Specific recognition of protein by deep eutectic solvent-based magnetic ß-cyclodextrin molecularly imprinted polymer.

A magnetic ß-cyclodextrin (MCD) surface molecularly imprinted polymer (MIP) based on deep eutectic solvents (DESs) as cross-linker and functional monomer (MCD@DES-MIP) was successfully synthesized for the specific recognition of bovine hemoglobin (BHb). The adsorption behavior of MCD@DES-MIP for BHb was investigated by adsorption thermodynamics, adsorption kinetics, and pH control experiments. The maximum adsorption capacity of MCD@DES-MIP for BHb under the optimized conditions was 195.94 mg g-1 and the imprinting factor was 4.68. In addition, the competitive adsorption experiments demonstrated that MCD@DES-MIP showed excellent selective extraction ability for BHb in the binary mixture of BHb and bovine serum albumin (BSA). The actual sample analysis manifested that MCD@DES-MIP effectively separated BHb from complex samples. The results of circular dichroism spectra proved that the secondary structure of BHb did not change during elution. The result indicated that MCD@DES-MIP can be used as a new imprinting material for the separation and purification of BHb.Graphical abstract Magnetic imprinted microspheres (MCD@DES-MIP) were prepared by free radical polymerization using magnetic ß-cyclodextrin (MCD) as carrier, deep eutectic solvents (DESs) as functional monomer and cross-linker. MCD@DES-MIP show high adsorption capacity and excellent selectivity for BHb.

Tubulin colchicine site binding agent LL01 displays potent antitumor efficiency both in vitro and in vivo with suitable drug-like properties.

Through rational drug design, we previously identified an indenoprazole derivative, 2-(6-ethoxy-3-(3-ethoxyphenylamino)-1-methyl-1,4-dihydroindeno[1,2-c]pyrazol-7-yloxy)acetamide (LL01), as a potent tubulin polymerization inhibitor targeting the tubulin colchicine binding site. In this study, we further demonstrated that LL01 was not a P-gp substrate. It potently inhibited the growth of a variety of tumor cells, including those with multidrug resistance, with GI50 values in the low nanomole ranges. In vitro liver microsome stability assay, LL01 was modest stable in the liver microsomes of human, mouse and rat, but was fast metabolized in dog. After single oral administration of LL01 at a dose of 10 mg/kg in SD male rats, LL01 showed acceptable PK properties with a mean bioavailability of 41%. In human HepG2 hepatoma xenograft, at the oral doses of 25 mg/kg/day and 12.5 mg/kg/day, LL01 inhibited the tumor growth by 61.27%, and 43.74%, respectively, which is much better than the positive drug sorafenib (29.45%; 30 mg/kg/day). Therefore, LL01 might be a potential drug candidate for further investigation for hepatocellular carcinoma therapy.

A composite consisting of a deep eutectic solvent and dispersed magnetic metal-organic framework (type UiO-66-NH2) for solid-phase extraction of RNA.

A cactus-shaped magnetic composite was prepared for solid-phase extraction of RNA. It is composed of the metal organic framework UiO-66-NH2 that was modified with Fe3O4 nanoparticles. The composite was then dispersed in a lactic acid-based deep eutectic solvent (DES, Fe3O4-COOH@UiO-66-NH2@DES). The structures of the sorbents were characterized by transmission electron microscopy, scanning electron microscopy, X-ray diffraction, Fourier transform infrared spectrometry, vibrating sample magnetometry and thermogravimetric analysis. The extraction performance of sorbents was optimized and the maximum extraction capacity reached 246 mg·g-1. Extraction is shown to mainly rely on chelation interaction, electrostatic interaction, hydrophobic interaction and hydrogen bonding interaction. The sorbent can selectively extract RNA over DNA, bovine hemoglobin and amino acids. Regeneration studies indicated that the sorbent can be re-used (after regenreation with DES) several times without obvious change of the extraction capacity. The successful extraction of RNA from yeast testified the practical application of the sorbent. Graphical abstractSchematic representation of the fabrication Fe3O4-COOH@UiO-66-NH2@DES, and its application in the magnetic solid phase extraction of RNA.

Preparation of magnetic molecularly imprinted polymers based on a deep eutectic solvent as the functional monomer for specific recognition of lysozyme.

A magnetized molecularly imprinted polymer (MIP) was prepared via a surface-imprinting technique. An allyl-based deep eutectic solvent was chosen as the functional monomer to obtain the polymer for specific recognition of lysozyme. It was deposited on silica-coated magnetite nanoparticles. The structure of the polymer was confirmed by X-ray diffraction, Fourier transform infrared spectrometry, transmission electron microscopy, thermogravimetric analysis and vibrating sample magnetometry. The maximum binding capacity of the imprinted polymer is found to be 108 mg·g-1, which is higher than that of non-imprinted polymer. Compared to reference proteins such as cytochrome C, bovine hemoglobin and bovine serum albumin, the MIP shows favorable selectivity for lysozyme. Besides, the imprinted polymer can be further used to specifically recognize lysozyme from the protein mixture and chicken egg white. Reusability studies demonstrate that the polymer can be recycled four times without significant loss of adsorption capacity. The LOD of the method is 12.8 µg·mL-1. The relative standard deviations (for n = 3) are 1.38% for precision and 2.76% for repeatability. Its facile synthesis, high adsorption performance and excellent selectivity to capture lysozyme make this polymer an attractive candidate to be applied in biomacromolecular purification. Graphical abstract Magnetic molecularly imprinted polymer (MIP) based on deep eutectic solvent as functional monomer was fabricated and applied for the specific recognition of lysozyme. The MIP exhibits high adsorption capacity and excellent selectivity for lysozyme.

Analysis of the Volatile Profile of Core Chinese Mango Germplasm by Headspace Solid-Phase Microextraction Coupled with Gas Chromatography-Mass Spectrometry.

Despite abundant published research on the volatile characterization of mango germplasm, the aroma differentiation of Chinese cultivars remains unclear. Using headspace solid phase microextraction (HS-SPME) coupled with gas chromatography⁻mass spectrometry (GC-MS), the composition and relative content of volatiles in 37 cultivars representing the diversity of Chinese mango germplasm were investigated. Results indicated that there are distinct differences in the components and content of volatile compounds among and within cultivars. In total, 114 volatile compounds, including 23 monoterpenes, 16 sesquiterpenes, 29 non-terpene hydrocarbons, 25 esters, 11 aldehydes, five alcohols and five ketones, were identified. The total volatile content among cultivars ranged from 211 to 26,022 µg/kg fresh weight (FW), with 123-fold variation. Terpene compounds were the basic background volatiles, and 34 cultivars exhibited abundant monoterpenes. On the basis of hierarchical cluster analysis (HCA) and principal component analysis (PCA), terpinolene and α-pinene were important components constituting the aroma of Chinese mango cultivars. Most obviously, a number of mango cultivars with high content of various aroma components were observed, and they can serve as potential germplasms for both breeding and direct use.

Expression of Nodal on Bronchial Epithelial Cells Influenced by Lung Microbes Through DNA Methylation Modulates the Differentiation of T-Helper Cells.

BACKGROUND/AIMS: The previous study in our lab showed that Nodal molecule on bronchial epithelial cells (BECs) was modulated by all kinds of lung microbes. The present study was designed to determine the effects of Nodal on proliferation of BECs and BECs-induced differentiation of T-helper (Th) cells. The epigenetic mechanisms of Nodal expression following treatments of different lung microbes were also identified. METHODS: Real-time polymerization chain reaction (PCR) and western blot were used to determine the expression of Nodal. Flow cytometry was used to observe the effects of proliferation of BECs and subsequent BECs-induced differentiation of Th cells. Methylation levels of CpG islands in Nodal promoters were also analyzed by time of flight mass spectrometry. RESULTS: The results showed that Nodal promoted proliferation of BECs and BECs-induced differentiation of Th cell from Th1 to Th2 and Th17. Nodal promoter showed a hyper-methylation in normal BECs. Through methylation modification in the promoter, P. aeruginosa or A.baumanni inhibited the expression of Nodal while RSV promoted the expression of Nodal. CONCLUSIONS: Our data showed that Nodal promoted Th2 and Th17 differentiation and inhibited Th1 differentiation which may cause imbalance of airway microenvironment. P. aeruginosa or A.baumanni may be hopeful for the treatment of airway hyperresponsveness by inhibition Nodal expression through DNA methylation modification in the promoter.

Magnetic multiwall carbon nanotubes modified with dual hydroxy functional ionic liquid for the solid-phase extraction of protein.

A novel adsorbent based on silica-coated magnetic multiwall carbon nanotubes (MWCNTs) surface modified by dual hydroxy functional ionic liquid (FIL) ([OH]-FIL-m-MWCNTs@SiO2) has been designed and used for the purification of lysozyme (Lys) by magnetic solid-phase extraction (MSPE). Fourier transform infrared spectroscopy (FTIR), a vibrating sample magnetometer (VSM), transmission electron microscopy (TEM), X-ray diffraction (XRD) and thermogravimetric analysis (TGA) were employed to characterize [OH]-FIL-m-MWCNTs@SiO2. After extraction, the concentration of Lys was determined by a UV-Vis spectrophotometer at 278 nm. A series of single-factor experiments were carried out to identify the optimal conditions of the extraction and the extraction amount could reach up to 94.6 mg g(-1). The RSD of the precision, the repeatability and the stability experiments were 0.37% (n = 3), 0.47% (n = 3) and 0.52% (n = 3), respectively. Comparison of [OH]-FIL-m-MWCNTs@SiO2 with silica-coated magnetic Fe3O4 (Fe3O4@SiO2), silica-coated magnetic multiwall carbon nanotubes (m-MWCNTs@SiO2) and alkyl quaternary ammonium ionic liquid-modified on m-MWCNTs@SiO2 was carried out by extracting Lys. The extraction of bovine serum albumin (BSA), trypsin (Try) and ovalbumin (OVA) was also done by the proposed method. Desorption of Lys was carried out by 0.005 mol L(-1) Na2HPO4-1 mol L(-1) NaCl as the eluent solution and the desorption ratio reached 91.6%. Nearly 97.8% of the [OH]-FIL-m-MWCNTs@SiO2 could be recovered from each run, and the extraction amount decreased less after five runs. The circular dichroism spectral experiment analysis indicated that the secondary structure of Lys was unchanged after extraction.

A green deep eutectic solvent modified magnetic titanium dioxide nanoparticles for the solid-phase extraction of chymotrypsin.

Magnetic titanium dioxide nanoparticles modified with green deep eutectic solvent (DES) composed of choline chloride (ChCl) and xylitol (Xyl) (Fe3O4@TiO2@[ChCl][Xyl]) were synthesized and applied to the solid-phase extraction(MSPE) of chymotrypsin (Chy). The physicochemical properties and morphology of Fe3O4@TiO2@[ChCl][Xyl] was characterized by Fourier transform infrared spectroscopy (FT-IR), thermogravimetric analysis (TGA), Zeta potential, X-ray diffraction (XRD), vibrating sample magnetometer (VSM) and transmission electron microscope (TEM). The experiment parameters such as initial concentration of Chy, extraction time, pH value, ionic strength, extraction temperature and sample matrix were effectively optimized. Under the optimal experimental conditions, the extraction capacity of Fe3O4@TiO2@[ChCl][Xyl] obtained a significantly improvement after the modification of Fe3O4@TiO2 nanoparticles by [ChCl][Xyl], and reached up to 347.8 mg g-1. In the elution experiment, 10% sodium dodecyl sulfate-acetic acid (SDS-HAc) was used as eluent, achieving an elution rate of 85.9% for the Chy on Fe3O4@TiO2@[ChCl][Xyl]. And the Fe3O4@TiO2@[ChCl][Xyl] still maintained a good extraction capacity for Chy after six times of reuse. The application result in the extraction of Chy from porcine pancreas crude extract showed a good practical application ability for Chy extraction. All the results indicated that the synthesized Fe3O4@TiO2@[ChCl][Xyl] has good application potential in the extraction of biomolecular molecules such as protein.

A deep eutectic solvent modified magnetic ß-cyclodextrin particle for solid-phase extraction of trypsin.

An environmentally friendly deep eutectic solvent (DES) composed of trifluoroacetamide and benzyltrimethylammonium chloride modified ß-cyclodextrin (CD) grafted magnetic beads (MB-NH2@CD@DES) were synthesized for the first time and applied to the solid-phase extraction of trypsin. Among the five trypsin extractants prepared in this work (MB, MB-NH2, MB-NH2@CD, MB-NH2@DES, MB-NH2@CD@DES), the extractant MB-NH2@CD@DES with higher extraction capacity for trypsin was selected as final extractant. The extraction capacity of MB-NH2@CD@DES for trypsin can reach up to 549.87 mg⋅g-1 under the optimized conditions. The Langmuir adsorption equilibrium was found fitted better with equilibrium relation between MB-NH2@CD@DES and trypsin than Freundlich adsorption equilibrium. And a superior extraction for trypsin was verified by comparing the extraction capacity of MB-NH2@CD@DES for trypsin and four other common proteins. Compared with some reported trypsin extractants, the MB-NH2@CD@DES had a shorter extraction process, higher extraction capacity, more convenient operation of separation, a safer and more environmentally friendly synthesis process. With the optimized eluent, a great elution rate (74.32%) of trypsin was achieved. The absolute recovery of trypsin in trypsin standard solution was calculated to be 16.8%. And the extraction capacity of MB-NH2@CD@DES toward trypsin still maintained well after ten times recycling and reuse. The detection limit (LOD) and quantitative limit (LOQ) were 0.072 mg⋅mL-1 and 0.240 mg⋅mL-1 respectively. By sodium dodecyl sulfate polyacrylamide gel electrophoresis experiment, the extraction ability of MB-NH2@CD@DES to trypsin from real sample was fully demonstrated. All above results showed the potential of fabricated MB-NH2@CD@DES as a superior extractant for trypsin from real complex samples.

A new magnetic molecularly imprinted polymer based on deep eutectic solvents as functional monomer and cross-linker for specific recognition of bovine hemoglobin.

A novel magnetic molecularly imprinted polymer only based on deep eutectic solvents (DESs-MMIP) was successfully synthesized. The DESs-MMIP was constructed by using 2-hydroxyethyl methacrylate/tetrabutylammonium chloride deep eutectic solvent (DES1) as functional monomer, arylamide/(3-acrylamidopropyl) trimethylammonium chloride deep eutectic solvent (DES2) as cross-linker and bovine hemoglobin (BHb) as template through surface imprinting technology. The obtained DESs-MMIP was characterized by transmission electron microscope, X-ray diffraction, fourier transform infrared spectrometry, thermal gravimetric analysis and vibrating sample magnetometer. Under the optimized conditions, the maximum adsorption capacity of DESs-MMIP on BHb was 229.54 mg g-1 and the imprinting factor reached up to 21.89. The selective adsorption experiments indicated that compared with seven references, DESs-MMIP showed significant selectivity for BHb. The new-type DESs-MMIP exhibited higher adsorption capacity and imprinting factor on BHb than molecularly imprinted polymers constructed with traditional functional monomer and cross-linker in reported methods. The recognition of BHb by DESs-MMIP in calf blood samples demonstrated the practicality of the particles. The DESs-MMIP only based on deep eutectic solvents with excellent selectivity is expected to become an ideal candidate for selective recognition of BHb in complicated samples.

Magnetic carbon nanotube modified with polymeric deep eutectic solvent for the solid phase extraction of bovine serum albumin.

This article described the fabrication of novel magnetic carbon nanotube modified with polymeric deep eutectic solvent (M-CNT@PDES) and its application as extractant for the magnetic solid phase extraction (MSPE) of bovine serum albumin (BSA). The physicochemical properties and morphology of M-CNT@PDES were characterized by X-ray diffraction (XRD), vibrating sample magnetometer (VSM), thermo-gravimetric analysis (TGA), zeta potentials, fourier transform infrared spectrometry (FT-IR) and transmission electron microscope (TEM). Afterwards, several parameters such as pH value, initial concentration of BSA, extraction time, ionic strength and extraction temperature were optimized. The results indicated that the modification of PDES significantly improved the extraction performance for BSA, and the maximum extraction capacity was 225.15 mg/g under the optimized conditions. In addition, 0.20 mol/L NaCl-PBS solution was chosen as the appropriate eluent, and favourable elution rate (81.22%) was obtained. Circular dichroism spectroscopy (CD) indicated that the secondary structure of BSA has not changed during extraction and elution. The regenerative experiment and application in real calf serum confirmed the outstanding durability and practical application ability of M-CNT@PDES. All of above verified that the proposed M-CNT@PDES coupled with MSPE method has great application potential for the pre-concentration of biomolecules.

Ionic liquids skeleton typed magnetic core-shell molecularly imprinted polymers for the specific recognition of lysozyme.

The novel ionic liquids skeleton typed magnetic core-shell molecularly imprinted polymers (Fe3O4-COOH@IL-MIP) were firstly constructed with 1-vinyl-3-aminoformylmethyl imidazolium chloride ionic liquid ([VAFMIM]Cl-IL) modified magnetic particles as the substrate materials, [VAFMIM]Cl-IL as functional monomer, 1,6-hexanediyl-3,3'-bis-1-vinylimidazolium dichloride ionic liquid as cross-linker and Lysozyme (Lys) as template protein via surface-imprinting technique. The structure of Fe3O4-COOH@IL-MIP were confirmed by transmission and scanning electron microscopy, dynamic light scattering, thermo-gravimetric analysis, fourier transform infrared spectrometry and X-ray diffraction. The adsorption mechanism was discussed from the perspective of amino acid residues of Lys. The maximum adsorption capacity of MIPs was 166.36 mg g-1 and imprinting factor was 2.67. The competitive adsorption experiments demonstrated the favorable recognition ability of MIPs toward Lys. Reusability studies indicated MIPs can be reused ten times without obvious loss of rebinding ability. The Lys conformation maintained intact after elution and the elution rate was as high as 74%. The adsorption experiment of egg white manifested that MIPs can effectively separate Lys in practical samples. Only ILs and Fe3O4 were utilized to fabricate MIPs, this strategy realized the goal of energy and cost saving while achieving simple synthesis of imprinted materials, and is expected to provide a new feasible idea to exploit synthetic methods for protein-MIPs.

Development of different deep eutectic solvent aqueous biphasic systems for the separation of proteins.

In this work, aqueous biphasic systems (ABSs) formed by different deep eutectic solvents (DESs) were prepared and applied to extract proteins. The five kinds of DESs comprised amino acids and polyols ([amino acids][polyols]). They were combined with another DES resulting from tetrabutylammonium chloride and polypropylene glycol 400 ([TBAC][PPG400]) to form ABSs. The phase-forming abilities of [TBAC][PPG400]/[amino acids][polyols] were compared with those of [TBAC][PPG400]/amino acids and [TBAC][PPG400]/polyols. The results exhibited that the biphasic formation ability of [amino acids][polyols] lies between those of amino acids and polyols when [TBAC][PPG400] acts as the other phase in ABSs. The systems comprising [TBAC][PPG400] and [l-proline][xylitol] ([Pro][Xyl]) were further investigated to optimize the extraction performance. It was found that 97.30% chymotrypsin tended to distribute into the [Pro][Xyl]-rich phase under optimum conditions. The practical application of the system was demonstrated by the extraction of chymotrypsin from porcine pancreas. Besides, UV-Vis spectrophotometry (UV-Vis), Fourier transform infrared spectroscopy (FT-IR), and circular dichroism (CD) spectroscopy proved that the conformation of proteins remained unchanged during the extraction process. The extraction mechanism of the formation of DES-protein aggregates was investigated via conductivity, dynamic light scattering (DLS), and transmission electron microscopy (TEM). The overall results suggest that the DES/DES-based ABSs have outstanding potential in the green extraction of proteins.

Ionic liquid modified molybdenum disulfide and reduced graphene oxide magnetic nanocomposite for the magnetic separation of dye from aqueous solution.

A novel ionic liquid (IL) modified magnetic nanocomposite (mag-MoS2-RGO), which was composed of molybdenum disulfide (MoS2) and reduced graphene oxide (RGO), was firstly synthesized and characterized (mag-MoS2-RGO-IL). Compared with mag-MoS2-RGO, the mag-MoS2-RGO-IL exhibited the higher adsorption capacity of 143.9 mg/g for methylene blue (MB) under optimum conditions. Experimental results were imitated by adsorption isotherms and kinetic models, revealing a monolayer adsorption on the homogeneous surface governed by chemisorption. The removal rate was maintained 78% after 5 cycles of regeneration, indicating its good reutilization. Methodological study suggested that the method possessed excellent repeatability, precision and stability. The recoveries from the real samples were in the range of 87.0-98.8% with relative standard deviation of ≤5.2%. Benefiting from composite structure and magnetic Fe3O4, the stable magnetic adsorbent can be quickly separated from aqueous solution. All of above proves that the prepared mag-MoS2-RGO-IL has great application potential in dye separation.

Poly(deep eutectic solvent)-functionalized magnetic metal-organic framework composites coupled with solid-phase extraction for the selective separation of cationic dyes.

Novel polymeric deep eutectic solvents (PDES) based on 3-acrylamidopropyl trimethylammonium chloride/D-sorbitol functionalized amino-magnetic (Fe3O4NH2) metal-organic framework (HKUST-1-MOF) composites (Fe3O4NH2@HKUST-1@PDES) were synthesized and characterized by field emission transmission electron microscope (FE-SEM), transmission electron microscope (TEM), X-ray diffraction (XRD), fourier transform infrared spectrometry (FT-IR), thermal gravimetric analysis (TGA), vibrating sample magnetometer (VSM) and zeta potentials. Then the composites were firstly utilized to selectively separate malachite green (MG) and crystal violet (CV) coupled with magnetic solid-phase extraction (MSPE). A response surface methodology (RSM) based on Latin hypercube sampling (LHS) was selected to analytically optimize the extraction parameters including initial concentration of dyes, extraction time, pH value and extraction temperature. The maximum extraction amount and optimal extraction conditions predicted by the RSM model matched well with the actual experimental results, and the extraction amount was 966.93 mg g-1 for MG and 788.90 mg g-1 for CV,respectively. The results indicated that the model possessed higher calculation accuracy through analyzing fewer sample points, thereby achieving theoretical prediction of extraction amount and conditions and being a prefect supplementary to actual experiments. The electrostatic interaction between the composites and cationic dyes played the main roll in the extraction process. The proposed extraction method exhibited lower limit of detection (98.19 ng mL-1 for MG and 23.97 ng mL-1 for CV) and preeminent precision (RSD ˂ 0.4%). Spiked recoveries of fish samples at three spiking levers ranged from 89.43% to 100.65% for MG and 95.29%-98.03% for CV. All results highlighted the excellent potential of Fe3O4NH2@HKUST-1@PDES-MSPE strategy in selective separation of cationic dyes in complex medium.

Fabrication of magnetic polymers based on deep eutectic solvent for separation of bovine hemoglobin via molecular imprinting technology.

In this work, molecularly imprinted polymers (MIPs) were prepared with vinyl-coated magnetic particles (Fe3O4@VTEO) as the support material, deep eutectic solvent (DES) based on vinyl as the functional monomer, respectively. N,N-methylenebisacrylamide (MBAAm) was used as the crosslinker on account of its abundant carbon-carbon double bonds. The MIPs were prepared with the addition of bovine hemoglobin (BHb) acted as the template. The MIPs particles can be collected quickly by a magnetic field. The composition and morphology of the MIPs particles were characterized by transmission electron microscopy (TEM), dynamic light scattering (DLS), fourier transform infrared spectrometry (FT-IR) and thermo-gravimetric analysis (TGA). X-ray diffraction (XRD) was used to illustrate the cubic inverse spinel structure of Fe3O4. Meanwhile, vibrating sample magnetometer (VSM) was applied to characterize the magnetism of the MIPs. Adsorption experiments were performed to attain the optimum adsorption conditions. Under the optimized conditions, the obtained maximum adsorption capacity (Q, mg·g-1) of the MIPs particles is found to be 164.20 mg g-1, and the imprinting factor (IF) is 4.93. Four reference molecules were used to test the selectivity of the MIPs particles, which indicates that the recognition sites can adsorb template molecules with selectivity. Furthermore, the prepared magnetic MIPs particles were applied to capture BHb from the real samples (calf blood) effectively.

Development of deep eutectic solvent-based aqueous biphasic system for the extraction of lysozyme.

Aqueous Biphasic Systems (ABSs) based on deep eutectic solvents (DESs) were determined and applied in the extraction of lysozyme from chicken egg white. Tetrabutylammonium bromide (TBAB) and benzyltributylammonium bromide were utilized as hydrogen-bond acceptors to synthesize six kinds of DESs with different carboxylic acids (such as glycolic acid, Gly). The phase-formation ability of these DESs was evaluated by combining several salts. The results revealed that the content of hydrophilic group and the alkyl side chain length of the carboxylic acids played a crucial role in phase separation process, and the introduce of the benzyl group for quaternary ammonium salt had an aptitude to promote two-phase splitting. Then the system comprising [TBAB][Gly] and Na2SO4 was used to appraise the effect of different experimental parameters on the extraction efficiency, including the amount of DES and salt, the temperature, the values of pH and the ionic strength. More than 98% of lysozyme was transferred into the DES-rich phase at the optimum condition. The activity of lysozyme after the process of extraction still retained 91.73% of initial activity, demonstrating high biocompatibility of the studied system. What's more, the proposed method was successfully utilized for the real sample analysis. Finally, UV-vis, FT-IR, circular dichroism spectra, dynamic light scattering and transmission electron microscopy were employed to investigate the extraction mechanism. All of these results verify the excellent feasibility of the proposed system in the analysis of biological samples.

Preparation of ionic liquid modified magnetic metal-organic frameworks composites for the solid-phase extraction of α-chymotrypsin.

A novel magnetic solid-phase extraction (MSPE) method based on 1-hexyl-3-methyl imidazolium chloride ionic liquid (IL) modified magnetic Fe3O4 nanoparticles, hydroxylated multiwall carbon nanotubes (MWCNTs-OH) and zeolitic imidazolate frameworks (ZIFs) nanocomposites (Fe3O4-MWCNTs-OH@ZIF-67@IL) were proposed and applied to extract α-chymotrypsin. The magnetic materials were synthesized successfully and characterized by X-ray diffraction (XRD), transmission electron microscope (TEM), thermal gravimetric analysis (TGA), fourier transform infrared spectrometry (FT-IR), vibrating sample magnetometer (VSM) and zeta potentials. Subsequently, the UV-vis spectrophotometer at about 280 nm was utilized to quantitatively analyze the α-chymotrypsin concentration in the supernatant. Furthermore, single factor experiments revealed that the extraction capacity was influenced by initial α-chymotrypsin concentration, ionic strength, extraction time, extraction temperature and pH value. The extraction capacity could reach up to about 635 mg g-1 under the optimized conditions, absolutely higher than that of extraction for Ovalbumin (OVA), Bovine serum albumin (BSA) and Bovine hemoglobin (BHb). In addition, the regeneration studies showed Fe3O4-MWCNTs-OH@ZIF-67@IL particles could be reused several times and kept a high extraction capacity. Besides, the study of enzymatic activity also indicated that the activity of the extracted α-chymotrypsin was well maintained 93% of initial activity. What's more, the proposed method was successfully applied to extract α-chymotrypsin in porcine pancreas crude extract with satisfactory results. All of above conclusions highlight the great potential of the proposed Fe3O4-MWCNTs-OH@ZIF-67@IL-MSPE method in the analysis of biomolecules.

Aqueous biphasic systems formed by deep eutectic solvent and new-type salts for the high-performance extraction of pigments.

Deep eutectic solvent (DES) composed of polypropylene glycol 400 (PPG 400) and tetrabutylammonium bromide (TBAB) was combined with a series of new-type salts such as quaternary ammonium salts, amino acid and polyols to form Aqueous Biphasic Systems (ABSs). Phase-forming ability of the salts was investigated firstly. The results showed that polyols had a relatively weak power to produce phases within studied scopes. And the shorter of carbon chain length of salts, the easier to obtain phase-splitting. Then partitioning of three pigments in PPG 400/betaine-based ABSs was addressed to investigate the effect of pigments' hydrophobicity on extraction efficiency. It was found that an increase in hydrophobicity contributed to the migration of pigments in the DES-rich phase. On the other hand, with a decline in phase-forming ability of salts, the extraction efficiency of the whole systems started to go down gradually. Based on the results, selective separation experiment was conducted successfully in the PPG 400/betaine-based systems, including more than 93.00% Sudan Ⅲ in the top phase and about 80.00% sunset yellow FCF/amaranth in the bottom phase. Additionally, ABSs constructed by DES/betaine for partitioning amaranth were further utilized to explore the performances of influence factors and back extraction. It can be concluded that after the optimization above 98.00% amaranth was transferred into the top phase. And 67.98% amaranth can be transferred into the bottom phase in back-extraction experiment. At last, dynamic light scattering (DLS) and transmission electron microscope (TEM) were applied to probe into extraction mechanism. The results demonstrated that hydrophobicity played an important role in the separation process of pigments. Through combining with new-type DES, this work was devoted to introducing plentiful salts as novel compositions of ABSs and providing an eco-friendly extraction way for partitioning pigments, which boosted development of ABSs in the monitoring food safety field.

A novel aqueous biphasic system formed by deep eutectic solvent and ionic liquid for DNA partitioning.

In this work, aqueous biphasic systems (ABSs) composed of ionic liquids (ILs) and deep eutectic solvents (DESs) were developed and utilized to efficiently extract DNA for the first time. Four kinds of ILs/DESs were mainly constituted by betaine/carboxylic acid and betaine/carbohydrates respectively. While another DES ([TBAB][PPG400]) was formed by polypropylene glycol 400 (PPG 400) and tetrabutylammonium bromide (TBAB). The phase-formation ability of the studied ABSs was evaluated by using [TBAB][PPG400]/(ILs/DESs) and [TBAB][PPG400]/inorganic salts. The results revealed that the phase forming ability of ABSs involved with the size of anion alky chain of ILs, the viscosity, the density and the hydrophilicity of DESs, ionic radius and ionic valence of inorganic salts. Then the system comprising [TBAB][PPG400]/IL was selected to ascertain the optimum extraction conditions for the extraction of DNA by the influence factor experiments. Meanwhile the maximum extraction efficiency could be attained 99.60%. Mixed sample experiments were implemented to separate DNA/cytochrome C (Cyt-c) and DNA/bovine hemoglobin (BHb), where the DNA mainly partitioned to IL-rich bottom phase. It turned out that the relationship between the isoelectric point of analytes and the pH of the system played an important role in the separation process. The result also showed that the studied system can be applied to selectively separate mixtures of nucleic acids and proteins in a single-step. Moreover, the developed system was successfully applied to the extraction of DNA from bovine whole blood with satisfactory result. Finally, the extraction mechanisms associated with the separation process were explored by FT-IR spectra, circular dichroism spectra (CD), dynamic light scattering (DLS), transmission electron microscopy (TEM). Overall, the novel systems have been proven to be a remarkable performance in the separation of DNA, which is expected to be widely used and provide further possibilities in separation fields.