摘要
论文研究了PEG/盐、浊点萃取、醇/盐和离子液体/盐四种双水相体系,并成功将其应用到萃取、分离和纯化芦荟中的蒽醌、多糖类物质。
首先,采用星点设计-响应面法分别优化了芦荟中的蒽醌和多糖类物质提取工艺。分别考察了乙醇浓度、提取温度和液固比对蒽醌得率的影响；提取温度、提取时间和液固比对多糖得率的影响。采用3因素5水平设计了实验,绘制出自变量三维效应曲面图。对结果进行预测分析,获得响应值最大的优化条件。
其次,采用PEG/盐双水相体系对芦荟蒽醌粗提液进行分离纯化。在筛选出最佳的PEG/盐体系后,考察了pH值、无机盐、温度对萃取的影响。在萃取过程中,芦荟蒽醌被萃取到PEG相,调节体系pH值来实现反萃取。
第三,采用基于非离子表面活性剂的浊点萃取体系分离纯化芦荟蒽醌类物质。考察了Triton X-114浓度、pH值、平衡温度和时间、添加剂对萃取效率的影响。蒽醌被萃取到表面活性剂相,通过反萃取能将蒽醌萃取到水相,同时表面活性剂能够得到回收利用。
第四,采用丙醇/硫酸铵体系萃取、纯化芦荟中的蒽醌类物质。考察了醇/盐的类型和浓度、温度、pH值对萃取的影响。在最佳条件下,大部分蒽醌被萃取到醇相,而大部分的杂质如多糖等被萃取到盐相。萃取完成后,丙醇通过旋蒸进行回收,盐相中的盐通过溶析结晶的方法进行回收。研究了蒽醌在体系中的分配机理,证明蒽醌在双水相体系的分配受疏水作用、盐析作用、氢键作用等作用力的影响。通过研究萃取过程的热力学,证明温度在很大程度上影响了蒽醌的分配。
第五,选择离子液体/盐双水相体系作为本实验中使用到的四种双水相体系的代表,研究了双水相体系的一些基本特征。首先合成了一系列碳链长度和阴离子不同的咪唑型离子液体,并合成了具有特定功能的功能化离子液体。采用红外和核磁等手段对合成出来的离子液体进行表征。然后将合成出来的离子液体与常见的无机盐构建双水相体系,研究了体系的相平衡,绘制出相图、系线等。
第六,将离子液体/盐双水相体系应用于同时萃取、分离芦荟多糖和蛋白质。多糖被萃取到盐相,然而大部分杂质如蛋白质等被萃取到离子液体相。通过研究多糖和蛋白质在双水相体系中分配行为来获得最佳的萃取条件,考察了盐类型和浓度、温度、pH值、添加剂等对萃取的影响。盐相中的多糖用透析法进一步纯化。1-丁基-3-甲基咪唑四氟硼酸盐([C4mim]BF4)通过溶剂萃取进行回收。
最后,[C4mim]BF4/Na2SO4双水相体系被应用于分离、纯化芦荟中的蒽醌类物质。考察了影响萃取的因素如离子液体的类型、盐的类型和浓度、萃取温度、pH值等。在最佳的萃取条件下,该方法通过一步萃取能将蒽醌萃取到离子液体相。萃取后,通过反萃取的方法来回收离子液体,能简单地将离子液体和蒽醌进行分离。采用高效液相色谱法分析芦荟蒽醌的主要成分芦荟大黄素和大黄酚在萃取前后浓度变化。
在萃取芦荟活性成分的应用时比较了四种双水相体系的优势和劣势,为其在分离、纯化植物活性成分应用研究奠定了理论和实践基础。
In this paper, we have studied four aqueous two-phase systems (ATPSs) of PEG/salt, cloud point extraction (CPE), alcohol/salt and ionic liquid (IL)/salt. Meanwhile, the four systems were used in extraction, separation and purification of aloe anthraquinones, polysaccharides.
Firstly, we have optimized the extraction process of aloe anthraquinones and polysaccharides by the central composite design and response surface methodology (CCD-RSM). The influencing factors to anthraquinones yield of concentration of ethanol, extraction time and liquid solid ratio were investigated. The influencing factors to polysaccharides yield of extraction temperature, extraction time and liquid solid ratio were investigated. The experiments were designed by3factors and5levels, the3D response surface plots were drawn. The optimal conditions were obtained by analysis and pridiction of the experimental results.
Secondly, the PEG/salt ATPS was used in separation and purification of aloe anthraquinones. The factors of pH, inorganic addictive and temperature were investigated by the optimal PEG/salt system. The anthraquinones were extracted into PEG-rich phase after ATPS extraction and then the back extraction was achieved by adjusting pH.
Thirdly, the CPE based on a non-ionic surfactant Triton X-114was used to separate and purify aloe anthraquinones. The influencing parameters of Triton X-114concentration, pH, equilibrium temperature and time and the addictive of NaCl were investigated. Aloe anthraquinones were extracted into aqueous phase by back extraction, and the surfactant can be recycled. The analytical characteristics and the validity of this method were analyzed.
Fourthly, the alcohol/salt ATPS was used in extraction and purification of aloe anthraquinones. The factors of the type and concentration of alcohol and salt, temperature and pH were investigated. Under the optimal condition, majority of anthraquinones were extracted into alcohol-rich phase and the impurities of polysaccharides, proteins and mineral substance were extracted into salt-rich phase. The alcohol in alcohol-rich phase can be recycled by evaporation and the salt in salt-rich phase can be recycle by dilution crystallization. The major constitutes of aloe anthraquinones were aloe-emodin by HPLC analysis. The distribution of anthraquinones in alcohol/salt ATPS were influenced by hydrophobicity, salting-out effect and hydrogen bond interaction through investigating the extraction mechanism. The thermodynamics also demonstrated that temperature can affect the distribution to a great extent.
Fifthly, a series of iminazole IL intermediate with different length of carbon chain were synthetized. Then ILs with different anion were synthetized by ion-change method. Furthermore, the functionalized ILs were synthetized. The ILs were charactered by IR and HMNR. The ATPSs were constructed using IL and salt, the phase equilibrium was studied and the phase diagram and tie line were drawn.
Sixthly, the IL/salt ATPS was used in simultaneous extraction and purification of aloe polysaccharides and proteins. Polysaccharides were extracted into salt-rich phase, while majority impurities of proteins, mineral substance and phenol compounds were extracted into IL-rich phase. The optimal condition was obtained by studying the distribution of polysaccharides and proteins in ATPS, the factors of salt type and concentration, temperature, pH, inorganic addictive were investigated. Polysaccharides after ATPS extraction were further purified by dialysis membrane, IL and salt were filtrated leaving polysaccharides in the dialysis tube. The TG analysis demonstrated this method can obtain polysaccharides with higher purity.[C4mim]BF4in IL-rich phase was extracted into CH2CI2,[C4mim]BF4can be recycled by evaporating CH2Cl2.
Lastly, the IL/salt ATPS was used in separation and purification of anthraquinones.[C4mim]BF4was chosen as the phase-forming IL due to its good phase-forming ability and extraction ability. The influencing system parameters of type of IL, type and concentration of salt, extraction temperature and pH were investigated. Under the optimal condition, anthraquinones can be extracted into IL-rich phase by a single-step extraction and majority impurities were extracted into salt-rich phase. The major components of aloe-emodin and chrysophanol were analyzed by HPLC before and after ILATPS. Compared with conventional liquid-liquid extraction, this ILATPS is more efficient and environment friendly.
The advantages and disadvantages of the four types of ATPSs were compared in this paper, which pvovide a theoretical direction in the separation field.
引文
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