离子液体双水相萃取分离苋菜红的研究

建立了由亲水性离子液体四氟硼酸1-丁基-3-甲基咪唑([Bmim]BF4)和NaH2PO4形成的双水相体系萃取分离苋菜红的新方法.研究了盐的浓度、离子液体浓度、溶液酸度、其它共存物质对苋菜红萃取率的影响.结果表明,NaH2PO4加入量在2～2.5 g,离子液量在1.0～2.0 mL,苋菜红溶液量在1.5 mL,溶液酸度在pH 4～6范围,离子液体双水相体系对苋菜红有较高的萃取率(E%＞90).用加入无机离子、不同类型表面活性剂和吸收光谱探讨了离子液体与苋菜红之间的作用.     

The influence of temperature on the phase behavior of ionic liquid aqueous two-phase systems

In this article, the equilibrium behavior of solutions of 1-ethyl-3-methylimidazolium dimethyl phosphate ([Emim]DMP) and ethyl acetate or acetone in aqueous two-phase system (ATPS) was discussed to understand the liquid–liquid equilibrium (LLE) behavior of these organic solvents. Thus, we determined phase diagrams and LLE data at 303.15, 308.15, 313.15, and 323.15 K for the investigated biphasic systems. Four empirical equations were used to study the tie lines. The results showed that for the [Emim]DMP?+?acetone?+?water biphasic systems within the investigated temperature range, temperature influences the phase behavior, but for the [Emim]DMP?+?ethyl acetate?+?water biphasic systems within the investigated temperature range, there are no notable changes on the phase behavior with rising temperature. The results may have important applications for the separation of antibiotics and for the recovery of ionic liquids (ILs).     

离子液体双水相萃取山楂黄酮和多糖的相行为研究

用分光度法研究了[Bmim]BF4/(NH4)2SO4双水相体系萃取山楂黄酮和多糖的相行为,考察了同时提取这两种组分时[Bmim]BF4的浓度、(NH4)2SO4的浓度、山楂的用量、超声萃取时间等因素对双水相的上下相体积以及分配系数的影响。结果表明:(1)离子液体浓度的增加,双水相的上、下相体积分别明显增大和减小。但黄酮和多糖在双水相中的分配系数仅有波动,均小于5%;(2)硫酸铵浓度的增加,双水相的上下相体积分别明显减小和增大。同时黄酮和多糖的分配系数均有较明显先降低后升高的趋势;(3)山楂质量的增减,不影响双水相的形成,但存在一个实验条件下的溶出饱和值,该值为0.15g,此时黄酮和多糖的分配系数最大;(4)超声萃取时间的延长或缩短,同样不影响双水相的形成,但存在对山楂有效成分达到饱和溶出的阈值,对于黄酮为25min,多糖为20min。     

Design of functional guanidinium ionic liquid aqueous two-phase systems for the efficient purification of protein

A series of novel cationic functional hexaalkylguanidinium ionic liquids and anionic functional tetraalkylguanidinium ionic liquids have been devised and synthesized based on 1,1,3,3-tetramethylguanidine. The structures of the ionic liquids (ILs) were confirmed by ¹H nuclear magnetic resonance (¹H NMR) and 13C nuclear magnetic resonance (13C NMR) and the production yields were all above 90%. Functional guanidinium ionic liquid aqueous two-phase systems (FGIL-ATPSs) have been first designed with these functional guanidinium ILs and phosphate solution for the purification of protein. After phase separation, proteins had transferred into the IL-rich phase and the concentrations of proteins were determined by measuring the absorbance at 278 nm using an ultra violet visible (UV–vis) spectrophotometer. The advantages of FGIL-ATPSs were compared with ordinary ionic liquid aqueous two-phase systems (IL-ATPSs). The proposed FGIL-ATPS has been applied to purify lysozyme, trypsin, ovalbumin and bovine serum albumin. Single factor experiments were used to research the effects of the process, such as the amount of ionic liquid (IL), the concentration of salt solution, temperature and the amount of protein. The purification efficiency reaches to 97.05%. The secondary structure of protein during the experimental process was observed upon investigation using UV–vis spectrophotometer, Fourier-transform infrared spectroscopy (FT-IR) and circular dichroism spectrum (CD spectrum). The precision, stability and repeatability of the process were investigated. The mechanisms of purification were researched by dynamic light scattering (DLS), determination of the conductivity and transmission electron microscopy (TEM). It was suggested that aggregation and embrace phenomenon play a significant role in the purification of proteins. All the results show that FGIL-ATPSs have huge potential to offer new possibility in the purification of proteins.     

Extraction of testosterone and epitestosterone in human urine using aqueous two-phase systems of ionic liquid and salt

Based on aqueous two-phase systems (ATPS) consisting of 1-butyl-3-methylimidazolium chloride, a hydrophilic ionic liquid (IL), and K2HPO4, a new and simple extraction technique, coupled with a reversed-phase high performance liquid chromatography (RP-HPLC), was developed for the simultaneous concentration and analysis of testosterone (T) and epitestosterone (ET) in human urine. Under the optimal conditions, the extraction efficiencies for both analytes were 80-90% in a one-step extraction. The method required only 3.0 mL of urine and a single hydrolysis/deproteinization/extraction step followed by direct injection of the IL-rich upper phase into HPLC system for analysis. The method has been satisfactorily applied to the analysis of T and ET in human urine with detection limits of 1 ng/mL and linear ranges of 10-500 ng/mL for both compounds. Compared with conventional liquid-liquid extraction or solid phase extraction, this new method is much "greener" due to no use of volatile organic solvent and low consumption of IL. The proposed extraction technique opens up new possibilities in the separation of other drugs.     

利用双水相分离回收离子液体的研究进展

研究离子液体的分离与回收对于减少离子液体对环境的影响、提高离子液体的利用效率、降低离子液体的应用成本、促进离子液体的工业应用具有重要的意义.本文重点综述了利用无机盐-离子液体双水相、糖-离子液体双水相、聚合物-离子液体双水相和CO2诱导的离子液体双水相技术分离回收离子液体的研究进展,分析了影响离子液体分离回收的关键因素,评价了不同离子液体双水相体系的优缺点,展望了该领域的发展方向及面临的挑战.     

Solvatochromic relationship: Prediction of distribution of ionic solutes in aqueous two-phase systems

Partition ratios of several ionic compounds in 20 different polymer/polymer aqueous two-phase systems (ATPS) containing 0.15 M NaCl in 0.01 M phosphate buffer, pH 7.4, were determined. The differences between the electrostatic properties of the phases in all the ATPS were estimated from partitioning of the homologous series of dinitrophenylated-amino acids. Also the solvatochromic solvent parameters characterizing the solvent dipolarity/polarizability (π*), solvent hydrogen-bond donor acidity (α), and solvent hydrogen-bond acceptor basicity (β) of aqueous media were measured in the coexisting phases of the ATPS. The solute-specific coefficients for the compounds examined were determined by the multiple linear regression analysis using the modified linear solvation energy relationship equation. The minimal number of ATPS necessary for determination of the coefficients was established and 10 ATPS were selected as a reference ATPS set. The solute-specific coefficients values obtained with this reference set of ATPS were used to predict the partition ratios for the compounds in 10 ATPS not included in the reference set. The predicted partition ratios values were compared to those determined experimentally and found to be in good agreement. It is concluded that the presented model of solute–solvent interactions as the driving force for solute partitioning in polymer/polymer ATPS describes experimental observations with 90–95% accuracy.     

Distribution behavior of some drug enantiomers in an aqueous two-phase system using counter-current extraction with protein

Summary The partial resolution of some drug enantiomers by counter-current extraction in an aqueous two-phase system using bovine serum albumin (BSA) or ovomucoid (OVM) as chiral discriminator is described. The phase system was prepared with dextran 40, polyethylene glycol and a protein. Ofloxacin (OFLX) enantiomer was well recognized by the BSA system, whereas the enantiomers of carvedilol (-blocker) and DG-5128 (a new oral hypoglycaemic drug) were hardly recognized by the OVM system. The distribution coefficients of (S)-, (R)- and rac-OFLX varied with pH. Good enantioselectivity for OFLX was obtained at relatively high pH ( pH 9). The crude optically enriched OFLX enantiomers were extracted from the aqueous fractions and finally purified by thin-layer chromatography.     

Selective separation of protein and saccharides by ionic liquids aqueous two-phase systems

In the present work, it was found that aqueous solution of a hydrophilic ionic liquid (IL), 1-butyl-3-methylimidazolium dicyanamide ([C₄mim][N(CN)₂]), could be separated into an aqueous two-phase system (ATPS) by inorganic salts such as K₂HPO₄ and K₃PO₄. The top phase is IL-rich, while the bottom phase is phosphate-rich. It was shown that 82.7%–100% bovine serum albumin (BSA) could be enriched into the top phase and almost quantitative saccharides (arabinose, glucose, sucrose, raffinose or dextran) were preferentially extracted into the bottom phase in a single-step extraction by [C₄mim][N(CN)₂] + K₂HPO₄ ATPS. The extraction efficiency of BSA from the aqueous saccharide solutions was influenced by the molecular structure of saccharides. The conductivity, dynamic light scattering (DLS) and transmission electron microscopy (TEM) were combined to investigate the microstructure of the IL-rich top phase and the possible mechanism for the selective separation. It is suggested that the formation of the IL aggregate and the IL aggregate-BSA complex plays a significant role in the separation of BSA from aqueous saccharide solutions. This is the first example for the selective separation by ILs-based ATPSs. It is expected that these findings would have potential applications in bio-analysis, separation, and IL recycle.     

Enantioselective extraction of fenvaleric acid enantiomers by two-phase (W/O) recognition chiral extraction

To establish an extraction method for fenvaleric acid (FA) enantiomers using l-iso-butyl-l-tartaric esters and hydroxypropyl-β-cyclodextrin (HP-β-CD) as chiral selector, the distribution of FA enantiomers was examined in methanol aqueous solution containing HP-β-CD and 1,2-dichloroethane organic solution containing l-iso-butyl-l-tartaric esters. The influences of the concentration of l-iso-butyl-l-tartaric esters and HP-β-CD, organic diluent, pH, extraction temperature and the concentration of methanol aqueous solution on the partition coefficient (k) and separation factor (α) of FA were investigated. The experiment results showed that the complex formed by l-iso-butyl-l-tartaric esters with S-enantiomer is stabler than that with R-enantiomer. With the increase of the concentration of l-iso-butyl-l-tartaric ester, k and α increased; With the increase of the concentration of HP-β-CD, k increased firstly, and then decreased, but α increased all the while, k was the highest when the concentration of HP-β-CD was 4 mmol L^?1. 1,2-dichloroethane organic diluent was better than the others. With the increase of pH, k and α decreased; with further increasing concentration of methanol aqueous solution, k and α decreased, k and α were the highest when the concentration of methanol aqueous solution was 10%. The extraction temperature had a great influence on k and α, too.     

In situ extraction of intracellular L-asparaginase using thermoseparating aqueous two-phase systems

The feasibility and generic applicability of directly integrating conventional discrete operations of cell disruption by high pressure homogenizer and the product capture by aqueous two-phase extraction (ATPE) system have been demonstrated for the extraction of intracellular L-asparaginase from E. coli. In a side-by-side comparison with the conventional ATPE process, including cell disruption, centrifugal clarification and following ATPE, purification of L-asparaginase via this novel in situ ATPE process yielded a product of L-asparaginase with a higher specific activity of 94.8 U/(mg protein) and a higher yield of 73.3%, both of which in the conventional ATPE process were 78.6 U/(mg protein) and 52.1%, respectively. In the purification of L-asparaginase (pI=4.9), product-debris interactions commonly diminish its recovery. It was demonstrated that immediate extraction of L-asparaginase in ATPE systems when it is released at pH 5.0 during cell disruption effectively increased its recovery in the top phase due to the reduced interaction between L-asparaginase and cell debris and the reduced degradation by contaminated protease. In addition, no clarification step and/or disruptate storage are required in this in situ ATPE, which reduced the number of unit operations and thus shortened the overall process time. This novel process has a good potential for the separation of other intracellular biological products.     

Effect of anionic structure on the phase formation and hydrophobicity of amino acid ionic liquids aqueous two-phase systems

Compared with the conventional ionic liquids, amino acid ionic liquids are more biodegradable and biocompatible, and can enhance stability of biomaterials. In this work, amino acid ionic liquids 1-butyl-3-methylimidazolium L-serine ([C(4)mim][Ser]), 1-butyl-3-methylimidazolium glycine ([C(4)mim][Gly]), 1-butyl-3-methylimidazolium L-alanine ([C(4)mim][Ala]) and 1-butyl-3-methylimidazolium L-leucine ([C(4)mim][Leu]) have been synthesized. These ionic liquids are found to form aqueous two-phase systems (ATPSs) by the salted-out of K(3)PO(4) in aqueous solutions. Phase diagram of the ATPSs and the Gibbs energies of transfer of methylene group from the bottom salt-rich phase to the top ionic liquid-rich phase have been determined at 298.15K and pH 14, and the effect of anionic structure of the ionic liquids on phase formation of the ATPSs and the relative hydrophobicity between the top and the bottom phases are then examined. In order to understand the effect of relative hydrophobicity of the phases in equilibrium in the ATPSs on the extraction/separation capability of biomolecules, the partition coefficients of cytochrome-c (as a model biomolecule) in the ATPSs are measured by spectrophotometry. It is suggested that hydrophobic interactions are mainly responsible for the higher partition coefficients of cytochrome-c in aqueous two-phase systems at pH 14, and the extraction and separation capacity of biomolecules can be improved by the modulation of the relative hydrophobicity of the phases and/or the pH of the system.     

Study of Cs extraction in an ionic liquid system using a common anion in both aqueous and ionic liquid phases

The effect of Tf₂N⁻, as a common anion in aqueous and ionic liquid (IL) phases, on Cs extraction in the IL system was investigated using C₂mimTf₂N as an IL and DCH18C6 as an extractant. The “common anion (Tf₂N⁻) effect” operated via the movement of Cs⁺·Tf₂N⁻ from the aqueous phase into the IL phase by extraction in the form of Cs⁺·extractant·Tf₂N⁻, without any traditional cation exchange. The extraction product was recovered as a precipitate using a small amount of IL, which facilitated precipitation via supersaturation.

     

Factors affecting the separation of mandelic acid enantiomers by capillary electrophoresis

Chromatographia - The separation of the optical isomers of mandelic acid by capillary electrophoresis using γ-cyclodextrin as a chiral selector is presented. The mobility of the enantiomers...     

Drop formation in aqueous two-phase systems

Extraction using aqueous two-phase systems (ATPSs) is a versatile technique for the downstream processing of various proteins/enzymes. The study of drop formation deals with the fundamental understanding of the behavior of liquid drops under the influence of various external body as well as surface forces. These studies provide a basis for designing of the extractions in column contactors in which liquid drops play a major role. Most of the drop formation studies reported so far is restricted to aqueous-organic systems. ATPSs, differ from aqueous-organic systems in their physical properties. In view of this, an attempt was made to develop a model for drop formation in ATPSs adopting the information available on aqueous-organic systems. In order to validate the model, experiments were performed by using polyethylene glycol (PEG)/salt systems of different phase compositions at various flow rates. At low flow rates the single stage model and at high flow rates the two stage model are able to predict the drop volume during its formation from tip of capillary. The experimental results were found to agree reasonably well with those predicted by the model.     

Microfluidics with aqueous two-phase systems

An overview is given about research activities in which aqueous two phase systems (ATPSs) are utilized in microfluidic setups. ATPSs consist of two immiscible aqueous phases and have traditionally been used for the separation and purification of biological material such as proteins or cells. Microfluidic implementations of such schemes are usually based on a number of co-flowing streams of immiscible phases in a microchannel, thereby replacing the standard batch by flow-through processes. Some aspects of the stability of such flow patterns and the recovery of the phases at the channel exit are reviewed. Furthermore, the diffusive mass transfer and sample partitioning between the phases are discussed, and corresponding applications are highlighted. When diffusion is superposed by an applied electric field normal to the liquid/liquid interface, the transport processes are accelerated, and under specific conditions the interface acts as a size-selective filter for molecules. Finally, the activities involving droplet microflows of ATPSs are reviewed. By either forming ATPS droplets in an organic phase or a droplet of one aqueous phase inside the other, a range of applications has been demonstrated, extending from separation/purification schemes to the patterning of surfaces covered with cells.     

聚乙二醇-硫酸铵双水相萃取次甲基蓝的研究

利用分光光度法研究了次甲基蓝在聚乙二醇-硫酸铵双水相体系的萃取行为,探索了质量配比(mPEc/m(NH4)2SO4)、温度和次甲基蓝的浓度对双水相及次甲基蓝萃取率(Y)的影响.实验表明:(1)在一定的温度和浓度下,随着mPEC/m(NH4)2SO4的减小,分配系数K( cup/clow)增加,萃取率Y略有减小;(2)在一定的质量配比和浓度下,分配系数K(cup/clow)随温度的升高而增大,但萃取率Y却基本保持不变;(3)在一定的质量配比和温度下,随着次甲基蓝浓度的增加,分配系数K(cup/clow)增大;萃取率Y略有增加.在mPEG/m( NH4) 2SO4为1.50∶1,c次甲基蓝=8.915×10-5mol/L、t=30℃的实验条件下,K(cup/clow)=22.14,萃取率(y)可达95.43％.