嵌入糖醛酸印迹聚合物的 cryogel 对透明质酸的分离与纯化。
Separation and purification of hyaluronic acid by embedded glucuronic acid imprinted polymers into cryogel.
机构信息
Department of Chemistry, Anadolu University, Eskişehir, Turkey.
出版信息
J Chromatogr B Analyt Technol Biomed Life Sci. 2013 Sep 1;934:46-52. doi: 10.1016/j.jchromb.2013.06.022. Epub 2013 Jul 9.
Hyaluronic acid (HA) has been used in many applications such as pharmaceutical, clinical and cosmetics, so its separation and purification is very important. In this study, firstly d-glucuronic acid imprinted polymers (MIPs) have been synthesized for the separation of HA which has glucuronic acid part in its structure. MIP particles have characterized by elemental analysis, Fourier Transform Infrared Spectroscopy (FT-IR) and swelling tests. Then, synthesized MIP particles have embedded into polyacrylamide based cryogel. Cryogel has prepared by free radical cryogelation process initiated by N,N,N',N'-Tetramethylethylenediamine (TEMED) and ammonium persulfate (APS) as redox initiators. This cryogel material was characterized by FT-IR, swelling tests, scanning electron microscopy (SEM) and surface adsorption analyze including pore size analyzer (BET) method. The adsorption of HA has investigated by spectrophotometric method using MIPs embedded into cryogel columns (GAIPEC) and the maximum HA adsorption capacity was found to be 318mgg(-1). The selectivity of GAIPEC column has estimated using N-acetylglucose amine as interfering agent since this molecule is a part of HA and the results have shown that GAIPEC has been nearly 35 times selective for HA than N-acetylglucose amine. The optimum chromatographic conditions for separation of HA were investigated. pH 7.0 buffer solution for elution and 0.1M of NaCl solution as desorption agent were used at 0.5mLmin(-1) flow rate. Also, recovery of GAIPEC was investigated and the results have shown that GAIPEC could be used many times without decreasing its adsorption capacity significantly. Here in, combining selectivity of MIP particles and mechanical properties of cryogel, a rigid and stable material was prepared for the separation and purification of HA. To point out this, HA has been isolated from fish eye and fermentation of Streptococcus equi RSKK 679 cell culture. After that, it has characterized and Fast Protein Liquid Chromatography (FPLC) applications have been investigated.
透明质酸（HA）已被广泛应用于医药、临床和化妆品等领域，因此其分离和纯化非常重要。在本研究中，首先合成了具有葡萄糖醛酸部分结构的 d-葡萄糖醛酸印迹聚合物（MIPs），用于分离 HA。MIP 颗粒通过元素分析、傅里叶变换红外光谱（FT-IR）和溶胀试验进行了表征。然后，将合成的 MIP 颗粒嵌入到聚丙烯酰胺基 cryogel 中。cryogel 通过自由基 cryogelation 过程制备，由 N，N，N'，N'-四甲基乙二胺（TEMED）和过硫酸铵（APS）作为氧化还原引发剂引发。这种 cryogel 材料通过 FT-IR、溶胀试验、扫描电子显微镜（SEM）和表面吸附分析（包括孔径分析仪（BET）方法）进行了表征。通过使用嵌入 cryogel 柱（GAIPEC）的 MIPs 的分光光度法研究了 HA 的吸附，发现最大 HA 吸附容量为 318mgg(-1)。使用 N-乙酰葡萄糖胺作为干扰剂估计了 GAIPEC 柱的选择性，因为该分子是 HA 的一部分，结果表明 GAIPEC 对 HA 的选择性几乎是 N-乙酰葡萄糖胺的 35 倍。还研究了分离 HA 的最佳色谱条件。使用 pH 7.0 缓冲溶液作为洗脱液，0.1M 的 NaCl 溶液作为洗脱剂，流速为 0.5mLmin(-1)。此外，还研究了 GAIPEC 的回收率，结果表明 GAIPEC 可以多次使用而不会明显降低其吸附能力。在这里，通过结合 MIP 颗粒的选择性和 cryogel 的机械性能，制备了一种刚性稳定的材料，用于 HA 的分离和纯化。为了指出这一点，从鱼眼和马链球菌 RSKK 679 细胞培养物的发酵液中分离出了 HA，然后对其进行了表征，并研究了快速蛋白液相色谱（FPLC）的应用。
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