用微載體,具有其獨(dú)特的優(yōu)勢。本研究以價廉易得的淀粉為原料,研究制備適合動物細(xì)胞貼壁培養(yǎng)用的微載體,旨在開發(fā)具有高附加值、可生物降解、可用淀粉酶消化以收獲細(xì)胞、非動物來源材料制備的淀粉微載體,避免在生物制藥領(lǐng)域使用動物源性材料和高成本微載體的弊端。本文探討了微載體的研究進(jìn)展、高分子微球的基本性質(zhì)、淀粉微球的制備方法及其研究應(yīng)用進(jìn)展。論證了淀粉微球作為動物細(xì)胞培養(yǎng)用微載體的可行性。在前人制備淀粉微球的基礎(chǔ)上,經(jīng)過制備工藝改進(jìn)和參數(shù)優(yōu)化,探討了制備淀粉微球的最佳工藝條件。結(jié)果表明:以機(jī)油和異辛烷為油相,6%糊化淀粉堿溶液為水相,聚乙二醇二縮水甘油醚為交聯(lián)劑制備交聯(lián)淀粉微球時,油水相體積比4:1,攪拌轉(zhuǎn)速500 rpm,交聯(lián)劑10%為制備交聯(lián)淀粉微球的最佳工藝條件,制備的交聯(lián)淀粉微球能夠耐受121℃C 30min的高壓滅菌。通過正交試驗(yàn)、配基篩選,在淀粉微球表面偶聯(lián)了二乙氨基乙基(DEAE),使淀粉微球表面具有了和商品化Cytodex 1微載體近似的電荷和電荷量。在測定、分析了所制交聯(lián)淀粉微球的基本理化性質(zhì)后,本研究以BHK-21、Marc-145和MDCK三種疫苗生產(chǎn)常用貼壁細(xì)胞為實(shí)驗(yàn)細(xì)胞,以Cytodex 1微載體為對照,進(jìn)行的微載體細(xì)胞培養(yǎng),并對實(shí)驗(yàn)結(jié)果進(jìn)行了總結(jié)分析。研究結(jié)果表明:所制備的淀粉微球,在偶聯(lián)DEAE后,能夠耐受高壓滅菌。在細(xì)胞培養(yǎng)后,偶聯(lián)了 DEAE、粒徑與Cytodex 1微載體相近的淀粉微球,上述三種細(xì)胞均能夠良好貼附、增殖。在相同懸浮培養(yǎng)條件下,淀粉微載體的細(xì)胞貼附、增殖量與Cytodex 1微載體近似,培養(yǎng)至144h,上述三種細(xì)胞的增殖量均達(dá)到最大值。其中,在自制淀粉微球上BHK-21細(xì)胞的最大增殖量為 2.5 ×106cells/mL,略大于 Cytodex 1 的 2.2 ×106cells/mL。Marc-145 細(xì)胞的最大增殖量為前者1.4×106cells/mL,后者1.6×106cells/mL,后者略大于前者;MDCK細(xì)胞的最大增殖量為前者2.2×106cells/mL,后者2.4×106cells/mL,后者略大于前者。用自制微載體培養(yǎng)細(xì)胞,三種貼壁細(xì)胞的生長與生化代謝曲線也和商品化的Cytodex 1相似。綜上可知,本研究制備的淀粉微球,通過偶聯(lián)DEAE后,能夠用于BHK-21、Marc-145和MDCK貼壁細(xì)胞的微載體懸浮培養(yǎng)。其細(xì)胞培養(yǎng)效果與Cytodex 1相似,有良好的開發(fā)與應(yīng)用前景。
[Abstract]:The application of starch microspheres in drug delivery, encapsulation and adsorbents has been reported, but few studies have been reported on the application of starch microspheres in cell culture of adherent animals. This study is based on the cheap and easily available starch as raw material to study the preparation of microcarriers suitable for animal cell adhesion culture in order to develop a high value-added biodegradable microcarriers. Starch microcarriers that can be digested by amylase to harvest cells and non-animal materials. To avoid the disadvantages of using animal-derived materials and high-cost microcarriers in the field of biopharmaceuticals, this paper discusses the research progress of microcarriers and the basic properties of polymer microspheres. The preparation and application of starch microspheres. The feasibility of starch microspheres as microcarriers for animal cell culture was demonstrated. On the basis of the previous preparation of starch microspheres, the preparation process was improved and the parameters were optimized. The optimum conditions for preparation of starch microspheres were discussed. The results showed that when oil and isooctane were used as oil phase and 6% gelatinized starch alkali solution as water phase, polyethylene glycol diglycidyl ether was used as crosslinking agent to prepare cross-linked starch microspheres. The optimum conditions for the preparation of cross-linked starch microspheres were oil / water phase volume ratio 4: 1, stirring speed 500rpm, crosslinking agent 10%. The cross-linked starch microspheres were able to withstand high pressure sterilization at 121 鈩，

本文編號：1468463